A site-specific genetic modification for induction of pluripotency and subsequent isolation of derived lung alveolar epithelial type II cells.

PubMed

Yan, Qing; Quan, Yuan; Sun, Huanhuan; Peng, Xinmiao; Zou, Zhengyun; Alcorn, Joseph L; Wetsel, Rick A; Wang, Dachun

2014-02-01

Human induced pluripotent stem cells (hiPSCs) have great therapeutic potential in repairing defective lung alveoli. However, genetic abnormalities caused by vector integrations and low efficiency in generating hiPSCs, as well as difficulty in obtaining transplantable hiPSC-derived cell types are still major obstacles. Here we report a novel strategy using a single nonviral site-specific targeting vector with a combination of Tet-On inducible gene expression system, Cre/lox P switching gene expression system, and alveolar epithelial type II cell (ATIIC)-specific Neomycin(R) transgene expression system. With this strategy, a single copy of all of the required transgenes can be specifically knocked into a site immediately downstream of β-2-microglobulin (B2M) gene locus at a high frequency, without causing B2M dysfunction. Thus, the expression of reprogramming factors, Oct4, Sox2, cMyc, and Klf4, can be precisely regulated for efficient reprogramming of somatic cells into random integration-free or genetic mutation-free hiPSCs. The exogenous reprogramming factor transgenes can be subsequently removed after reprogramming by transient expression of Cre recombinase, and the resulting random integration-free and exogenous reprogramming factor-free hiPSCs can be selectively differentiated into a homogenous population of ATIICs. In addition, we show that these hiPSC-derived ATIICs exhibit ultrastructural characteristics and biological functions of normal ATIICs. When transplanted into bleomycin-challenged mice lungs, hiPSC-derived ATIICs efficiently remain and re-epithelialize injured alveoli to restore pulmonary function, preventing lung fibrosis and increasing survival without tumorigenic side effect. This strategy allows for the first time efficient generation of patient-specific ATIICs for possible future clinical applications. © 2013 AlphaMed Press.

A site-specific genetic modification for induction of pluripotency and subsequent isolation of derived lung alveolar epithelial type II cells

PubMed Central

Yan, Qing; Quan, Yuan; Sun, Huanhuan; Peng, Xinmiao; Zou, Zhengyun; Alcorn, Joseph L.; Wetsel, Rick A.; Wang, Dachun

2013-01-01

Human induced pluripotent stem cells (hiPSCs) have great therapeutic potential in repairing defective lung alveoli. However, genetic abnormalities caused by vector-integrations and low efficiency in generating hiPSCs, as well as difficulty in obtaining transplantable hiPSC-derived cell types, are still major obstacles. Here we report a novel strategy using a single non-viral site-specific-targeting vector with a combination of Tet-On inducible gene expression system, Cre/lox P switching gene expression system, and alveolar epithelial type II cell (ATIIC)-specific NeomycinR trangene expression system. With this strategy, a single copy of all of the required transgenes can be specifically knocked into a site immediately downstream of beta-2-microglobulin (B2M) gene locus at a high frequency, without causing B2M dysfunction. Thus, the expression of reprogramming factors, Oct4, Sox2, cMyc and Klf4, can be precisely regulated for efficient reprogramming of somatic cells into random-integration-free or genetic mutation-free hiPSCs. The exogenous reprogramming factor transgenes can be subsequently removed after reprogramming by transient expression of Cre recombinase, and the resulting random-integration-free and exogenous reprogramming-factor-free hiPSCs can be selectively differentiated into a homogenous population of ATIICs. In addition, we show that these hiPSC-derived ATIICs exhibit ultra-structural characteristics and biological functions of normal ATIICs. When transplanted into bleomycin-challenged mice lungs, hiPSC-derived ATIICs efficiently remain and re-epithelialize injured alveoli to restore pulmonary function, preventing lung fibrosis and increasing survival without tumorigenic side effect. This strategy allows for the first time efficient generation of patient-specific ATIICs for possible future clinical applications. PMID:24123810

Deriving site-specific clean-up criteria to protect ecological receptors (plants and soil invertebrates) exposed to metal or metalloid soil contaminants via the direct contact exposure pathway

PubMed Central

Checkai, Ron; Van Genderen, Eric; Sousa, José Paulo; Stephenson, Gladys; Smolders, Erik

2014-01-01

Soil contaminant concentration limits for the protection of terrestrial plants and soil invertebrates are commonly based on thresholds derived using data from laboratory ecotoxicity tests. A comprehensive assessment has been made for the derivation of ecological soil screening levels (Eco-SSL) in the United States; however, these limits are conservative because of their focus on high bioavailability scenarios. Here, we explain and evaluate approaches to soil limit derivation taken by 4 jurisdictions, 2 of which allow for correction of data for factors affecting bioavailability among soils, and between spiked and field-contaminated soils (Registration Evaluation Authorisation and Restriction of Chemicals [REACH] Regulation, European Union [EU], and the National Environment Protection Council [NEPC], Australia). Scientifically advanced features from these methods have been integrated into a newly developed method for deriving soil clean-up values (SCVs) within the context of site-specific baseline ecological risk assessment. Resulting site-specific SCVs that account for bioavailability may permit a greater residual concentration in soil when compared to generic screening limit concentrations (e.g., Eco-SSL), while still affording acceptable protection. Two choices for selecting the level of protection are compared (i.e., allowing higher effect levels per species, or allowing a higher percentile of species that are potentially unprotected). Implementation of this new method is presented for the jurisdiction of the United States, with a focus on metal and metalloid contaminants; however, the new method can be used in any jurisdiction. A case study for molybdate shows the large effect of bioavailability corrections and smaller effects of protection level choices when deriving SCVs. Integr Environ Assess Manag 2014;10:346–357. PMID:24470189

Deriving sediment Interstitial Water Remediation Goals ...

EPA Pesticide Factsheets

Background/Objectives. Passive sampling is becoming a frequently used measurement technique at Superfund sites with contaminated sediments. Passive sampling measures the concentrations of freely dissolved chemicals (Cfrees) in the sediment interstitial water. The freely dissolved chemical is a good surrogate for and a very practical means for estimating the concentrations of bioavailable chemical in the sediments. Building from this approach, a methodology is proposed to derive sediment Interstitial Water Remediation Goals (IWRGs) for the protection of benthic organisms from direct toxicity using Cfrees measured with passive sampling.Approach/Activities. In the early 2000s, EPA developed and released Equilibrium Partitioning Sediment Benchmarks (ESBs) for a series of chemicals. ESBs are intended to be chemical concentrations below which unacceptable toxicity to benthic organisms does not occur. The ESBs (expressed with the units of ug/g OC) were derived using the equations:ESB= K_OC×FCV where K_OC=0.00028+0.983K_OWThe KOC is the organic carbon normalized sediment-water chemical partition coefficient, FCV is the Final Chronic Value from EPA’s ambient water quality criteria for the protection of aquatic life, and KOW is the n-octanol/water partition coefficient for the chemical. At a specific site, the remedial goal (CS:ESB µg/kg-dw) in sediment are then derived using the site-specific fraction of organic carbon in the sediment (fOC:SS) at the site:C_

Influence of quasi-specific sites on kinetics of target DNA search by a sequence-specific DNA-binding protein.

PubMed

Kemme, Catherine A; Esadze, Alexandre; Iwahara, Junji

2015-11-10

Functions of transcription factors require formation of specific complexes at particular sites in cis-regulatory elements of genes. However, chromosomal DNA contains numerous sites that are similar to the target sequences recognized by transcription factors. The influence of such "quasi-specific" sites on functions of the transcription factors is not well understood at present by experimental means. In this work, using fluorescence methods, we have investigated the influence of quasi-specific DNA sites on the efficiency of target location by the zinc finger DNA-binding domain of the inducible transcription factor Egr-1, which recognizes a 9 bp sequence. By stopped-flow assays, we measured the kinetics of Egr-1's association with a target site on 143 bp DNA in the presence of various competitor DNAs, including nonspecific and quasi-specific sites. The presence of quasi-specific sites on competitor DNA significantly decelerated the target association by the Egr-1 protein. The impact of the quasi-specific sites depended strongly on their affinity, their concentration, and the degree of their binding to the protein. To quantitatively describe the kinetic impact of the quasi-specific sites, we derived an analytical form of the apparent kinetic rate constant for the target association and used it for fitting to the experimental data. Our kinetic data with calf thymus DNA as a competitor suggested that there are millions of high-affinity quasi-specific sites for Egr-1 among the 3 billion bp of genomic DNA. This study quantitatively demonstrates that naturally abundant quasi-specific sites on DNA can considerably impede the target search processes of sequence-specific DNA-binding proteins.

Inhibition of Super-Enhancer Activity in Autoinflammatory Site-Derived T Cells Reduces Disease-Associated Gene Expression.

PubMed

Peeters, Janneke G C; Vervoort, Stephin J; Tan, Sander C; Mijnheer, Gerdien; de Roock, Sytze; Vastert, Sebastiaan J; Nieuwenhuis, Edward E S; van Wijk, Femke; Prakken, Berent J; Creyghton, Menno P; Coffer, Paul J; Mokry, Michal; van Loosdregt, Jorg

2015-09-29

The underlying molecular mechanisms for many autoimmune diseases are poorly understood. Juvenile idiopathic arthritis (JIA) is an exceptionally well-suited model for studying autoimmune diseases due to its early onset and the possibility to analyze cells derived from the site of inflammation. Epigenetic profiling, utilizing primary JIA patient-derived cells, can contribute to the understanding of autoimmune diseases. With H3K27ac chromatin immunoprecipitation, we identified a disease-specific, inflammation-associated, typical enhancer and super-enhancer signature in JIA patient synovial-fluid-derived CD4(+) memory/effector T cells. RNA sequencing of autoinflammatory site-derived patient T cells revealed that BET inhibition, utilizing JQ1, inhibited immune-related super-enhancers and preferentially reduced disease-associated gene expression, including cytokine-related processes. Altogether, these results demonstrate the potential use of enhancer profiling to identify disease mediators and provide evidence for BET inhibition as a possible therapeutic approach for the treatment of autoimmune diseases. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

The phage integrase vector pIPI03 allows RecA-independent, site-specific labelling of Staphylococcus lugdunensis strains.

PubMed

Heilbronner, Simon; Monk, Ian R; Foster, Timothy J

2013-11-01

Staphylococcus lugdunensis is a coagulase negative staphylococcus that is a commensal of man and an opportunistic pathogen. A site-specific integrative plasmid for the use in S. lugdunensis was constructed and validated. The integrase gene ccrB of bacteriophage ϕSL01 together with its attachment site was cloned into the thermosensitive plasmid pIMAY. The resulting plasmid pIPI03 integrated RecA-independently, site-specifically and irreversibly into the S. lugdunensis chromosome. Two IPTG-inducible antibiotic resistance determinants were cloned into pIPI03 and the derivatives were used to construct strains suitable for competitive growth experiments in both in vitro and in vivo. Copyright © 2013 Elsevier Inc. All rights reserved.

Novel IgE Inhibitors for the Treatment of Food Allergies

DTIC Science & Technology

2016-10-01

antibodies using phage display 1-12 Completed Subtask 5: Functional studies with phage-derived Fabs 12-24 In progess Subtask 6: Production and...characterization of bifunctional antibodies using phage-derived Fab 12-36 Not yet started Subtask 7: Elicitation of site specific antibodies from boost

Photoaffinity labeling of protoporphyrinogen oxidase, the molecular target of diphenylether-type herbicides.

PubMed

Camadro, J M; Matringe, M; Thome, F; Brouillet, N; Mornet, R; Labbe, P

1995-05-01

Diphenylether-type herbicides are extremely potent inhibitors of protoporphyrinogen oxidase, a membrane-bound enzyme involved in the heme and chlorophyll biosynthesis pathways. Tritiated acifluorfen and a diazoketone derivative of tritiated acifluorfen were specifically bound to a single class of high-affinity binding sites on yeast mitochondrial membranes with apparent dissociation constants of 7 nM and 12.5 nM, respectively. The maximum density of specific binding sites, determined by Scatchard analysis, was 3 pmol.mg-1 protein. Protoporphyrinogen oxidase specific activity was estimated to be 2500 nmol protoporphyrinogen oxidized h-1.mol-1 enzyme. The diazoketone derivative of tritiated acifluorfen was used to specifically photolabel yeast protoporphyrinogen oxidase. The specifically labeled polypeptide in wild-type mitochondrial membranes had an apparent molecular mass of 55 kDa, identical to the molecular mass of the purified enzyme. This photolabeled polypeptide was not detected in a protoporphyrinogen-oxidase-deficient yeast strain, but the membranes contained an equivalent amount of inactive immunoreactive protoporphyrinogen oxidase protein.

Influence of Quasi-Specific Sites on Kinetics of Target DNA Search by a Sequence-Specific DNA-Binding Protein

PubMed Central

2015-01-01

Functions of transcription factors require formation of specific complexes at particular sites in cis-regulatory elements of genes. However, chromosomal DNA contains numerous sites that are similar to the target sequences recognized by transcription factors. The influence of such “quasi-specific” sites on functions of the transcription factors is not well understood at present by experimental means. In this work, using fluorescence methods, we have investigated the influence of quasi-specific DNA sites on the efficiency of target location by the zinc finger DNA-binding domain of the inducible transcription factor Egr-1, which recognizes a 9 bp sequence. By stopped-flow assays, we measured the kinetics of Egr-1’s association with a target site on 143 bp DNA in the presence of various competitor DNAs, including nonspecific and quasi-specific sites. The presence of quasi-specific sites on competitor DNA significantly decelerated the target association by the Egr-1 protein. The impact of the quasi-specific sites depended strongly on their affinity, their concentration, and the degree of their binding to the protein. To quantitatively describe the kinetic impact of the quasi-specific sites, we derived an analytical form of the apparent kinetic rate constant for the target association and used it for fitting to the experimental data. Our kinetic data with calf thymus DNA as a competitor suggested that there are millions of high-affinity quasi-specific sites for Egr-1 among the 3 billion bp of genomic DNA. This study quantitatively demonstrates that naturally abundant quasi-specific sites on DNA can considerably impede the target search processes of sequence-specific DNA-binding proteins. PMID:26502071

Engineering peptide ligase specificity by proteomic identification of ligation sites.

PubMed

Weeks, Amy M; Wells, James A

2018-01-01

Enzyme-catalyzed peptide ligation is a powerful tool for site-specific protein bioconjugation, but stringent enzyme-substrate specificity limits its utility. We developed an approach for comprehensively characterizing peptide ligase specificity for N termini using proteome-derived peptide libraries. We used this strategy to characterize the ligation efficiency for >25,000 enzyme-substrate pairs in the context of the engineered peptide ligase subtiligase and identified a family of 72 mutant subtiligases with activity toward N-terminal sequences that were previously recalcitrant to modification. We applied these mutants individually for site-specific bioconjugation of purified proteins, including antibodies, and in algorithmically selected combinations for sequencing of the cellular N terminome with reduced sequence bias. We also developed a web application to enable algorithmic selection of the most efficient subtiligase variant(s) for bioconjugation to user-defined sequences. Our methods provide a new toolbox of enzymes for site-specific protein modification and a general approach for rapidly defining and engineering peptide ligase specificity.

Preparation and Characterization of Fluorescent Derivatives of Lysozyme

NASA Technical Reports Server (NTRS)

Smith, Lori; Pusey, Marc

1998-01-01

Fluorescence is one of the most versatile and powerful tools for the study of macromolecules. However, its use in macromolecular crystal growth studies is hampered by the necessity of preparing fluorescent derivatives where the probe does not markedly affect the crystal packing. Alternatively, one can prepare derivatives of limited utility if it is known that they will not affect the specific goals of a given study. We have prepared a number of fluorescent derivatives of chicken egg white lysozyme, covalently attaching fluorescent probes to two different sites on the protein molecule. The first site is the side chain carboxyl group of ASP 101. Amine containing probes such as lucifer yellow, cascade blue, and 5- (2-aminoethyl) aminonapthalene-l-sulfonic acid (EDANS) have been attached using a carbodiimide coupling procedure. ASP 101 lies within the active site cleft, and it is believed that the probes are "buried" within that cleft. This is supported by the fact that all such derivatives have been found to crystallize, with the crystals being fluorescent. Tetragonal crystals of the lucifer yellow derivative have been found to diffract to at least 1.9 A resolution. X-ray diffraction data has been acquired and we are now working on the structure of this derivative. The second group of derivatives is to the N-terminal amine group. The derivatization reaction is performed by using a succinimidyl ester of the probe to be attached. Fluorescent probes such as pyrene acetic acid, 5-carboxyfluorescein, and Oregon green have been attached to this site. We have had little success in crystallizing these derivatives, probably because this site is part of the contact region between the 43 helix chains. However, these sites do not interfere with formation of the 43 helices and the derivatives are suitable for study of their formation in solution. The derivatives are being characterized by steady state and lifetime fluorescence methods, and the presentation will discuss these results.

Distinct p53 genomic binding patterns in normal and cancer-derived human cells

PubMed Central

McCorkle, Sean R; McCombie, WR; Dunn, John J

2011-01-01

Here, we report genome-wide analysis of the tumor suppressor p53 binding sites in normal human cells. 743 high-confidence ChIP-seq peaks representing putative genomic binding sites were identified in normal IMR90 fibroblasts using a reference chromatin sample. More than 40% were located within 2 kb of a transcription start site (TSS), a distribution similar to that documented for individually studied, functional p53 binding sites and, to date, not observed by previous p53 genome-wide studies. Nearly half of the high-confidence binding sites in the IMR90 cells reside in CpG islands in marked contrast to sites reported in cancer-derived cells. The distinct genomic features of the IMR90 binding sites do not reflect a distinct preference for specific sequences, since the de novo developed p53 motif based on our study is similar to those reported by genome-wide studies of cancer cells. More likely, the different chromatin landscape in normal, compared with cancer-derived cells, influences p53 binding via modulating availability of the sites. We compared the IMR90 ChIP-seq peaks to the recently published IMR90 methylome1 and demonstrated that they are enriched at hypomethylated DNA. Our study represents the first genome-wide, de novo mapping of p53 binding sites in normal human cells and reveals that p53 binding sites reside in distinct genomic landscapes in normal and cancer-derived human cells. PMID:22127205

Restoring and Enhancing Productivity of Degraded Tephra-Derived Soils

Treesearch

Chuck Bulmer; Jim Archuleta; Mike Curran

2007-01-01

Soil restoration (sometimes termed enhancement) is an important strategy for sustaining the productivity of managed forest landscapes. Tephra-derived soils have unique physical and chemical characteristics that affect their response to disturbance and restoration. A variety of factors reduce forest productivity on degraded soils. Site-specific information on soil...

CAGEd-oPOSSUM: motif enrichment analysis from CAGE-derived TSSs.

PubMed

Arenillas, David J; Forrest, Alistair R R; Kawaji, Hideya; Lassmann, Timo; Wasserman, Wyeth W; Mathelier, Anthony

2016-09-15

With the emergence of large-scale Cap Analysis of Gene Expression (CAGE) datasets from individual labs and the FANTOM consortium, one can now analyze the cis-regulatory regions associated with gene transcription at an unprecedented level of refinement. By coupling transcription factor binding site (TFBS) enrichment analysis with CAGE-derived genomic regions, CAGEd-oPOSSUM can identify TFs that act as key regulators of genes involved in specific mammalian cell and tissue types. The webtool allows for the analysis of CAGE-derived transcription start sites (TSSs) either provided by the user or selected from ∼1300 mammalian samples from the FANTOM5 project with pre-computed TFBS predicted with JASPAR TF binding profiles. The tool helps power insights into the regulation of genes through the study of the specific usage of TSSs within specific cell types and/or under specific conditions. The CAGEd-oPOSUM web tool is implemented in Perl, MySQL and Apache and is available at http://cagedop.cmmt.ubc.ca/CAGEd_oPOSSUM CONTACTS: anthony.mathelier@ncmm.uio.no or wyeth@cmmt.ubc.ca Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press.

CAGEd-oPOSSUM: motif enrichment analysis from CAGE-derived TSSs

PubMed Central

Arenillas, David J.; Forrest, Alistair R. R.; Kawaji, Hideya; Lassmann, Timo; Wasserman, Wyeth W.; Mathelier, Anthony

2016-01-01

With the emergence of large-scale Cap Analysis of Gene Expression (CAGE) datasets from individual labs and the FANTOM consortium, one can now analyze the cis-regulatory regions associated with gene transcription at an unprecedented level of refinement. By coupling transcription factor binding site (TFBS) enrichment analysis with CAGE-derived genomic regions, CAGEd-oPOSSUM can identify TFs that act as key regulators of genes involved in specific mammalian cell and tissue types. The webtool allows for the analysis of CAGE-derived transcription start sites (TSSs) either provided by the user or selected from ∼1300 mammalian samples from the FANTOM5 project with pre-computed TFBS predicted with JASPAR TF binding profiles. The tool helps power insights into the regulation of genes through the study of the specific usage of TSSs within specific cell types and/or under specific conditions. Availability and Implementation: The CAGEd-oPOSUM web tool is implemented in Perl, MySQL and Apache and is available at http://cagedop.cmmt.ubc.ca/CAGEd_oPOSSUM. Contacts: anthony.mathelier@ncmm.uio.no or wyeth@cmmt.ubc.ca Supplementary information: Supplementary data are available at Bioinformatics online. PMID:27334471

TALEN-mediated generation and genetic correction of disease-specific human induced pluripotent stem cells.

PubMed

Ramalingam, Sivaprakash; Annaluru, Narayana; Kandavelou, Karthikeyan; Chandrasegaran, Srinivasan

2014-01-01

Generation and precise genetic correction of patient-derived hiPSCs have great potential in regenerative medicine. Such targeted genetic manipulations can now be achieved using gene-editing nucleases. Here, we report generation of cystic fibrosis (CF) and Gaucher's disease (GD) hiPSCs respectively from CF (homozygous for CFTRΔF508 mutation) and Type II GD [homozygous for β-glucocerebrosidase (GBA) 1448T>C mutation] patient fibroblasts, using CCR5- specific TALENs. Site-specific addition of loxP-flanked Oct4/Sox2/Klf4/Lin28/Nanog/eGFP gene cassette at the endogenous CCR5 site of patient-derived disease-specific primary fibroblasts induced reprogramming, giving rise to both monoallele (heterozygous) and biallele CCR5-modified hiPSCs. Subsequent excision of the donor cassette was done by treating CCR5-modified CF and GD hiPSCs with Cre. We also demonstrate site-specific correction of sickle cell disease (SCD) mutations at the endogenous HBB locus of patient-specific hiPSCs [TNC1 line that is homozygous for mutated β- globin alleles (βS/βS)], using HBB-specific TALENs. SCD-corrected hiPSC lines showed gene conversion of the mutated βS to the wild-type βA in one of the HBB alleles, while the other allele remained a mutant phenotype. After excision of the loxP-flanked DNA cassette from the SCD-corrected hiPSC lines using Cre, we obtained secondary heterozygous βS/βA hiPSCs, which express the wild-type (βA) transcript to 30-40% level as compared to uncorrected (βS/βS) SCD hiPSCs when differentiated into erythroid cells. Furthermore, we also show that TALEN-mediated generation and genetic correction of disease-specific hiPSCs did not induce any off-target mutations at closely related sites.

Soil type-depending effect of paddy management: composition and distribution of soil organic matter

NASA Astrophysics Data System (ADS)

Urbanski, Livia; Kölbl, Angelika; Lehndorff, Eva; Houtermans, Miriam; Schad, Peter; Zhang, Gang-Lin; Rahayu Utami, Sri; Kögel-Knabner, Ingrid

2016-04-01

Paddy soil management is assumed to promote soil organic matter accumulation and specifically lignin caused by the resistance of the aromatic lignin structure against biodegradation under anaerobic conditions during inundation of paddy fields. The present study investigates the effect of paddy soil management on soil organic matter composition compared to agricultural soils which are not used for rice production (non-paddy soils). A variety of major soil types, were chosen in Indonesia (Java), including Alisol, Andosol and Vertisol sites (humid tropical climate of Java, Indonesia) and in China Alisol sites (humid subtropical climate, Nanjing). This soils are typically used for rice cultivation and represent a large range of soil properties to be expected in Asian paddy fields. All topsoils were analysed for their soil organic matter composition by solid-state 13C nuclear magnetic resonance spectroscopy and lignin-derived phenols by CuO oxidation method. The soil organic matter composition, revealed by solid-state 13C nuclear magnetic resonance, was similar for the above named different parent soil types (non-paddy soils) and was also not affected by the specific paddy soil management. The contribution of lignin-related carbon groups to total SOM was similar in the investigated paddy and non-paddy soils. A significant proportion of the total aromatic carbon in some paddy and non-paddy soils was attributed to the application of charcoal as a common management practise. The extraction of lignin-derived phenols revealed low VSC (vanillyl, syringyl, cinnamyl) values for all investigated soils, being typical for agricultural soils. An inherent accumulation of lignin-derived phenols due to paddy management was not found. Lignin-derived phenols seem to be soil type-dependent, shown by different VSC concentrations between the parent soil types. The specific paddy management only affects the lignin-derived phenols in Andosol-derived paddy soils which are characterized by significantly higher VSC values compared to their parent soil types. However, the higher organic carbon concentrations in Andosol and Alisol (China)-derived paddy soils compared to their parent soil types, could not be explained by an enrichment of lignin-derived phenols. It seems that site specific incorporation of crop residues and properties of the parent soil types are likely more important for organic carbon contents and soil organic matter composition than the effect of paddy management itself.

Fracture induced mobilization and incorporation of bone marrow-derived endothelial progenitor cells for bone healing.

PubMed

Matsumoto, Tomoyuki; Mifune, Yutaka; Kawamoto, Atsuhiko; Kuroda, Ryosuke; Shoji, Taro; Iwasaki, Hiroto; Suzuki, Takahiro; Oyamada, Akira; Horii, Miki; Yokoyama, Ayumi; Nishimura, Hiromi; Lee, Sang Yang; Miwa, Masahiko; Doita, Minoru; Kurosaka, Masahiro; Asahara, Takayuki

2008-04-01

We recently reported that systemic administration of peripheral blood (PB) CD34+ cells, an endothelial progenitor cell (EPC)-enriched population, contributed to fracture healing via vasculogenesis/angiogenesis. However, pathophysiological role of EPCs in fracture healing process has not been fully clarified. Therefore, we investigated the hypothesis whether mobilization and incorporation of bone marrow (BM)-derived EPCs may play a pivotal role in appropriate fracture healing. Serial examinations of Laser doppler perfusion imaging and histological capillary density revealed that neovascularization activity at the fracture site peaked at day 7 post-fracture, the early phase of endochondral ossifification. Fluorescence-activated cell sorting (FACS) analysis demonstrated that the frequency of BM cKit+Sca1+Lineage- (Lin-) cells and PB Sca1+Lin- cells, which are EPC-enriched fractions, significantly increased post-fracture. The Sca1+ EPC-derived vasuculogenesis at the fracture site was confirmed by double immunohistochemistry for CD31 and Sca1. BM transplantation from transgenic donors expressing LacZ transcriptionally regulated by endothelial cell-specific Tie-2 promoter into wild type also provided direct evidence that EPCs contributing to enhanced neovascularization at the fracture site were specifically derived from BM. Animal model of systemic administration of PB Sca1+Lin- Green Fluorescent Protein (GFP)+ cells further confirmed incorporation of the mobilized EPCs into the fracture site for fracture healing. These findings indicate that fracture may induce mobilization of EPCs from BM to PB and recruitment of the mobilized EPCs into fracture sites, thereby augment neovascularization during the process of bone healing. EPCs may play an essential role in fracture healing by promoting a favorable environment through neovascularization in damaged skeletal tissue. (c) 2008 Wiley-Liss, Inc.

Seven diverse human embryonic stem cell-derived chondrogenic clonal embryonic progenitor cell lines display site-specific cell fates.

PubMed

Sternberg, Hal; Kidd, Jennifer; Murai, James T; Jiang, Jianjie; Rinon, Ariel; Erickson, Isaac E; Funk, Walter D; Wang, Qian; Chapman, Karen B; Vangsness, C Thomas; West, Michael D

2013-03-01

The transcriptomes of seven diverse clonal human embryonic progenitor cell lines with chondrogenic potential were compared with that of bone marrow-derived mesenchymal stem cells (MSCs). The cell lines 4D20.8, 7PEND24, 7SMOO32, E15, MEL2, SK11 and SM30 were compared with MSCs using immunohistochemical methods, gene expression microarrays and quantitative real-time PCR. In the undifferentiated progenitor state, each line displayed unique combinations of site-specific markers, including AJAP1, ALDH1A2, BMP5, BARX1, HAND2, HOXB2, LHX1, LHX8, PITX1, TBX15 and ZIC2, but none of the lines expressed the MSC marker CD74. The lines showed diverse responses when differentiated in the presence of combinations of TGF-β3, BMP2, 4, 6 and 7 and GDF5, with the lines 4D20.8, SK11, SM30 and MEL2 showing osteogenic markers in some differentiation conditions. The line 7PEND24 showed evidence of regenerating articular cartilage and, in some conditions, markers of tendon differentiation. The scalability of site-specific clonal human embryonic stem cell-derived embryonic progenitor cell lines may provide novel models for the study of differentiation and methods for preparing purified and identified cells types for use in therapy.

In and out of the rRNA genes: characterization of Pokey elements in the sequenced Daphnia genome

PubMed Central

2013-01-01

Background Only a few transposable elements are known to exhibit site-specific insertion patterns, including the well-studied R-element retrotransposons that insert into specific sites within the multigene rDNA. The only known rDNA-specific DNA transposon, Pokey (superfamily: piggyBac) is found in the freshwater microcrustacean, Daphnia pulex. Here, we present a genome-wide analysis of Pokey based on the recently completed whole genome sequencing project for D. pulex. Results Phylogenetic analysis of Pokey elements recovered from the genome sequence revealed the presence of four lineages corresponding to two divergent autonomous families and two related lineages of non-autonomous miniature inverted repeat transposable elements (MITEs). The MITEs are also found at the same 28S rRNA gene insertion site as the Pokey elements, and appear to have arisen as deletion derivatives of autonomous elements. Several copies of the full-length Pokey elements may be capable of producing an active transposase. Surprisingly, both families of Pokey possess a series of 200 bp repeats upstream of the transposase that is derived from the rDNA intergenic spacer (IGS). The IGS sequences within the Pokey elements appear to be evolving in concert with the rDNA units. Finally, analysis of the insertion sites of Pokey elements outside of rDNA showed a target preference for sites similar to the specific sequence that is targeted within rDNA. Conclusions Based on the target site preference of Pokey elements and the concerted evolution of a segment of the element with the rDNA unit, we propose an evolutionary path by which the ancestors of Pokey elements have invaded the rDNA niche. We discuss how specificity for the rDNA unit may have evolved and how this specificity has played a role in the long-term survival of these elements in the subgenus Daphnia. PMID:24059783

Identification and removal of low-complexity sites in allele-specific analysis of ChIP-seq data.

PubMed

Waszak, Sebastian M; Kilpinen, Helena; Gschwind, Andreas R; Orioli, Andrea; Raghav, Sunil K; Witwicki, Robert M; Migliavacca, Eugenia; Yurovsky, Alisa; Lappalainen, Tuuli; Hernandez, Nouria; Reymond, Alexandre; Dermitzakis, Emmanouil T; Deplancke, Bart

2014-01-15

High-throughput sequencing technologies enable the genome-wide analysis of the impact of genetic variation on molecular phenotypes at unprecedented resolution. However, although powerful, these technologies can also introduce unexpected artifacts. We investigated the impact of library amplification bias on the identification of allele-specific (AS) molecular events from high-throughput sequencing data derived from chromatin immunoprecipitation assays (ChIP-seq). Putative AS DNA binding activity for RNA polymerase II was determined using ChIP-seq data derived from lymphoblastoid cell lines of two parent-daughter trios. We found that, at high-sequencing depth, many significant AS binding sites suffered from an amplification bias, as evidenced by a larger number of clonal reads representing one of the two alleles. To alleviate this bias, we devised an amplification bias detection strategy, which filters out sites with low read complexity and sites featuring a significant excess of clonal reads. This method will be useful for AS analyses involving ChIP-seq and other functional sequencing assays. The R package abs filter for library clonality simulations and detection of amplification-biased sites is available from http://updepla1srv1.epfl.ch/waszaks/absfilter

A novel site-specific recombination system derived from bacteriophage phiMR11.

PubMed

Rashel, Mohammad; Uchiyama, Jumpei; Ujihara, Takako; Takemura, Iyo; Hoshiba, Hiroshi; Matsuzaki, Shigenobu

2008-04-04

We report identification of a novel site-specific DNA recombination system that functions in both in vivo and in vitro, derived from lysogenic Staphylococcus aureus phage phiMR11. In silico analysis of the phiMR11 genome indicated orf1 as a putative integrase gene. Phage and bacterial attachment sites (attP and attB, respectively) and attachment junctions were determined and their nucleotide sequences decoded. Sequences of attP and attB were mostly different to each other except for a two bp common core that was the crossover point. We found several inverted repeats adjacent to the core sequence of attP as potential protein binding sites. The precise and efficient integration properties of phiMR11 integrase were shown on attP and attB in Escherichia coli and the minimum size of attP was found to be 34bp. In in vitro assays using crude or purified integrase, only buffer and substrate DNAs were required for the recombination reaction, indicating that other bacterially encoded factors are not essential for activity.

Antigenic characterization of a formalin-inactivated poliovirus vaccine derived from live-attenuated Sabin strains.

PubMed

Tano, Yoshio; Shimizu, Hiroyuki; Martin, Javier; Nishimura, Yorihiro; Simizu, Bunsiti; Miyamura, Tatsuo

2007-10-10

A candidate inactivated poliovirus vaccine derived from live-attenuated Sabin strains (sIPV), which are used in the oral poliovirus vaccine (OPV), was prepared in a large-production scale. The modification of viral antigenic epitopes during the formalin inactivation process was investigated by capture ELISA assays using type-specific and antigenic site-specific monoclonal antibodies (MoAbs). The major antigenic site 1 was modified during the formalin inactivation of Sabin 1. Antigenic sites 1-3 were slightly modified during the formalin inactivation of Sabin 2 strain. Sites 1 and 3 were altered on inactivated Sabin 3 virus. These alterations were different to those shown by wild-type Saukett strain, used in conventional IPV (cIPV). It has been previously reported that type 1 sIPV showed higher immunogenicity to type 1 cIPV whereas types 2 and 3 sIPV induced lower level of immunogenicity than their cIPV counterparts. Our results suggest that the differences in epitope structure after formalin inactivation may account, at least in part, for the observed differences in immunogenicity between Sabin and wild-type inactivated poliovaccines.

Genetic Incorporation of Twelve meta-Substituted Phenylalanine Derivatives Using A Single Pyrrolysyl-tRNA Synthetase

PubMed Central

Wang, Yane-Shih; Fang, Xinqiang; Chen, Hsueh-Ying; Wu, Bo; Wang, Zhiyong U.; Hilty, Christian; Liu, Wenshe R.

2012-01-01

When coexpressed with its cognate amber suppressing tRNACUAPyl, a pyrrolysyl-tRNA synthetase mutant N346A/C348A is able to genetically incorporate twelve meta-substituted phenylalanine derivatives into proteins site-specifically at amber mutation sites in Escherichia coli. These genetically encoded noncanonical amino acids resemble phenylalanine in size and contain diverse bioorthogonal functional groups such as halide, trifluoromethyl, nitrile, nitro, ketone, alkyne, and azide moieties. The genetic installation of these functional groups in proteins provides multiple ways to site-selectively label proteins with biophysical and biochemical probes for their functional investigations. We demonstrate that a genetically incorporated trifluoromethyl group can be used as a sensitive 19F NMR probe to study protein folding/unfolding, and that genetically incorporated reactive functional groups such as ketone, alkyne, and azide moieties can be applied to site-specifically label proteins with florescent probes. This critical discovery allows the synthesis of proteins with diverse bioorthogonal functional groups for a variety of basic studies and biotechnology development using a single recombinant expression system. PMID:23138887

Quantitative chemoproteomics for site-specific analysis of protein alkylation by 4-hydroxy-2-nonenal in cells.

PubMed

Yang, Jing; Tallman, Keri A; Porter, Ned A; Liebler, Daniel C

2015-03-03

Protein alkylation by 4-hydroxy-2-nonenal (HNE), an endogenous lipid derived electrophile, contributes to stress signaling and cellular toxicity. Although previous work has identified protein targets for HNE alkylation, the sequence specificity of alkylation and dynamics in a cellular context remain largely unexplored. We developed a new quantitative chemoproteomic platform, which uses isotopically tagged, photocleavable azido-biotin reagents to selectively capture and quantify the cellular targets labeled by the alkynyl analogue of HNE (aHNE). Our analyses site-specifically identified and quantified 398 aHNE protein alkylation events (386 cysteine sites and 12 histidine sites) in intact cells. This data set expands by at least an order of magnitude the number of such modification sites previously reported. Although adducts formed by Michael addition are thought to be largely irreversible, we found that most aHNE modifications are lost rapidly in situ. Moreover, aHNE adduct turnover occurs only in intact cells and loss rates are site-selective. This quantitative chemoproteomics platform provides a versatile general approach to map bioorthogonal-chemically engineered post-translational modifications and their cellular dynamics in a site-specific and unbiased manner.

Streptococci Engage TLR13 on Myeloid Cells in a Site-Specific Fashion.

PubMed

Kolter, Julia; Feuerstein, Reinhild; Spoeri, Evelyne; Gharun, Kourosh; Elling, Roland; Trieu-Cuot, Patrick; Goldmann, Tobias; Waskow, Claudia; Chen, Zhijian J; Kirschning, Carsten J; Deshmukh, Sachin D; Henneke, Philipp

2016-03-15

Streptococci are common human colonizers with a species-specific mucocutaneous distribution. At the same time, they are among the most important and most virulent invasive bacterial pathogens. Thus, site-specific cellular innate immunity, which is predominantly executed by resident and invading myeloid cells, has to be adapted with respect to streptococcal sensing, handling, and response. In this article, we show that TLR13 is the critical mouse macrophage (MΦ) receptor in the response to group B Streptococcus, both in bone marrow-derived MΦs and in mature tissue MΦs, such as those residing in the lamina propria of the colon and the dermis, as well as in microglia. In contrast, TLR13 and its chaperone UNC-93B are dispensable for a potent cytokine response of blood monocytes to group B Streptococcus, although monocytes serve as the key progenitors of intestinal and dermal MΦs. Furthermore, a specific role for TLR13 with respect to MΦ function is supported by the response to staphylococci, where TLR13 and UNC-93B limit the cytokine response in bone marrow-derived MΦs and microglia, but not in dermal MΦs. In summary, TLR13 is a critical and site-specific receptor in the single MΦ response to β-hemolytic streptococci. Copyright © 2016 by The American Association of Immunologists, Inc.

Characterization of bacteriophage communities and CRISPR profiles from dental plaque

PubMed Central

2014-01-01

Background Dental plaque is home to a diverse and complex community of bacteria, but has generally been believed to be inhabited by relatively few viruses. We sampled the saliva and dental plaque from 4 healthy human subjects to determine whether plaque was populated by viral communities, and whether there were differences in viral communities specific to subject or sample type. Results We found that the plaque was inhabited by a community of bacteriophage whose membership was mostly subject-specific. There was a significant proportion of viral homologues shared between plaque and salivary viromes within each subject, suggesting that some oral viruses were present in both sites. We also characterized Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) in oral streptococci, as their profiles provide clues to the viruses that oral bacteria may be able to counteract. While there were some CRISPR spacers specific to each sample type, many more were shared across sites and were highly subject specific. Many CRISPR spacers matched viruses present in plaque, suggesting that the evolution of CRISPR loci may have been specific to plaque-derived viruses. Conclusions Our findings of subject specificity to both plaque-derived viruses and CRISPR profiles suggest that human viral ecology may be highly personalized. PMID:24981669

Site-specific protein labeling with PRIME and chelation-assisted Click chemistry

PubMed Central

Uttamapinant, Chayasith; Sanchez, Mateo I.; Liu, Daniel S.; Yao, Jennifer Z.; White, Katharine A.; Grecian, Scott; Clarke, Scott; Gee, Kyle R.; Ting, Alice Y.

2016-01-01

This protocol describes an efficient method to site-specifically label cell-surface or purified proteins with chemical probes in two steps: PRobe Incorporation Mediated by Enzymes (PRIME) followed by chelation-assisted copper-catalyzed azide-alkyne cycloaddition (CuAAC). In the PRIME step, Escherichia coli lipoic acid ligase site-specifically attaches a picolyl azide derivative to a 13-amino acid recognition sequence that has been genetically fused onto the protein of interest. Proteins bearing picolyl azide are chemoselectively derivatized with an alkyne-probe conjugate by chelation-assisted CuAAC in the second step. We describe herein the optimized protocols to synthesize picolyl azide, perform PRIME labeling, and achieve CuAAC derivatization of picolyl azide on live cells, fixed cells, and purified proteins. Reagent preparations, including synthesis of picolyl azide probes and expression of lipoic acid ligase, take 12 d, while the procedure to perform site-specific picolyl azide ligation and CuAAC on cells or on purified proteins takes 40 min-3 h. PMID:23887180

Cobalt Oxide Hollow Nanoparticles Derived by Bio-Templating

NASA Technical Reports Server (NTRS)

Kim, Jae-Woo; Choi, Sang H.; Lillehei, Peter T.; Chu, Sang-Hyon; King, Glen C.; Watt, Gerald D.

2005-01-01

We present here the first fabrication of hollow cobalt oxide nanoparticles produced by a protein-regulated site-specific reconstitution process in aqueous solution and describe the metal growth mechanism in the ferritin interior.

HOX and TALE signatures specify human stromal stem cell populations from different sources.

PubMed

Picchi, Jacopo; Trombi, Luisa; Spugnesi, Laura; Barachini, Serena; Maroni, Giorgia; Brodano, Giovanni Barbanti; Boriani, Stefano; Valtieri, Mauro; Petrini, Mario; Magli, Maria Cristina

2013-04-01

Human stromal stem cell populations reside in different tissues and anatomical sites, however a critical question related to their efficient use in regenerative medicine is whether they exhibit equivalent biological properties. Here, we compared cellular and molecular characteristics of stromal stem cells derived from the bone marrow, at different body sites (iliac crest, sternum, and vertebrae) and other tissues (dental pulp and colon). In particular, we investigated whether homeobox genes of the HOX and TALE subfamilies might provide suitable markers to identify distinct stromal cell populations, as HOX proteins control cell positional identity and, together with their co-factors TALE, are involved in orchestrating differentiation of adult tissues. Our results show that stromal populations from different sources, although immunophenotypically similar, display distinct HOX and TALE signatures, as well as different growth and differentiation abilities. Stromal stem cells from different tissues are characterized by specific HOX profiles, differing in the number and type of active genes, as well as in their level of expression. Conversely, bone marrow-derived cell populations can be essentially distinguished for the expression levels of specific HOX members, strongly suggesting that quantitative differences in HOX activity may be crucial. Taken together, our data indicate that the HOX and TALE profiles provide positional, embryological and hierarchical identity of human stromal stem cells. Furthermore, our data suggest that cell populations derived from different body sites may not represent equivalent cell sources for cell-based therapeutical strategies for regeneration and repair of specific tissues. Copyright © 2012 Wiley Periodicals, Inc.

Evaluating meteorological data from weather stations, and from satellites and global models for a multi-site epidemiological study.

PubMed

Colston, Josh M; Ahmed, Tahmeed; Mahopo, Cloupas; Kang, Gagandeep; Kosek, Margaret; de Sousa Junior, Francisco; Shrestha, Prakash Sunder; Svensen, Erling; Turab, Ali; Zaitchik, Benjamin

2018-04-21

Longitudinal and time series analyses are needed to characterize the associations between hydrometeorological parameters and health outcomes. Earth Observation (EO) climate data products derived from satellites and global model-based reanalysis have the potential to be used as surrogates in situations and locations where weather-station based observations are inadequate or incomplete. However, these products often lack direct evaluation at specific sites of epidemiological interest. Standard evaluation metrics of correlation, agreement, bias and error were applied to a set of ten hydrometeorological variables extracted from two quasi-global, commonly used climate data products - the Global Land Data Assimilation System (GLDAS) and Climate Hazards Group InfraRed Precipitation with Stations (CHIRPS) - to evaluate their performance relative to weather-station derived estimates at the specific geographic locations of the eight sites in a multi-site cohort study. These metrics were calculated for both daily estimates and 7-day averages and for a rotavirus-peak-season subset. Then the variables from the two sources were each used as predictors in longitudinal regression models to test their association with rotavirus infection in the cohort after adjusting for covariates. The availability and completeness of station-based validation data varied depending on the variable and study site. The performance of the two gridded climate models varied considerably within the same location and for the same variable across locations, according to different evaluation criteria and for the peak-season compared to the full dataset in ways that showed no obvious pattern. They also differed in the statistical significance of their association with the rotavirus outcome. For some variables, the station-based records showed a strong association while the EO-derived estimates showed none, while for others, the opposite was true. Researchers wishing to utilize publicly available climate data - whether EO-derived or station based - are advised to recognize their specific limitations both in the analysis and the interpretation of the results. Epidemiologists engaged in prospective research into environmentally driven diseases should install their own weather monitoring stations at their study sites whenever possible, in order to circumvent the constraints of choosing between distant or incomplete station data or unverified EO estimates. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Platelets secrete stromal cell–derived factor 1α and recruit bone marrow–derived progenitor cells to arterial thrombi in vivo

PubMed Central

Massberg, Steffen; Konrad, Ildiko; Schürzinger, Katrin; Lorenz, Michael; Schneider, Simon; Zohlnhoefer, Dietlind; Hoppe, Katharina; Schiemann, Matthias; Kennerknecht, Elisabeth; Sauer, Susanne; Schulz, Christian; Kerstan, Sandra; Rudelius, Martina; Seidl, Stefan; Sorge, Falko; Langer, Harald; Peluso, Mario; Goyal, Pankaj; Vestweber, Dietmar; Emambokus, Nikla R.; Busch, Dirk H.; Frampton, Jon; Gawaz, Meinrad

2006-01-01

The accumulation of smooth muscle and endothelial cells is essential for remodeling and repair of injured blood vessel walls. Bone marrow–derived progenitor cells have been implicated in vascular repair and remodeling; however, the mechanisms underlying their recruitment to the site of injury remain elusive. Here, using real-time in vivo fluorescence microscopy, we show that platelets provide the critical signal that recruits CD34+ bone marrow cells and c-Kit+ Sca-1+ Lin− bone marrow–derived progenitor cells to sites of vascular injury. Correspondingly, specific inhibition of platelet adhesion virtually abrogated the accumulation of both CD34+ and c-Kit+ Sca-1+ Lin− bone marrow–derived progenitor cells at sites of endothelial disruption. Binding of bone marrow cells to platelets involves both P-selectin and GPIIb integrin on platelets. Unexpectedly, we found that activated platelets secrete the chemokine SDF-1α, thereby supporting further primary adhesion and migration of progenitor cells. These findings establish the platelet as a major player in the initiation of vascular remodeling, a process of fundamental importance for vascular repair and pathological remodeling after vascular injury. PMID:16618794

Platelets secrete stromal cell-derived factor 1alpha and recruit bone marrow-derived progenitor cells to arterial thrombi in vivo.

PubMed

Massberg, Steffen; Konrad, Ildiko; Schürzinger, Katrin; Lorenz, Michael; Schneider, Simon; Zohlnhoefer, Dietlind; Hoppe, Katharina; Schiemann, Matthias; Kennerknecht, Elisabeth; Sauer, Susanne; Schulz, Christian; Kerstan, Sandra; Rudelius, Martina; Seidl, Stefan; Sorge, Falko; Langer, Harald; Peluso, Mario; Goyal, Pankaj; Vestweber, Dietmar; Emambokus, Nikla R; Busch, Dirk H; Frampton, Jon; Gawaz, Meinrad

2006-05-15

The accumulation of smooth muscle and endothelial cells is essential for remodeling and repair of injured blood vessel walls. Bone marrow-derived progenitor cells have been implicated in vascular repair and remodeling; however, the mechanisms underlying their recruitment to the site of injury remain elusive. Here, using real-time in vivo fluorescence microscopy, we show that platelets provide the critical signal that recruits CD34+ bone marrow cells and c-Kit+ Sca-1+ Lin- bone marrow-derived progenitor cells to sites of vascular injury. Correspondingly, specific inhibition of platelet adhesion virtually abrogated the accumulation of both CD34+ and c-Kit+ Sca-1+ Lin- bone marrow-derived progenitor cells at sites of endothelial disruption. Binding of bone marrow cells to platelets involves both P-selectin and GPIIb integrin on platelets. Unexpectedly, we found that activated platelets secrete the chemokine SDF-1alpha, thereby supporting further primary adhesion and migration of progenitor cells. These findings establish the platelet as a major player in the initiation of vascular remodeling, a process of fundamental importance for vascular repair and pathological remodeling after vascular injury.

Microzonation Mapping Of The Yanbu Industrial City, Western Saudi Arabia: A Multicriteria Decision Analysis Approach

NASA Astrophysics Data System (ADS)

Moustafa, Sayed, Sr.; Alarifi, Nassir S.; Lashin, Aref A.

2016-04-01

Urban areas along the western coast of Saudi Arabia are susceptible to natural disasters and environmental damages due to lack of planning. To produce a site-specific microzonation map of the rapidly growing Yanbu industrial city, spatial distribution of different hazard entities are assessed using the Analytical Hierarchal Process (AHP) together with Geographical Information System (GIS). For this purpose six hazard parameter layers are considered, namely; fundamental frequency, site amplification, soil strength in terms of effective shear-wave velocity, overburden sediment thickness, seismic vulnerability index and peak ground acceleration. The weight and rank values are determined during AHP and are assigned to each layer and its corresponding classes, respectively. An integrated seismic microzonation map was derived using GIS platform. Based on the derived map, the study area is classified into five hazard categories: very low, low, moderate high, and very high. The western and central parts of the study area, as indicated from the derived microzonation map, are categorized as a high hazard zone as compared to other surrounding places. The produced microzonation map of the current study is envisaged as a first-level assessment of the site specific hazards in the Yanbu city area, which can be used as a platform by different stakeholders in any future land-use planning and environmental hazard management.

Specific interactions between mycobacterial FtsZ protein and curcumin derivatives: Molecular docking and ab initio molecular simulations

NASA Astrophysics Data System (ADS)

Fujimori, Mitsuki; Sogawa, Haruki; Ota, Shintaro; Karpov, Pavel; Shulga, Sergey; Blume, Yaroslav; Kurita, Noriyuki

2018-01-01

Filamentous temperature-sensitive Z (FtsZ) protein plays essential role in bacteria cell division, and its inhibition prevents Mycobacteria reproduction. Here we adopted curcumin derivatives as candidates of novel inhibitors and investigated their specific interactions with FtsZ, using ab initio molecular simulations based on protein-ligand docking, classical molecular mechanics and ab initio fragment molecular orbital (FMO) calculations. Based on FMO calculations, we specified the most preferable site of curcumin binding to FtsZ and highlighted the key amino acid residues for curcumin binding at an electronic level. The result will be useful for proposing novel inhibitors against FtsZ based on curcumin derivatives.

Site-selective post-translational modification of proteins using an unnatural amino acid, 3-azidotyrosine.

PubMed

Ohno, Satoshi; Matsui, Megumi; Yokogawa, Takashi; Nakamura, Masashi; Hosoya, Takamitsu; Hiramatsu, Toshiyuki; Suzuki, Masaaki; Hayashi, Nobuhiro; Nishikawa, Kazuya

2007-03-01

An efficient method for site-selective modification of proteins using an unnatural amino acid, 3-azidotyrosine has been developed. This method utilizes the yeast amber suppressor tRNA(Tyr)/mutated tyrosyl-tRNA synthetase pair as a carrier of 3-azidotyrosine in an Escherichia coli cell-free translation system, and triarylphosphine derivatives for specific modification of the azido group. Using rat calmodulin (CaM) as a model protein, we prepared several unnatural CaM molecules, each carrying an azidotyrosine at predetermined positions 72, 78, 80 or 100, respectively. Post-translational modification of these proteins with a conjugate compound of triarylphosphine and biotin produced site-selectively biotinylated CaM molecules. Reaction efficiency was similar among these proteins irrespective of the position of introduction, and site-specificity of biotinylation was confirmed using mass spectrometry. In addition, CBP-binding activity of the biotinylated CaMs was confirmed to be similar to that of wild-type CaM. This method is intrinsically versatile in that it should be easily applicable to introducing any other desirable compounds (e.g., probes and cross-linkers) into selected sites of proteins as far as appropriate derivative compounds of triarylphosphine could be chemically synthesized. Elucidation of molecular mechanisms of protein functions and protein-to-protein networks will be greatly facilitated by making use of these site-selectively modified proteins.

Designing and Testing of Novel Taxanes to Probe the Highly Complex Mechanisms by Which Taxanes Bind to Microtubules and Cause Cytotoxicity to Cancer Cells

PubMed Central

St. George, Marc; Ayoub, Ahmed T.; Banerjee, Asok; Churchill, Cassandra D. M.; Winter, Philip; Klobukowski, Mariusz; Cass, Carol E.; Ludueña, Richard F.; Tuszynski, Jack A.; Damaraju, Sambasivarao

2015-01-01

Our previous work identified an intermediate binding site for taxanes in the microtubule nanopore. The goal of this study was to test derivatives of paclitaxel designed to bind to this intermediate site differentially depending on the isotype of β-tubulin. Since β-tubulin isotypes have tissue-dependent expression—specifically, the βIII isotype is very abundant in aggressive tumors and much less common in normal tissues—this is expected to lead to tubulin targeted drugs that are more efficacious and have less side effects. Seven derivatives of paclitaxel were designed and four of these were amenable for synthesis in sufficient purity and yield for further testing in breast cancer model cell lines. None of the derivatives studied were superior to currently used taxanes, however computer simulations provided insights into the activity of the derivatives. Our results suggest that neither binding to the intermediate binding site nor the final binding site is sufficient to explain the activities of the derivative taxanes studied. These findings highlight the need to iteratively improve on the design of taxanes based on their activity in model systems. Knowledge gained on the ability of the engineered drugs to bind to targets and bring about activity in a predictable manner is a step towards personalizing therapies. PMID:26052950

Biological Evaluation in Vitro and in Silico of Azetidin-2-one Derivatives as Potential Anticancer Agents.

PubMed

Olazaran, Fabián E; Rivera, Gildardo; Pérez-Vázquez, Alondra M; Morales-Reyes, Cynthia M; Segura-Cabrera, Aldo; Balderas-Rentería, Isaías

2017-01-12

Potential anticancer activity of 16 azetidin-2-one derivatives was evaluated showing that compound 6 [ N -( p -methoxy-phenyl)-2-( p -methyl-phenyl)-3-phenoxy-azetidin-2-one] presented cytotoxic activity in SiHa cells and B16F10 cells. The caspase-3 assay in B16F10 cells displayed that azetidin-2-one derivatives induce apoptosis. Microarray and molecular analysis showed that compound 6 was involved on specific gene overexpression of cytoskeleton regulation and apoptosis due to the inhibition of some cell cycle genes. From the 16 derivatives, compound 6 showed the highest selectivity to neoplastic cells, it was an inducer of apoptosis, and according to an in silico analysis of chemical interactions with colchicine binding site of human α/β-tubulin, the mechanism of action could be a molecular interaction involving the amino acids outlining such binding site.

Biological Evaluation in Vitro and in Silico of Azetidin-2-one Derivatives as Potential Anticancer Agents

PubMed Central

2016-01-01

Potential anticancer activity of 16 azetidin-2-one derivatives was evaluated showing that compound 6 [N-(p-methoxy-phenyl)-2-(p-methyl-phenyl)-3-phenoxy-azetidin-2-one] presented cytotoxic activity in SiHa cells and B16F10 cells. The caspase-3 assay in B16F10 cells displayed that azetidin-2-one derivatives induce apoptosis. Microarray and molecular analysis showed that compound 6 was involved on specific gene overexpression of cytoskeleton regulation and apoptosis due to the inhibition of some cell cycle genes. From the 16 derivatives, compound 6 showed the highest selectivity to neoplastic cells, it was an inducer of apoptosis, and according to an in silico analysis of chemical interactions with colchicine binding site of human α/β-tubulin, the mechanism of action could be a molecular interaction involving the amino acids outlining such binding site. PMID:28105271

Heritable Individual-Specific and Allele-Specific Chromatin Signatures in Humans

PubMed Central

McDaniell, Ryan; Lee, Bum-Kyu; Song, Lingyun; Liu, Zheng; Boyle, Alan P.; Erdos, Michael R.; Scott, Laura J.; Morken, Mario A.; Kucera, Katerina S.; Battenhouse, Anna; Keefe, Damian; Collins, Francis S.; Willard, Huntington F.; Lieb, Jason D.; Furey, Terrence S.; Crawford, Gregory E.; Iyer, Vishwanath R.; Birney, Ewan

2010-01-01

The extent to which variation in chromatin structure and transcription factor binding may influence gene expression, and thus underlie or contribute to variation in phenotype, is unknown. To address this question, we cataloged both individual-to-individual variation and differences between homologous chromosomes within the same individual (allele-specific variation) in chromatin structure and transcription factor binding in lymphoblastoid cells derived from individuals of geographically diverse ancestry. Ten percent of active chromatin sites were individual-specific; a similar proportion were allele-specific. Both individual-specific and allele-specific sites were commonly transmitted from parent to child, which suggests that they are heritable features of the human genome. Our study shows that heritable chromatin status and transcription factor binding differ as a result of genetic variation and may underlie phenotypic variation in humans. PMID:20299549

Validation of ozone response functions for annual Mediterranean pasture species using close-to-field-conditions experiments.

PubMed

González-Fernández, Ignacio; Sanz, Javier; Calvete-Sogo, Héctor; Elvira, Susana; Alonso, Rocío; Bermejo-Bermejo, Victoria

2017-12-01

Ozone (O 3 ) critical levels have been established under the Long-Range Transboundary Air Pollution Convention to assess the risk of O 3 effects in European vegetation. A recent review study has led to the development of O 3 critical levels for annual Mediterranean pasture species using plants growing in well-watered pots at a coastal site and under low levels of competition. However, uncertainties remain in the extrapolation of the O 3 sensitivity of these species under natural conditions. The response of two O 3 -sensitive annual Mediterranean pasture Trifolium species at the coastal site was compared with the response of the same species growing at a continental site, in natural soil and subject to water-stress and inter-specific competition, representing more closely their natural habitat. The slopes of exposure- and dose-response relationships derived for the two sites showed differences in the response to O 3 between sites attributed to differences in environmental growing conditions, growing medium and the level of inter-specific competition, but the effect of the individual factors could not be assessed separately. Dose-based O 3 indices partially explained differences due to environmental growing conditions between sites. The slopes showed that plants were more sensitive to O 3 at the continental site, but homogeneity of slopes tests revealed that results from both experimental sites may be combined. Although more experimental data considering complex inter-specific competition situations and the effect of important interactive factors such as nitrogen would be needed, these results confirm the validity of applying the current flux-based O 3 critical level under close to natural growing conditions. The AOT40-based O 3 critical level derived at the coastal site was also considered a suitable risk indicator in close to natural growing conditions in the absence of soil moisture limitations on plant growth.

Site-specific standard request for Underground Storage Tanks 1219-U, 1222-U, 2082-U, and 2068-U at the Rust Garage Facility Buildings 9754-1 and 9720-15

DOE Office of Scientific and Technical Information (OSTI.GOV)

Not Available

1994-08-01

This document is a site-specific standard request for underground storage tanks located at the Rust Garage Facility. These standards are justified based on conclusion derived from the exposure assessment that indicates there is no current or forseeable future human health risk associated with petroleum contaminants on the site, that current and future ecological risks would be generally limited to subsurface species and plant life with roots extending into the area, and that most of the impacted area at the site is covered by asphalt or concrete. The vertical and horizontal extent of soil and ground water contamination are limited tomore » immediate area of the Rust Garage Facility.« less

Chemoproteomics Reveals Chemical Diversity and Dynamics of 4-Oxo-2-nonenal Modifications in Cells.

PubMed

Sun, Rui; Fu, Ling; Liu, Keke; Tian, Caiping; Yang, Yong; Tallman, Keri A; Porter, Ned A; Liebler, Daniel C; Yang, Jing

2017-10-01

4-Oxo-2-nonenal (ONE) derived from lipid peroxidation modifies nucleophiles and transduces redox signaling by its reactions with proteins. However, the molecular interactions between ONE and complex proteomes and their dynamics in situ remain largely unknown. Here we describe a quantitative chemoproteomic analysis of protein adduction by ONE in cells, in which the cellular target profile of ONE is mimicked by its alkynyl surrogate. The analyses reveal four types of ONE-derived modifications in cells, including ketoamide and Schiff-base adducts to lysine, Michael adducts to cysteine, and a novel pyrrole adduct to cysteine. ONE-derived adducts co-localize and exhibit crosstalk with many histone marks and redox sensitive sites. All four types of modifications derived from ONE can be reversed site-specifically in cells. Taken together, our study provides much-needed mechanistic insights into the cellular signaling and potential toxicities associated with this important lipid derived electrophile. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Variable effects of plant colonization on black slate uptake into microbial PLFAs

NASA Astrophysics Data System (ADS)

Seifert, Anne-Gret; Trumbore, Susan; Xu, Xiaomei; Zhang, Dachung; Gleixner, Gerd

2013-04-01

Microbial degradation of carbon derived from black shale and slate has been shown in vitro. However, in natural settings where other labile carbon sources are likely to exist, this has not been previously demonstrated. We investigated the uptake of ancient carbon derived from slate weathering and from recently photosynthesised organic matter by different groups of microorganisms. Therefore we isolated microbial biomarkers (phospholipid fatty acids, PLFAs) from black slates collected at a chronosequence of waste piles which differed in age and vegetation cover. We quantified the amount of PLFAs and performed stable isotope and radiocarbon measurements on individual or grouped PLFAs to quantify the fraction of slate derived carbon. We used black slate from a pile heaped in the 1950s with either uncovered black slate material (bare site) or material slightly colonized by small plants (greened site) and from a forested leaching pile (forested site) used for alum-mining in the 19th century. Colonization by plants influenced the amount and composition of the microbial community. Greater amounts of PLFAs (5410 ng PLFA/g dw) were extracted from slate sampled at the forested site as opposed to the bare site (960 ng PLFAs/g dw) or the greened (annual grasses and mosses) rock waste pile (1050 ng PLFAs/g dw). We found the highest proportion of PLFAs representing Gram-negative bacteria on the forested site and the highest proportion of PLFAs representing Gram-positive bacteria on the bare site. The fungal PLFA was most abundant at the greened site. Sites with less plant colonization (bare and greened site) tended to have more depleted δ13C values compared to the forested site. Radiocarbon measurements on PLFAs indicated that fungi and Gram-positive bacteria were best adapted to black slate carbon uptake. In the fungal PLFA (combined bare and greened waste pile sample) and in PLFAs of Gram-positive bacteria (greened site) we measured 39.7% and 28.9% ancient carbon uptake, respectively. Our results prove that black slate degradation followed by carbon uptake takes place in situ. Results imply that plant colonization might additionally affect this process. Slight colonization with few plants increased slate derived carbon uptake in PLFAs of Gram-positive bacteria. Evidently, Gram-positive bacteria represented by specific PLFAs from the greened site held more ancient carbon than from the bare site. In contrast, no black slate derived carbon was used by microorganisms at the forested site with 2-3 times greater carbon content. Results suggest that the use of ancient slate derived carbon dominates mainly in early stages of microbial colonization of surfaces and that with increasing ecosystem development recycling of plant derived carbon dominates.

Geophysical constraints on contaminant transport modeling in a heterogeneous fluvial aquifer.

PubMed

Bowling, Jerry C; Zheng, Chunmiao; Rodriguez, Antonio B; Harry, Dennis L

2006-05-05

Approximately 3000 measurements of hydraulic conductivity in over 50 flowmeter boreholes were available at the Macro-Dispersion Experiment (MADE) site in Columbus, Mississippi, USA to quantify the heterogeneity in hydraulic conductivity at the site scale. This high-density measurement approach is perhaps infeasible for time and expense in typical groundwater remediation sites. A natural-gradient tracer experiment from the MADE site was simulated by a groundwater flow and solute transport model incorporating direct-current (DC) resistivity data collected over the observed plume location. Hydraulic conductivity from one borehole collected during the original site characterization was used to calibrate the electrical resistivity data to hydraulic conductivity using a previously derived log-log relationship. Application of this relationship, using site-specific empirical constants determined from the data, transforms the 3D electrical resistivity data into a 3D description of hydraulic conductivity that can be used in groundwater models. The validity of this approach was tested by using the geophysically derived hydraulic conductivity representation in numerical simulations of the natural-gradient tracer experiment. The agreement between the simulated and observed tracer plumes was quantified to gauge the effectiveness of geophysically derived and flowmeter based representations of the hydraulic conductivity field. This study demonstrates that a highly heterogeneous aquifer can be modeled with minimal hydrological data supplemented with geophysical data at least as well as previous models of the site using purely hydrologic data.

In silico site-directed mutagenesis informs species-specific predictions of chemical susceptibility derived from the Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS) tool

EPA Science Inventory

The Sequence Alignment to Predict Across Species Susceptibility (SeqAPASS) tool was developed to address needs for rapid, cost effective methods of species extrapolation of chemical susceptibility. Specifically, the SeqAPASS tool compares the primary sequence (Level 1), functiona...

Joint maximum-likelihood magnitudes of presumed underground nuclear test explosions

NASA Astrophysics Data System (ADS)

Peacock, Sheila; Douglas, Alan; Bowers, David

2017-08-01

Body-wave magnitudes (mb) of 606 seismic disturbances caused by presumed underground nuclear test explosions at specific test sites between 1964 and 1996 have been derived from station amplitudes collected by the International Seismological Centre (ISC), by a joint inversion for mb and station-specific magnitude corrections. A maximum-likelihood method was used to reduce the upward bias of network mean magnitudes caused by data censoring, where arrivals at stations that do not report arrivals are assumed to be hidden by the ambient noise at the time. Threshold noise levels at each station were derived from the ISC amplitudes using the method of Kelly and Lacoss, which fits to the observed magnitude-frequency distribution a Gutenberg-Richter exponential decay truncated at low magnitudes by an error function representing the low-magnitude threshold of the station. The joint maximum-likelihood inversion is applied to arrivals from the sites: Semipalatinsk (Kazakhstan) and Novaya Zemlya, former Soviet Union; Singer (Lop Nor), China; Mururoa and Fangataufa, French Polynesia; and Nevada, USA. At sites where eight or more arrivals could be used to derive magnitudes and station terms for 25 or more explosions (Nevada, Semipalatinsk and Mururoa), the resulting magnitudes and station terms were fixed and a second inversion carried out to derive magnitudes for additional explosions with three or more arrivals. 93 more magnitudes were thus derived. During processing for station thresholds, many stations were rejected for sparsity of data, obvious errors in reported amplitude, or great departure of the reported amplitude-frequency distribution from the expected left-truncated exponential decay. Abrupt changes in monthly mean amplitude at a station apparently coincide with changes in recording equipment and/or analysis method at the station.

The λ Integrase Site-specific Recombination Pathway

PubMed Central

LANDY, ARTHUR

2017-01-01

The site-specific recombinase encoded by bacteriophage λ (Int) is responsible for integrating and excising the viral chromosome into and out of the chromosome of its Escherichia coli host. Int carries out a reaction that is highly directional, tightly regulated, and depends upon an ensemble of accessory DNA bending proteins acting on 240 bp of DNA encoding 16 protein binding sites. This additional complexity enables two pathways, integrative and excisive recombination, whose opposite, and effectively irreversible, directions are dictated by different physiological and environmental signals. Int recombinase is a heterobivalent DNA binding protein and each of the four Int protomers, within a multiprotein 400 kDa recombinogenic complex, is thought to bind and, with the aid of DNA bending proteins, bridge one arm- and one core-type DNA site. In the 12 years since the publication of the last review focused solely on the λ site-specific recombination pathway in Mobile DNA II, there has been a great deal of progress in elucidating the molecular details of this pathway. The most dramatic advances in our understanding of the reaction have been in the area of X-ray crystallography where protein-DNA structures have now been determined for of all of the DNA-protein interfaces driving the Int pathway. Building on this foundation of structures, it has been possible to derive models for the assembly of components that determine the regulatory apparatus in the P-arm, and for the overall architectures that define excisive and integrative recombinogenic complexes. The most fundamental additional mechanistic insights derive from the application of hexapeptide inhibitors and single molecule kinetics. PMID:26104711

Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen*

PubMed Central

Terajima, Masahiko; Taga, Yuki; Chen, Yulong; Cabral, Wayne A.; Hou-Fu, Guo; Srisawasdi, Sirivimol; Nagasawa, Masako; Sumida, Noriko; Hattori, Shunji; Kurie, Jonathan M.; Marini, Joan C.; Yamauchi, Mitsuo

2016-01-01

Covalent intermolecular cross-linking provides collagen fibrils with stability. The cross-linking chemistry is tissue-specific and determined primarily by the state of lysine hydroxylation at specific sites. A recent study on cyclophilin B (CypB) null mice, a model of recessive osteogenesis imperfecta, demonstrated that lysine hydroxylation at the helical cross-linking site of bone type I collagen was diminished in these animals (Cabral, W. A., Perdivara, I., Weis, M., Terajima, M., Blissett, A. R., Chang, W., Perosky, J. E., Makareeva, E. N., Mertz, E. L., Leikin, S., Tomer, K. B., Kozloff, K. M., Eyre, D. R., Yamauchi, M., and Marini, J. C. (2014) PLoS Genet. 10, e1004465). However, the extent of decrease appears to be tissue- and molecular site-specific, the mechanism of which is unknown. Here we report that although CypB deficiency resulted in lower lysine hydroxylation in the helical cross-linking sites, it was increased in the telopeptide cross-linking sites in tendon type I collagen. This resulted in a decrease in the lysine aldehyde-derived cross-links but generation of hydroxylysine aldehyde-derived cross-links. The latter were absent from the wild type and heterozygous mice. Glycosylation of hydroxylysine residues was moderately increased in the CypB null tendon. We found that CypB interacted with all lysyl hydroxylase isoforms (isoforms 1–3) and a putative lysyl hydroxylase-2 chaperone, 65-kDa FK506-binding protein. Tendon collagen in CypB null mice showed severe size and organizational abnormalities. The data indicate that CypB modulates collagen cross-linking by differentially affecting lysine hydroxylation in a site-specific manner, possibly via its interaction with lysyl hydroxylases and associated molecules. This study underscores the critical importance of collagen post-translational modifications in connective tissue formation. PMID:26934917

Cyclophilin-B Modulates Collagen Cross-linking by Differentially Affecting Lysine Hydroxylation in the Helical and Telopeptidyl Domains of Tendon Type I Collagen.

PubMed

Terajima, Masahiko; Taga, Yuki; Chen, Yulong; Cabral, Wayne A; Hou-Fu, Guo; Srisawasdi, Sirivimol; Nagasawa, Masako; Sumida, Noriko; Hattori, Shunji; Kurie, Jonathan M; Marini, Joan C; Yamauchi, Mitsuo

2016-04-29

Covalent intermolecular cross-linking provides collagen fibrils with stability. The cross-linking chemistry is tissue-specific and determined primarily by the state of lysine hydroxylation at specific sites. A recent study on cyclophilin B (CypB) null mice, a model of recessive osteogenesis imperfecta, demonstrated that lysine hydroxylation at the helical cross-linking site of bone type I collagen was diminished in these animals (Cabral, W. A., Perdivara, I., Weis, M., Terajima, M., Blissett, A. R., Chang, W., Perosky, J. E., Makareeva, E. N., Mertz, E. L., Leikin, S., Tomer, K. B., Kozloff, K. M., Eyre, D. R., Yamauchi, M., and Marini, J. C. (2014) PLoS Genet 10, e1004465). However, the extent of decrease appears to be tissue- and molecular site-specific, the mechanism of which is unknown. Here we report that although CypB deficiency resulted in lower lysine hydroxylation in the helical cross-linking sites, it was increased in the telopeptide cross-linking sites in tendon type I collagen. This resulted in a decrease in the lysine aldehyde-derived cross-links but generation of hydroxylysine aldehyde-derived cross-links. The latter were absent from the wild type and heterozygous mice. Glycosylation of hydroxylysine residues was moderately increased in the CypB null tendon. We found that CypB interacted with all lysyl hydroxylase isoforms (isoforms 1-3) and a putative lysyl hydroxylase-2 chaperone, 65-kDa FK506-binding protein. Tendon collagen in CypB null mice showed severe size and organizational abnormalities. The data indicate that CypB modulates collagen cross-linking by differentially affecting lysine hydroxylation in a site-specific manner, possibly via its interaction with lysyl hydroxylases and associated molecules. This study underscores the critical importance of collagen post-translational modifications in connective tissue formation. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

Site-specific accumulation and dynamic change of flavonoids in Apocyni Veneti Folium.

PubMed

Chen, Cui-Hua; Xu, Hu; Liu, Xun-Hong; Zou, Li-Si; Wang, Mei; Liu, Zi-Xiu; Fu, Xing-Sheng; Zhao, Hui; Yan, Ying

2017-12-01

Site-specific accumulation of flavonoids in Apocyni Veneti Folium was determined by laser scanning confocal microscope (LSCM) and the localization of catechins also was observed via vanillin-HCl staining under the conventional optical microscope. The contents of five flavonoids in Apocyni Veneti Folium from different harvest times and growth parts were measured using HPLC method. LSCM observation showed that flavonoids are accumulated in cuticle of epidermal cells and vessel walls, especially in protoplasts and nucleolus of the collenchyma cells and the epidermal cells. Catechins are localized in the palisade parenchyma cells and vessel walls, particularly in the laticifers found in the phloem. On the basis of the difference of the maximal emission wavelength between quercetin and kaempferol derivatives which have fluorescence behavior by appropriate treatment, kaempferol and its derivatives are localized exclusively in the cuticle. Results showed that the content of astragalin in Apocyni Veneti Folium from different parts revealed the decreasing trend, while hyperin and isoquercitrin were higher in June and July analyzed by HPLC. In summary, the site-specific accumulation of flavonoids in Apocyni Veneti Folium can be determined by LSCM and vanillin-HCl staining. The contents of flavonoids in Apocyni Veneti Folium are correlated with harvest times and growth parts. © 2017 Wiley Periodicals, Inc.

Derivation of site-specific surface water quality criteria for the protection of aquatic ecosystems near a Korean military training facility.

PubMed

Jeong, Seung-Woo; An, Youn-Joo

2014-01-01

This study suggested the first Korean site-specific ecological surface water quality criteria for the protection of ecosystems near an artillery range at a Korean military training facility. Surface water quality (SWQ) criteria in Korea address human health protection but do not encompass ecological criteria such as limits for metals and explosives. The first objective of this study was to derive site-specific SWQ criteria for the protection of aquatic ecosystems in Hantan River, Korea. The second objective was to establish discharge criteria for the artillery range to protect the aquatic ecosystems of Hantan River. In this study, we first identified aquatic organisms living in the Hantan River, including fishes, reptiles, invertebrates, phytoplankton, zooplankton, and amphibians. Second, we collected ecotoxicity data for these aquatic organisms and constructed an ecotoxicity database for Cd, Cu, Zn, TNT, and RDX. This study determined the ecological maximum permissible concentrations for metals and explosives based on the ecotoxicity database and suggested ecological surface water quality criteria for the Hantan River by considering analytical detection limits. Discharge limit criteria for the shooting range were determined based on the ecological surface water quality criteria suggested for Hantan River with further consideration of the dilution of the contaminants discharged into the river.

Identification of amino acids in the N-terminal SH2 domain of phospholipase C gamma 1 important in the interaction with epidermal growth factor receptor.

PubMed

Gergel, J R; McNamara, D J; Dobrusin, E M; Zhu, G; Saltiel, A R; Miller, W T

1994-12-13

Photoaffinity labeling and site-directed mutagenesis have been used to identify amino acid residues of the phospholipase C gamma 1 (PLC gamma 1) N-terminal SH2 domain involved in recognition of the activated epidermal growth factor receptor (EGFR). The photoactive amino acid p-benzoylphenylalanine (Bpa) was incorporated into phosphotyrosine-containing peptides derived from EGFR autophosphorylation sites Tyr992 and Tyr1068. Irradiation of these labels in the presence of SH2 domains showed cross-linking which was time-dependent and specific; labeling was inhibited with non-Bpa-containing peptides from EGFR in molar excess. The phosphotyrosine residue on the peptides was important for SH2 recognition, as dephosphorylated peptides did not cross-link. Radiolabeled peptides were used to identify sites of cross-linking to the N-terminal SH2 of PLC gamma 1. Bpa peptide-SH2 complexes were digested with trypsin, and radioactive fragments were purified by HPLC and analyzed by Edman sequencing. These experiments showed Arg562 and an additional site in the alpha A-beta B region of the SH2 domain, most likely Glu587, to be labeled by the Tyr992-derived peptide. Similar analysis of the reaction with the Tyr1068-derived photoaffinity label identified Leu653 as the cross-linked site. Mutation of the neighboring residues of Glu587 decreased photo-cross-linking, emphasizing the importance of this region of the molecule for recognition. These results are consistent with evidence from the v-Src crystal structure and implicate the loop spanning residues Gln640-Ser654 of PLC gamma 1 in specific recognition of phosphopeptides.

Chemical structure determines target organ carcinogenesis in rats

PubMed Central

Carrasquer, C. A.; Malik, N.; States, G.; Qamar, S.; Cunningham, S.L.; Cunningham, A.R.

2012-01-01

SAR models were developed for 12 rat tumour sites using data derived from the Carcinogenic Potency Database. Essentially, the models fall into two categories: Target Site Carcinogen – Non-Carcinogen (TSC-NC) and Target Site Carcinogen – Non-Target Site Carcinogen (TSC-NTSC). The TSC-NC models were composed of active chemicals that were carcinogenic to a specific target site and inactive ones that were whole animal non-carcinogens. On the other hand, the TSC-NTSC models used an inactive category also composed of carcinogens but to any/all other sites but the target site. Leave one out validations produced an overall average concordance value for all 12 models of 0.77 for the TSC-NC models and 0.73 for the TSC-NTSC models. Overall, these findings suggest that while the TSC-NC models are able to distinguish between carcinogens and non-carcinogens, the TSC-NTSC models are identifying structural attributes that associate carcinogens to specific tumour sites. Since the TSC-NTSC models are composed of active and inactive compounds that are genotoxic and non-genotoxic carcinogens, the TSC-NTSC models may be capable of deciphering non-genotoxic mechanisms of carcinogenesis. Together, models of this type may also prove useful in anticancer drug development since they essentially contain chemicals moieties that target specific tumour site. PMID:23066888

Site-Specific Genome Engineering in Human Pluripotent Stem Cells.

PubMed

Merkert, Sylvia; Martin, Ulrich

2016-06-24

The possibility to generate patient-specific induced pluripotent stem cells (iPSCs) offers an unprecedented potential of applications in clinical therapy and medical research. Human iPSCs and their differentiated derivatives are tools for diseases modelling, drug discovery, safety pharmacology, and toxicology. Moreover, they allow for the engineering of bioartificial tissue and are promising candidates for cellular therapies. For many of these applications, the ability to genetically modify pluripotent stem cells (PSCs) is indispensable, but efficient site-specific and safe technologies for genetic engineering of PSCs were developed only recently. By now, customized engineered nucleases provide excellent tools for targeted genome editing, opening new perspectives for biomedical research and cellular therapies.

Human-specific protein isoforms produced by novel splice sites in the human genome after the human-chimpanzee divergence.

PubMed

Kim, Dong Seon; Hahn, Yoonsoo

2012-11-13

Evolution of splice sites is a well-known phenomenon that results in transcript diversity during human evolution. Many novel splice sites are derived from repetitive elements and may not contribute to protein products. Here, we analyzed annotated human protein-coding exons and identified human-specific splice sites that arose after the human-chimpanzee divergence. We analyzed multiple alignments of the annotated human protein-coding exons and their respective orthologous mammalian genome sequences to identify 85 novel splice sites (50 splice acceptors and 35 donors) in the human genome. The novel protein-coding exons, which are expressed either constitutively or alternatively, produce novel protein isoforms by insertion, deletion, or frameshift. We found three cases in which the human-specific isoform conferred novel molecular function in the human cells: the human-specific IMUP protein isoform induces apoptosis of the trophoblast and is implicated in pre-eclampsia; the intronization of a part of SMOX gene exon produces inactive spermine oxidase; the human-specific NUB1 isoform shows reduced interaction with ubiquitin-like proteins, possibly affecting ubiquitin pathways. Although the generation of novel protein isoforms does not equate to adaptive evolution, we propose that these cases are useful candidates for a molecular functional study to identify proteomic changes that might bring about novel phenotypes during human evolution.

Sources and transport of microbial tetraether membrane lipids in Karst Systems

NASA Astrophysics Data System (ADS)

Jex, C.; Blyth, A. J.; McDonald, J.; Woltering, M.; Khan, S.; Baker, A.

2014-12-01

Speleothems preserve organic biomarkers, proxies for surface climate. Microbial-derived lipids, specifically glycerol dialkyl glycerol tetraetheral (GDGT) lipids have been identified in cave deposits and shown to correlate well with surface air temperature using the archaea-derived isoprenoid '(i)GDGT' index of TEX86 and the bacteria derived branched '(b)GDGT' index of MBT/CBT of modern speleothems [1]. Two competing sources for GDGTs in karst systems have been suggested: 1) A soil derived microbial signal dominated by bGDGTs; and 2) An in situ signal dominated by iGDGTs, representative of archaea existing within the cave or overlying bedrock [2]. These findings are yet to be thoroughly tested by characterising the seasonal nature of GDGTs in caves to establish the source and transport pathways within these complex fractured rock systems. Here, we address this and present the results of a yearlong monitoring campaign of GDGTs within two contrasting cave sites from the Yarrangobilly Caves in Kosciuszko national park, SE Australia. The caves are located at a high altitude, semi-arid site. Harriewood cave is dominated by discrete infiltration events throughout the year. Above the cave there are thin soils consisting of loose shallow scree, steep slopes and sparse shrub vegetation. The surface above Jillabenan is characterised by thick red clays of moderate to no slope and Eucalypt dominated forest. As such, these caves provide ideal test sites to characterise the variability in GDGT signals that may be a result of non-temperature related factors, including varying inputs (groundwater vs. in situ growth) or site-specific hydrological conditions. We present data obtained from within the cave: drip waters and in situ collection of GDGTs formed on filter papers left inside the cave throughout the year, and externally sourced signals from soils and their leachates. We also identify key differences in soil pH and cave air temperatures that are best predicted by using cave specific GDGT calibrations of [1]. [1] Blyth et al. 2013. Calibrating the glycerol dialkyl glycerol tetraether temperature signal in speleothems. Geochim Cosmochim Ac. 109, 312-328. [2] Blyth et al. 2014. Contrasting distributions of glycerol dialkyl glycerol tetraethers (GDGTs) in speleothems and associated soils, Org Geochem, 69, 1-10.

Isoprene emission potentials from European oak forests derived from canopy flux measurements: an assessment of uncertainties and inter-algorithm variability

NASA Astrophysics Data System (ADS)

Langford, Ben; Cash, James; Acton, W. Joe F.; Valach, Amy C.; Hewitt, C. Nicholas; Fares, Silvano; Goded, Ignacio; Gruening, Carsten; House, Emily; Kalogridis, Athina-Cerise; Gros, Valérie; Schafers, Richard; Thomas, Rick; Broadmeadow, Mark; Nemitz, Eiko

2017-12-01

Biogenic emission algorithms predict that oak forests account for ˜ 70 % of the total European isoprene budget. Yet the isoprene emission potentials (IEPs) that underpin these model estimates are calculated from a very limited number of leaf-level observations and hence are highly uncertain. Increasingly, micrometeorological techniques such as eddy covariance are used to measure whole-canopy fluxes directly, from which isoprene emission potentials can be calculated. Here, we review five observational datasets of isoprene fluxes from a range of oak forests in the UK, Italy and France. We outline procedures to correct the measured net fluxes for losses from deposition and chemical flux divergence, which were found to be on the order of 5-8 and 4-5 %, respectively. The corrected observational data were used to derive isoprene emission potentials at each site in a two-step process. Firstly, six commonly used emission algorithms were inverted to back out time series of isoprene emission potential, and then an average isoprene emission potential was calculated for each site with an associated uncertainty. We used these data to assess how the derived emission potentials change depending upon the specific emission algorithm used and, importantly, on the particular approach adopted to derive an average site-specific emission potential. Our results show that isoprene emission potentials can vary by up to a factor of 4 depending on the specific algorithm used and whether or not it is used in a big-leaf or canopy environment (CE) model format. When using the same algorithm, the calculated average isoprene emission potential was found to vary by as much as 34 % depending on how the average was derived. Using a consistent approach with version 2.1 of the Model for Emissions of Gases and Aerosols from Nature (MEGAN), we derive new ecosystem-scale isoprene emission potentials for the five measurement sites: Alice Holt, UK (10 500 ± 2500 µg m-2 h-1); Bosco Fontana, Italy (1610 ± 420 µg m-2 h-1); Castelporziano, Italy (121 ± 15 µg m-2 h-1); Ispra, Italy (7590 ± 1070 µg m-2 h-1); and the Observatoire de Haute Provence, France (7990 ± 1010 µg m-2 h-1). Ecosystem-scale isoprene emission potentials were then extrapolated to the leaf-level and compared to previous leaf-level measurements for Quercus robur and Quercus pubescens, two species thought to account for 50 % of the total European isoprene budget. The literature values agreed closely with emission potentials calculated using the G93 algorithm, which were 85 ± 75 and 78 ± 25 µg g-1 h-1 for Q. robur and Q. pubescens, respectively. By contrast, emission potentials calculated using the G06 algorithm, the same algorithm used in a previous study to derive the European budget, were significantly lower, which we attribute to the influence of past light and temperature conditions. Adopting these new G06 specific emission potentials for Q. robur (55 ± 24 µg g-1 h-1) and Q. pubescens (47 ± 16 µg g-1 h-1) reduced the projected European budget by ˜ 17 %. Our findings demonstrate that calculated isoprene emission potentials vary considerably depending upon the specific approach used in their calculation. Therefore, it is our recommendation that the community now adopt a standardised approach to the way in which micrometeorological flux measurements are corrected and used to derive isoprene, and other biogenic volatile organic compounds, emission potentials.

Ursolic acid derivatives as potential antidiabetic agents: In vitro, in vivo, and in silico studies.

PubMed

Guzmán-Ávila, Ricardo; Flores-Morales, Virginia; Paoli, Paolo; Camici, Guido; Ramírez-Espinosa, Juan José; Cerón-Romero, Litzia; Navarrete-Vázquez, Gabriel; Hidalgo-Figueroa, Sergio; Yolanda Rios, Maria; Villalobos-Molina, Rafael; Estrada-Soto, Samuel

2018-03-01

Hit, Lead & Candidate Discovery Protein tyrosine phosphatase 1B (PTP-1B) has attracted interest as a novel target for the treatment of type 2 diabetes, this because its role in the insulin-signaling pathway as a negative regulator. Thus, the aim of current work was to obtain seven ursolic acid derivatives as potential antidiabetic agents with PTP-1B inhibition as main mechanism of action. Furthermore, derivatives 1-7 were submitted in vitro to enzymatic PTP-1B inhibition being 3, 5, and 7 the most active compounds (IC 50  = 5.6, 4.7, and 4.6 μM, respectively). In addition, results were corroborated with in silico docking studies with PTP-1B orthosteric site A and extended binding site B, showed that 3 had polar and Van der Waals interactions in both sites with Lys120, Tyr46, Ser216, Ala217, Ile219, Asp181, Phe182, Gln262, Val49, Met258, and Gly259, showing a docking score value of -7.48 Kcal/mol, being more specific for site A. Moreover, compound 7 showed polar interaction with Gln262 and Van der Waals interactions with Ala217, Phe182, Ile219, Arg45, Tyr46, Arg47, Asp48, and Val49 with a predictive docking score of -6.43 kcal/mol, suggesting that the potential binding site could be localized in the site B adjacent to the catalytic site A. Finally, derivatives 2 and 7 (50 mg/kg) were selected to establish their in vivo antidiabetic effect using a noninsulin-dependent diabetes mice model, showing significant blood glucose lowering compared with control group (p < .05). © 2018 Wiley Periodicals, Inc.

An Lnc RNA (GAS5)/SnoRNA-derived piRNA induces activation of TRAIL gene by site-specifically recruiting MLL/COMPASS-like complexes

PubMed Central

He, Xin; Chen, Xinxin; Zhang, Xue; Duan, Xiaobing; Pan, Ting; Hu, Qifei; Zhang, Yijun; Zhong, Fudi; Liu, Jun; Zhang, Hong; Luo, Juan; Wu, Kang; Peng, Gao; Luo, Haihua; Zhang, Lehong; Li, Xiaoxi; Zhang, Hui

2015-01-01

PIWI-interacting RNA (piRNA) silences the transposons in germlines or induces epigenetic modifications in the invertebrates. However, its function in the mammalian somatic cells remains unknown. Here we demonstrate that a piRNA derived from Growth Arrest Specific 5, a tumor-suppressive long non-coding RNA, potently upregulates the transcription of tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), a proapoptotic protein, by inducing H3K4 methylation/H3K27 demethylation. Interestingly, the PIWIL1/4 proteins, which bind with this piRNA, directly interact with WDR5, resulting in a site-specific recruitment of the hCOMPASS-like complexes containing at least MLL3 and UTX (KDM6A). We have indicated a novel pathway for piRNAs to specially activate gene expression. Given that MLL3 or UTX are frequently mutated in various tumors, the piRNA/MLL3/UTX complex mediates the induction of TRAIL, and consequently leads to the inhibition of tumor growth. PMID:25779046

Preparing the “Soil”: The Premetastatic Niche

PubMed Central

Kaplan, Rosandra N.; Rafii, Shahin; Lyden, David

2010-01-01

Current focus on cancer metastasis has centered on the intrinsic factors regulating the cell autonomous homing of the tumor cells to the metastatic site. Specific up-regulation of fibronectin and clustering of bone marrow–derived cellular infiltrates coexpressing matrix metalloproteinases in distant tissue sites before tumor cell arrival are proving to be indispensable for the initial stages of metastasis. These bone marrow–derived hematopoietic progenitors that express vascular endothelial growth factor receptor 1 mobilize in response to the unique array of growth factors produced by the primary tumor. Their arrival in distant sites represents early changes in the local microenvironment, termed the “premetastatic niche,” which dictate the pattern of metastatic spread. Focus on the early cellular and molecular events in cancer dissemination and selectivity will likely lead to new approaches to detect and prevent metastasis at its earliest inception. PMID:17145848

A-BOMB SURVIVOR SITE-SPECIFIC RADIOGENIC CANCER RISKS ESTIMATES

EPA Science Inventory

A draft manuscript is being prepared that describes ways to improve estimates of risk from radiation that have been derived from A-bomb survivors. The work has been published in the journal Radiation Research volume 169, pages 87-98.

Path spectra derived from inversion of source and site spectra for earthquakes in Southern California

NASA Astrophysics Data System (ADS)

Klimasewski, A.; Sahakian, V. J.; Baltay, A.; Boatwright, J.; Fletcher, J. B.; Baker, L. M.

2017-12-01

A large source of epistemic uncertainty in Ground Motion Prediction Equations (GMPEs) is derived from the path term, currently represented as a simple geometric spreading and intrinsic attenuation term. Including additional physical relationships between the path properties and predicted ground motions would produce more accurate and precise, region-specific GMPEs by reclassifying some of the random, aleatory uncertainty as epistemic. This study focuses on regions of Southern California, using data from the Anza network and Southern California Seismic network to create a catalog of events magnitude 2.5 and larger from 1998 to 2016. The catalog encompasses regions of varying geology and therefore varying path and site attenuation. Within this catalog of events, we investigate several collections of event region-to-station pairs, each of which share similar origin locations and stations so that all events have similar paths. Compared with a simple regional GMPE, these paths consistently have high or low residuals. By working with events that have the same path, we can isolate source and site effects, and focus on the remaining residual as path effects. We decompose the recordings into source and site spectra for each unique event and site in our greater Southern California regional database using the inversion method of Andrews (1986). This model represents each natural log record spectra as the sum of its natural log event and site spectra, while constraining each record to a reference site or Brune source spectrum. We estimate a regional, path-specific anelastic attenuation (Q) and site attenuation (t*) from the inversion site spectra and corner frequency from the inversion event spectra. We then compute the residuals between the observed record data, and the inversion model prediction (event*site spectra). This residual is representative of path effects, likely anelastic attenuation along the path that varies from the regional median attenuation. We examine the residuals for our different sets independently to see how path terms differ between event-to-station collections. The path-specific information gained from this can inform development of terms for regional GMPEs, through understanding of these seismological phenomena.

Efficient Site-Specific Labeling of Proteins via Cysteines

PubMed Central

Kim, Younggyu; Ho, Sam O.; Gassman, Natalie R.; Korlann, You; Landorf, Elizabeth V.; Collart, Frank R.; Weiss, Shimon

2011-01-01

Methods for chemical modifications of proteins have been crucial for the advancement of proteomics. In particular, site-specific covalent labeling of proteins with fluorophores and other moieties has permitted the development of a multitude of assays for proteome analysis. A common approach for such a modification is solvent-accessible cysteine labeling using thiol-reactive dyes. Cysteine is very attractive for site-specific conjugation due to its relative rarity throughout the proteome and the ease of its introduction into a specific site along the protein's amino acid chain. This is achieved by site-directed mutagenesis, most often without perturbing the protein's function. Bottlenecks in this reaction, however, include the maintenance of reactive thiol groups without oxidation before the reaction, and the effective removal of unreacted molecules prior to fluorescence studies. Here, we describe an efficient, specific, and rapid procedure for cysteine labeling starting from well-reduced proteins in the solid state. The efficacy and specificity of the improved procedure are estimated using a variety of single-cysteine proteins and thiol-reactive dyes. Based on UV/vis absorbance spectra, coupling efficiencies are typically in the range 70–90%, and specificities are better than ~95%. The labeled proteins are evaluated using fluorescence assays, proving that the covalent modification does not alter their function. In addition to maleimide-based conjugation, this improved procedure may be used for other thiol-reactive conjugations such as haloacetyl, alkyl halide, and disulfide interchange derivatives. This facile and rapid procedure is well suited for high throughput proteome analysis. PMID:18275130

Efficient site-specific labeling of proteins via cysteines.

PubMed

Kim, Younggyu; Ho, Sam O; Gassman, Natalie R; Korlann, You; Landorf, Elizabeth V; Collart, Frank R; Weiss, Shimon

2008-03-01

Methods for chemical modifications of proteins have been crucial for the advancement of proteomics. In particular, site-specific covalent labeling of proteins with fluorophores and other moieties has permitted the development of a multitude of assays for proteome analysis. A common approach for such a modification is solvent-accessible cysteine labeling using thiol-reactive dyes. Cysteine is very attractive for site-specific conjugation due to its relative rarity throughout the proteome and the ease of its introduction into a specific site along the protein's amino acid chain. This is achieved by site-directed mutagenesis, most often without perturbing the protein's function. Bottlenecks in this reaction, however, include the maintenance of reactive thiol groups without oxidation before the reaction, and the effective removal of unreacted molecules prior to fluorescence studies. Here, we describe an efficient, specific, and rapid procedure for cysteine labeling starting from well-reduced proteins in the solid state. The efficacy and specificity of the improved procedure are estimated using a variety of single-cysteine proteins and thiol-reactive dyes. Based on UV/vis absorbance spectra, coupling efficiencies are typically in the range 70-90%, and specificities are better than approximately 95%. The labeled proteins are evaluated using fluorescence assays, proving that the covalent modification does not alter their function. In addition to maleimide-based conjugation, this improved procedure may be used for other thiol-reactive conjugations such as haloacetyl, alkyl halide, and disulfide interchange derivatives. This facile and rapid procedure is well suited for high throughput proteome analysis.

Integrating proximal and satellite optical data for the analysis of ecosystem carbon uptake and plant phenology at the European larch Specnet site

NASA Astrophysics Data System (ADS)

Galvagno, Marta; Gamon, John; Cremonese, Edoardo; Garrity, Steven; Huemmrich, K. Fred; Filippa, Gianluca; Morra di Cella, Umberto; Rossini, Micol

2017-04-01

Automated canopy-level optical sampling in tandem with ecosystem-atmosphere flux observations is continuously carried on at a variety of ecosystems through the Specnet network (http://specnet.info/). Specifically, 9 sites within US and Europe were selected since 2015, to investigate the use of novel NDVI and PRI low-cost sensors for the analysis of ecosystem functioning and phenology. Different plant functional types, such as grasslands, deciduous, and evergreen forests belong to the network, here we present specific data from the larch (Larix decidua Mill.) forest Italian site. Three automated NDVI and three automated PRI spectral reflectance sensors (Decagon Devices Inc.) were installed in 2015 on the top of the 20-meters eddy covariance tower, pointing toward the west, north, and east orientations. An additional system, composed by one NDVI and PRI system was installed to monitor the understory component. The objective of this analysis is the comparison between these in-situ inexpensive sensors, independent NDVI and PRI sensors (Skye Instruments) previously installed on the 20-meters tower and satellite-derived NDVI. Both MODIS and Sentinel NDVI data were used for the comparison. Moreover, the newly derived chlorophyll/carotenoid index (CCI, Gamon et al. 2016), computed as the normalized difference between the NDVI red band and PRI 532 nm band, was tested to estimate the seasonal pattern of daily Gross Primary Productivity (GPP) of the larch forest. Results showed that the seasonality of NDVI was comparable among in-situ sensors and satellite data, though orientation-specific differences were observed. Both NDVI and CCI tracked daily GPP, but with different sensitivity to its seasonality. Future analysis will be directed toward a comparison between this site-based results with the other sites within the Specnet network.

A Model-based Approach to Scaling GPP and NPP in Support of MODIS Land Product Validation

NASA Astrophysics Data System (ADS)

Turner, D. P.; Cohen, W. B.; Gower, S. T.; Ritts, W. D.

2003-12-01

Global products from the Earth-orbiting MODIS sensor include land cover, leaf area index (LAI), FPAR, 8-day gross primary production (GPP), and annual net primary production (NPP) at the 1 km spatial resolution. The BigFoot Project was designed specifically to validate MODIS land products, and has initiated ground measurements at 9 sites representing a wide array of vegetation types. An ecosystem process model (Biome-BGC) is used to generate estimates of GPP and NPP for each 5 km x 5 km BigFoot site. Model inputs include land cover and LAI (from Landsat ETM+), daily meteorological data (from a centrally located eddy covariance flux tower), and soil characteristics. Model derived outputs are validated against field-measured NPP and flux tower-derived GPP. The resulting GPP and NPP estimates are then aggregated to the 1 km resolution for direct spatial comparison with corresponding MODIS products. At the high latitude sites (tundra and boreal forest), the MODIS GPP phenology closely tracks the BigFoot GPP, but there is a high bias in the MODIS GPP. In the temperate zone sites, problems with the timing and magnitude of the MODIS FPAR introduce differences in MODIS GPP compared to the validation data at some sites. However, the MODIS LAI/FPAR data are currently being reprocessed (=Collection 4) and new comparisons will be made for 2002. The BigFoot scaling approach permits precise overlap in spatial and temporal resolution between the MODIS products and BigFoot products, and thus permits the evaluation of specific components of the MODIS NPP algorithm. These components include meteorological inputs from the NASA Data Assimilation Office, LAI and FPAR from other MODIS algorithms, and biome-specific parameters for base respiration rate and light use efficiency.

Specificity of the Antibody Receptor Site to D-Lysergamide: Model of a Physiological Receptor for Lysergic Acid Diethylamide

PubMed Central

Vunakis, Helen Van; Farrow, John T.; Gjika, Hilda B.; Levine, Lawrence

1971-01-01

Antibodies to D-lysergic acid have been produced in rabbits and guinea pigs and a radioimmunoassay for the hapten was developed. The specificity of this lysergamide-antilysergamide reaction was determined by competitive binding with unlabeled lysergic acid diethylamide (LSD), psychotomimetic drugs, neurotransmitters, and other compounds with diverse structures. LSD and several related ergot alkaloids were potent competitors, three to seven times more potent than lysergic acid itself. The N,N-dimethyl derivatives of several compounds, including tryptamine, 5-hydroxytryptamine, 4-hydroxytryptamine, 5-methoxytryptamine, tyramine, and mescaline, were only about ten times less effective than lysergic acid, even though these compounds lack some of the ring systems of lysergic acid. The pattern of inhibition by related compounds with various substituents suggests that the antibody receptor site recognizes structural features resembling the LSD molecule. In particular, the aromatic nucleus and the dimethylated ethylamine side chain in phenylethylamine and tryptamine derivatives may assume in solution a conformation resembling ring A and the methylated nitrogen in ring C of LSD. Among the tryptamine derivatives, a large percentage of the most potent competitors are also psychotomimetic compounds. PMID:5283939

Specific interactions between amyloid-β peptide and curcumin derivatives: Ab initio molecular simulations

NASA Astrophysics Data System (ADS)

Ishimura, Hiromi; Kadoya, Ryushi; Suzuki, Tomoya; Murakawa, Takeru; Shulga, Sergiy; Kurita, Noriyuki

2015-07-01

Alzheimer's disease is caused by accumulation of amyloid-β (Aβ) peptides in a brain. To suppress the production of Aβ peptides, it is effective to inhibit the cleavage of amyloid precursor protein (APP) by secretases. However, because the secretases also play important roles to produce vital proteins for human body, inhibitors for the secretases may have side effects. To propose new agents for protecting the cleavage site of APP from the attacking of the γ-secretase, we have investigated here the specific interactions between a short APP peptide and curcumin derivatives, using protein-ligand docking as well as ab initio molecular simulations.

Potential soil cleanup objectives for nitrogen-containing fertilizers at agrichemical facilities

USGS Publications Warehouse

Roy, W.R.; Krapac, I.G.

2006-01-01

Accidental and incidental chemical releases of nitrogen-containing fertilizers occur at retail agrichemical facilities. Because contaminated soil may threaten groundwater quality, the facility may require some type of site remediation. The purpose of this study was to apply the concepts of the Soil Screening Levels of the U.S. Environmental Protection Agency to derive soil cleanup objectives (SCO) that are protective of groundwater quality in Illinois for nitrogen as nitrate and as ammonium. The Soil Screening Levels are based on the solute transport mechanisms of sorption, volatilization, and groundwater dilution, and the contaminant-specific groundwater cleanup objective used to derive the SCO. Because nitrate is relatively unreactive, only groundwater dilution could be taken into account in the derivation of a SCO. Using a default groundwater objective for potable groundwater, an SCO of 38 mg N-NO3/kg was derived. For ammonium, however, the extent of sorption was measured using an uncontaminated, surface-soil sample (0 to 15 cm) of 10 different soil types that occur in Illinois and three gravel-fill samples from three different agrichemical facilities. Using a default groundwater objective, an SCO was derived for each soil type. The median SCO was 989 mg N-NH4/kg. The SCO calculated for each of the 10 soil and 3 fill samples was positively correlated with cation exchange capacity, clay content, and surface area. It was concluded that this approach can be used to derive either default of site-specific SCOs for nitrogen as nitrate and as ammonium for chemical releases. Copyright ?? Taylor & Francis Group, LLC.

Soil Segregation Methods for Reducing Transportation and Disposal Costs - 13544

DOE Office of Scientific and Technical Information (OSTI.GOV)

Frothingham, David; Andrews, Shawn; Barker, Michelle

2013-07-01

At Formerly Utilized Sites Remedial Action Program (FUSRAP) sites where the selected alternative for contaminated soil is excavation and off-site disposal, the most significant budget items of the remedial action are the costs for transportation and disposal of soil at an off-site facility. At these sites, the objective is to excavate and dispose of only those soils that exceed derived concentration guideline levels. In situ soil segregation using gross gamma detectors to guide the excavation is often challenging at sites where the soil contamination is overlain by clean soil or where the contaminated soil is located in isolated, subsurface pockets.more » In addition, data gaps are often identified during the alternative evaluation and selection process, resulting in increased uncertainty in the extent of subsurface contamination. In response, the U.S. Army Corps of Engineers, Buffalo District is implementing ex situ soil segregation methods. At the remediated Painesville Site, soils were excavated and fed through a conveyor-belt system, which automatically segregated them into above- and below-cleanup criteria discharge piles utilizing gamma spectroscopy. At the Linde Site and the Shallow Land Disposal Area (SLDA) Site, which are both in the remediation phase, soils are initially segregated during the excavation process using gross gamma detectors and then transported to a pad for confirmatory manual surveying and sampling. At the Linde Site, the ex situ soils are analyzed on the basis of a site-specific method, to establish compliance with beneficial reuse criteria that were developed for the Linde remediation. At the SLDA Site, the ex situ soils are surveyed and sampled based on Multi-Agency Radiation Survey and Site Investigation Manual (MARSSIM) final status survey guidance to demonstrate compliance with the derived concentration guideline levels. At all three sites, the ex situ soils that meet the site- specific DCGLs are retained on-site and used as backfill material. This paper describes the ex situ soil segregation methods, the considerations of each method, and the estimated cost savings from minimizing the volume of soil requiring transportation and off-site disposal. (authors)« less

Management of extravasation of oxaliplatin by mimicking its biotransformation.

PubMed

Bahadori, F; Demiray, M

2018-04-27

Although oxaliplatin (Oxali) plays a key role in the treatment of many types of cancer and has been reported to be an irritant, there is no specific and effective method for its extravasation and failure in Oxali extravasation management results in the need for plastic surgery. In the body, Oxali bio-transforms upon dilution in chloride-containing buffer salts to its di-chloro derivative and loses an oxalate molecule. Consequently, the chloride ions exchange with water molecules in the intracellular environment to produce the di-aqua derivative, which is the most active biotransformation product of Oxali in terms of forming the DNA adducts. Thus, inhibiting transformation of di-chloro to di-aqua derivatives by accumulating chloride ions at the site of extravasation and saturating the Oxali molecule with these ions is a strategy that could help manage extravasation. Injecting normal saline at this site is a simple yet effective way to achieve this goal.

The Biodistribution and Immune Suppressive Effects of Breast Cancer-Derived Exosomes.

PubMed

Wen, Shu Wen; Sceneay, Jaclyn; Lima, Luize Goncalves; Wong, Christina S F; Becker, Melanie; Krumeich, Sophie; Lobb, Richard J; Castillo, Vanessa; Wong, Ke Ni; Ellis, Sarah; Parker, Belinda S; Möller, Andreas

2016-12-01

Small membranous secretions from tumor cells, termed exosomes, contribute significantly to intercellular communication and subsequent reprogramming of the tumor microenvironment. Here, we use optical imaging to determine that exogenously administered fluorescently labeled exosomes derived from highly metastatic murine breast cancer cells distributed predominantly to the lung of syngeneic mice, a frequent site of breast cancer metastasis. At the sites of accumulation, exosomes were taken up by CD45 + bone marrow-derived cells. Subsequent long-term conditioning of naïve mice with exosomes from highly metastatic breast cancer cells revealed the accumulation of myeloid-derived suppressor cells in the lung and liver. This favorable immune suppressive microenvironment was capable of promoting metastatic colonization in the lung and liver, an effect not observed from exosomes derived from nonmetastatic cells and liposome control vesicles. Furthermore, we determined that breast cancer exosomes directly suppressed T-cell proliferation and inhibited NK cell cytotoxicity, and hence likely suppressed the anticancer immune response in premetastatic organs. Together, our findings provide novel insight into the tissue-specific outcomes of breast cancer-derived exosome accumulation and their contribution to immune suppression and promotion of metastases. Cancer Res; 76(23); 6816-27. ©2016 AACR. ©2016 American Association for Cancer Research.

Endogenous lipid- and peptide-derived anti-inflammatory pathways generated with glucocorticoid and aspirin treatment activate the lipoxin A4 receptor

PubMed Central

Perretti, Mauro; Chiang, Nan; La, Mylinh; Fierro, Iolanda M.; Marullo, Stefano; Getting, Stephen J; Solito, Egle; Serhan, Charles N.

2009-01-01

Aspirin (ASA) and dexamethasone (DEX) are widely used anti-inflammatory agents yet their mechanism(s) for blocking polymorphonuclear neutrophil (PMN) accumulation at sites of inflammation remains unclear. Here, we report that inhibition of PMN infiltration by ASA and DEX is a property shared by aspirin-triggered lipoxins (ATL) and the glucocorticoid-induced annexin 1 (ANXA1)-derived peptides that are both generated in vivo and act at the lipoxin A4 receptor (ALXR/FPRL1) to halt PMN diapedesis. These structurally diverse ligands specifically interact directly with recombinant human ALXR demonstrated by specific radioligand binding and function as well as immunoprecipitation of PMN receptors. In addition, the combination of both ATL and ANXA1-derived peptides limited PMN infiltration and reduced production of inflammatory mediators (that is, prostaglandins and chemokines) in vivo. Together, these results indicate functional redundancies in endogenous lipid and peptide anti-inflammatory circuits that are spatially and temporally separate, where both ATL and specific ANXA1-derived peptides act in concert at ALXR to downregulate PMN recruitment to inflammatory loci. PMID:12368905

Human-specific protein isoforms produced by novel splice sites in the human genome after the human-chimpanzee divergence

PubMed Central

2012-01-01

Background Evolution of splice sites is a well-known phenomenon that results in transcript diversity during human evolution. Many novel splice sites are derived from repetitive elements and may not contribute to protein products. Here, we analyzed annotated human protein-coding exons and identified human-specific splice sites that arose after the human-chimpanzee divergence. Results We analyzed multiple alignments of the annotated human protein-coding exons and their respective orthologous mammalian genome sequences to identify 85 novel splice sites (50 splice acceptors and 35 donors) in the human genome. The novel protein-coding exons, which are expressed either constitutively or alternatively, produce novel protein isoforms by insertion, deletion, or frameshift. We found three cases in which the human-specific isoform conferred novel molecular function in the human cells: the human-specific IMUP protein isoform induces apoptosis of the trophoblast and is implicated in pre-eclampsia; the intronization of a part of SMOX gene exon produces inactive spermine oxidase; the human-specific NUB1 isoform shows reduced interaction with ubiquitin-like proteins, possibly affecting ubiquitin pathways. Conclusions Although the generation of novel protein isoforms does not equate to adaptive evolution, we propose that these cases are useful candidates for a molecular functional study to identify proteomic changes that might bring about novel phenotypes during human evolution. PMID:23148531

New Methods for the Site-Selective Placement of Peptides on a Microelectrode Array: Probing VEGF-v107 Binding as Proof of Concept.

PubMed

Graaf, Matthew D; Marquez, Bernadette V; Yeh, Nai-Hua; Lapi, Suzanne E; Moeller, Kevin D

2016-10-21

Cu(I)-catalyzed "click" reactions cannot be performed on a borate ester derived polymer coating on a microelectrode array because the Cu(II) precursor for the catalyst triggers background reactions between both acetylene and azide groups with the polymer surface. Fortunately, the Cu(II)-background reaction can itself be used to site-selectively add the acetylene and azide nucleophiles to the surface of the array. In this way, molecules previously functionalized for use in "click" reactions can be added directly to the array. In a similar fashion, activated esters can be added site-selectively to a borate ester coated array. The new chemistry can be used to explore new biological interactions on the arrays. Specifically, the binding of a v107 derived peptide with both human and murine VEGF was probed using a functionalized microelectrode array.

Globally Averaged Atmospheric CFC-11 Concentrations: Monthly and Annual Data for the Period 1975-1992 (DB1010)

DOE Data Explorer

Khalil, M. A.K. [Oregon Graduate Institute of Science and Technology Portland, Oregon (USA); Rasmussen, R. A. [Oregon Graduate Institute of Science and Technology Portland, Oregon

1996-01-01

This data set presents globally averaged atmospheric concentrations of chlorofluorocarbon 11, known also as CFC-11 or F-11 (chemical name: trichlorofluoromethane; formula: CCl3F). The monthly global average data are derived from flask air samples collected at eight sites in six locations over the period August 1980-July 1992. The sites are Barrow (Alaska), Cape Meares (Oregon), Cape Kumukahi and Mauna Loa (Hawaii), Cape Matatula (American Samoa), Cape Grim (Tasmania), Palmer Station, and the South Pole (Antarctica). At each collection site, monthly averages were obtained from three flask samples collected every week. In addition to the monthly global averages available for 1980-992, this data set also contains annual global average data for 1975-1985. These annual global averages were derived from January measurements at the South Pole and in the Pacific Northwest of the United States (specifically, Washington state and the Oregon coast).

Site-specific acid-base properties of pholcodine and related compounds.

PubMed

Kovács, Z; Hosztafi, S; Noszál, B

2006-11-01

The acid-base properties of pholcodine, a cough-depressant agent, and related compounds including metabolites were studied by 1H NMR-pH titrations, and are characterised in terms of macroscopic and microscopic protonation constants. New N-methylated derivatives were also synthesized in order to quantitate site- and nucleus-specific protonation shifts and to unravel microscopic acid-base equilibria. The piperidine nitrogen was found to be 38 and 400 times more basic than its morpholine counterpart in pholcodine and norpholcodine, respectively. The protonation data show that the molecule of pholcodine bears an average of positive charge of 1.07 at physiological pH, preventing it from entering the central nervous system, a plausible reason for its lack of analgesic or addictive properties. The protonation constants of pholcodine and its derivatives are interpreted by comparing with related molecules of pharmaceutical interest. The pH-dependent relative concentrations of the variously protonated forms of pholcodine and morphine are depicted in distribution diagrams.

Amerindian mitochondrial DNAs have rare Asian mutations at high frequencies, suggesting they derived from four primary maternal lineages.

PubMed Central

Schurr, T G; Ballinger, S W; Gan, Y Y; Hodge, J A; Merriwether, D A; Lawrence, D N; Knowler, W C; Weiss, K M; Wallace, D C

1990-01-01

The mitochondrial DNA (mtDNA) sequence variation of the South American Ticuna, the Central American Maya, and the North American Pima was analyzed by restriction-endonuclease digestion and oligonucleotide hybridization. The analysis revealed that Amerindian populations have high frequencies of mtDNAs containing the rare Asian RFLP HincII morph 6, a rare HaeIII site gain, and a unique AluI site gain. In addition, the Asian-specific deletion between the cytochrome c oxidase subunit II (COII) and tRNA(Lys) genes was also prevalent in both the Pima and the Maya. These data suggest that Amerindian mtDNAs derived from at least four primary maternal lineages, that new tribal-specific variants accumulated as these mtDNAs became distributed throughout the Americas, and that some genetic variation may have been lost when the progenitors of the Ticuna separated from the North and Central American populations. Images Figure 1 PMID:1968708

Two signaling molecules share a phosphotyrosine-containing binding site in the platelet-derived growth factor receptor.

PubMed

Nishimura, R; Li, W; Kashishian, A; Mondino, A; Zhou, M; Cooper, J; Schlessinger, J

1993-11-01

Autophosphorylation sites of growth factor receptors with tyrosine kinase activity function as specific binding sites for Src homology 2 (SH2) domains of signaling molecules. This interaction appears to be a crucial step in a mechanism by which receptor tyrosine kinases relay signals to downstream signaling pathways. Nck is a widely expressed protein consisting exclusively of SH2 and SH3 domains, the overexpression of which causes cell transformation. It has been shown that various growth factors stimulate the phosphorylation of Nck and its association with autophosphorylated growth factor receptors. A panel of platelet-derived growth factor (PDGF) receptor mutations at tyrosine residues has been used to identify the Nck binding site. Here we show that mutation at Tyr-751 of the PDGF beta-receptor eliminates Nck binding both in vitro and in living cells. Moreover, the Y751F PDGF receptor mutant failed to mediate PDGF-stimulated phosphorylation of Nck in intact cells. A phosphorylated Tyr-751 is also required for binding of phosphatidylinositol-3 kinase to the PDGF receptor. Hence, the SH2 domains of p85 and Nck share a binding site in the PDGF receptor. Competition experiments with different phosphopeptides derived from the PDGF receptor suggest that binding of Nck and p85 is influenced by different residues around Tyr-751. Thus, a single tyrosine autophosphorylation site is able to link the PDGF receptor to two distinct SH2 domain-containing signaling molecules.

Site-specific cleavage of the transactivation response site of human immunodeficiency virus RNA with a tat-based chemical nuclease.

PubMed Central

Jayasena, S D; Johnston, B H

1992-01-01

tat, an essential transactivator of gene transcription in the human immunodeficiency virus (HIV), is believed to activate viral gene expression by binding to the transactivation response (TAR) site located at the 5' end of all viral mRNAs. The TAR element forms a stem-loop structure containing a 3-nucleotide bulge that is the site for tat binding and is required for transactivation. Here we report the synthesis of a site-specific chemical ribonuclease based on the TAR binding domain of the HIV type 1 (HIV-1) tat. A peptide consisting of this 24-amino acid domain plus an additional C-terminal cysteine residue was chemically synthesized and covalently linked to 1,10-phenanthroline at the cysteine residue. The modified peptide binds to TAR sequences of both HIV-1 and HIV-2 and, in the presence of cupric ions and a reducing agent, cleaves these RNAs at specific sites. Cleavage sites on TAR sequences are consistent with peptide binding to the 3-nucleotide bulge, and the relative displacement of cleavage sites on the two strands suggests peptide binding to the major groove of the RNA. These results and existing evidence of the rapid cellular uptake of tat-derived peptides suggest that chemical nucleases based on tat may be useful for inactivating HIV mRNA in vivo. Images PMID:1565648

Protein-Protein Interaction Site Predictions with Three-Dimensional Probability Distributions of Interacting Atoms on Protein Surfaces

PubMed Central

Chen, Ching-Tai; Peng, Hung-Pin; Jian, Jhih-Wei; Tsai, Keng-Chang; Chang, Jeng-Yih; Yang, Ei-Wen; Chen, Jun-Bo; Ho, Shinn-Ying; Hsu, Wen-Lian; Yang, An-Suei

2012-01-01

Protein-protein interactions are key to many biological processes. Computational methodologies devised to predict protein-protein interaction (PPI) sites on protein surfaces are important tools in providing insights into the biological functions of proteins and in developing therapeutics targeting the protein-protein interaction sites. One of the general features of PPI sites is that the core regions from the two interacting protein surfaces are complementary to each other, similar to the interior of proteins in packing density and in the physicochemical nature of the amino acid composition. In this work, we simulated the physicochemical complementarities by constructing three-dimensional probability density maps of non-covalent interacting atoms on the protein surfaces. The interacting probabilities were derived from the interior of known structures. Machine learning algorithms were applied to learn the characteristic patterns of the probability density maps specific to the PPI sites. The trained predictors for PPI sites were cross-validated with the training cases (consisting of 432 proteins) and were tested on an independent dataset (consisting of 142 proteins). The residue-based Matthews correlation coefficient for the independent test set was 0.423; the accuracy, precision, sensitivity, specificity were 0.753, 0.519, 0.677, and 0.779 respectively. The benchmark results indicate that the optimized machine learning models are among the best predictors in identifying PPI sites on protein surfaces. In particular, the PPI site prediction accuracy increases with increasing size of the PPI site and with increasing hydrophobicity in amino acid composition of the PPI interface; the core interface regions are more likely to be recognized with high prediction confidence. The results indicate that the physicochemical complementarity patterns on protein surfaces are important determinants in PPIs, and a substantial portion of the PPI sites can be predicted correctly with the physicochemical complementarity features based on the non-covalent interaction data derived from protein interiors. PMID:22701576

Site-specific recombination in the chicken genome using Flipase recombinase-mediated cassette exchange.

PubMed

Lee, Hong Jo; Lee, Hyung Chul; Kim, Young Min; Hwang, Young Sun; Park, Young Hyun; Park, Tae Sub; Han, Jae Yong

2016-02-01

Targeted genome recombination has been applied in diverse research fields and has a wide range of possible applications. In particular, the discovery of specific loci in the genome that support robust and ubiquitous expression of integrated genes and the development of genome-editing technology have facilitated rapid advances in various scientific areas. In this study, we produced transgenic (TG) chickens that can induce recombinase-mediated gene cassette exchange (RMCE), one of the site-specific recombination technologies, and confirmed RMCE in TG chicken-derived cells. As a result, we established TG chicken lines that have, Flipase (Flp) recognition target (FRT) pairs in the chicken genome, mediated by piggyBac transposition. The transgene integration patterns were diverse in each TG chicken line, and the integration diversity resulted in diverse levels of expression of exogenous genes in each tissue of the TG chickens. In addition, the replaced gene cassette was expressed successfully and maintained by RMCE in the FRT predominant loci of TG chicken-derived cells. These results indicate that targeted genome recombination technology with RMCE could be adaptable to TG chicken models and that the technology would be applicable to specific gene regulation by cis-element insertion and customized expression of functional proteins at predicted levels without epigenetic influence. © FASEB.

Evolution of educational inequalities in site-specific cancer mortality among Belgian men between the 1990s and 2000s using a "fundamental cause" perspective.

PubMed

Vanthomme, Katrien; Vandenheede, Hadewijch; Hagedoorn, Paulien; Gadeyne, Sylvie

2017-07-05

According to the "fundamental cause" theory, emerging knowledge on health-enhancing behaviours and technologies results in health disparities. This study aims to assess (trends in) educational inequalities in site-specific cancer mortality in Belgian men in the 1990s and the 2000s using this framework. Data were derived from record linkage between the Belgian censuses of 1991 and 2001 and register data on mortality. The study population comprised all Belgian men aged 50-79 years during follow-up. Both absolute and relative inequality measures have been calculated. Despite an overall downward trend in cancer mortality, educational differences are observed for the majority of cancer sites in the 2000s. Generally, inequalities are largest for mortality from preventable cancers. Trends over time in inequalities are rather stable compared with the 1990s. Educational differences in site-specific cancer mortality persist in the 2000s in Belgium, mainly for cancers related to behavioural change and medical interventions. Policy efforts focussing on behavioural change and healthcare utilization remain crucial in order to tackle these increasing inequalities.

Genome Integration and Excision by a New Streptomyces Bacteriophage, ϕJoe

PubMed Central

Haley, Joshua A.; Stark, W. Marshall

2016-01-01

ABSTRACT Bacteriophages are the source of many valuable tools for molecular biology and genetic manipulation. In Streptomyces, most DNA cloning vectors are based on serine integrase site-specific DNA recombination systems derived from phage. Because of their efficiency and simplicity, serine integrases are also used for diverse synthetic biology applications. Here, we present the genome of a new Streptomyces phage, ϕJoe, and investigate the conditions for integration and excision of the ϕJoe genome. ϕJoe belongs to the largest Streptomyces phage cluster (R4-like) and encodes a serine integrase. The attB site from Streptomyces venezuelae was used efficiently by an integrating plasmid, pCMF92, constructed using the ϕJoe int-attP locus. The attB site for ϕJoe integrase was occupied in several Streptomyces genomes, including that of S. coelicolor, by a mobile element that varies in gene content and size between host species. Serine integrases require a phage-encoded recombination directionality factor (RDF) to activate the excision reaction. The ϕJoe RDF was identified, and its function was confirmed in vivo. Both the integrase and RDF were active in in vitro recombination assays. The ϕJoe site-specific recombination system is likely to be an important addition to the synthetic biology and genome engineering toolbox. IMPORTANCE Streptomyces spp. are prolific producers of secondary metabolites, including many clinically useful antibiotics. Bacteriophage-derived integrases are important tools for genetic engineering, as they enable integration of heterologous DNA into the Streptomyces chromosome with ease and high efficiency. Recently, researchers have been applying phage integrases for a variety of applications in synthetic biology, including rapid assembly of novel combinations of genes, biosensors, and biocomputing. An important requirement for optimal experimental design and predictability when using integrases, however, is the need for multiple enzymes with different specificities for their integration sites. In order to provide a broad platform of integrases, we identified and validated the integrase from a newly isolated Streptomyces phage, ϕJoe. ϕJoe integrase is active in vitro and in vivo. The specific recognition site for integration is present in a wide range of different actinobacteria, including Streptomyces venezuelae, an emerging model bacterium in Streptomyces research. PMID:28003200

Localization of premature ventricular contractions from the papillary muscles using the standard 12-lead electrocardiogram: a feasibility study using a novel cardiac isochrone positioning system.

PubMed

van Dam, Peter M; Boyle, Noel G; Laks, Michael M; Tung, Roderick

2016-12-01

The precise localization of the site of origin of a premature ventricular contraction (PVC) prior to ablation can facilitate the planning and execution of the electrophysiological procedure. In clinical practice, the targeted ablation site is estimated from the standard 12-lead ECG. The accuracy of this qualitative estimation has limitations, particularly in the localization of PVCs originating from the papillary muscles. Clinical available electrocardiographic imaging (ECGi) techniques that incorporate patient-specific anatomy may improve the localization of these PVCs, but require body surface maps with greater specificity for the epicardium. The purpose of this report is to demonstrate that a novel cardiac isochrone positioning system (CIPS) program can accurately detect the specific location of the PVC on the papillary muscle using only a 12-lead ECG. Cardiac isochrone positioning system uses three components: (i) endocardial and epicardial cardiac anatomy and torso geometry derived from MRI, (ii) the patient-specific electrode positions derived from an MRI model registered 3D image, and (iii) the 12-lead ECG. CIPS localizes the PVC origin by matching the anatomical isochrone vector with the ECG vector. The predicted PVC origin was compared with the site of successful ablation or stimulation. Three patients who underwent electrophysiological mapping and ablation of PVCs originating from the papillary muscles were studied. CIPS localized the PVC origin for all three patients to the correct papillary muscle and specifically to the base, mid, or apical region. A simplified form of ECGi utilizing only 12 standard electrocardiographic leads may facilitate accurate localization of the origin of papillary muscle PVCs. Published on behalf of the European Society of Cardiology. All rights reserved. © The Author 2016. For Permissions, please email: journals.permissions@oup.com.

Recruiting Human Microbiome Shotgun Data to Site-Specific Reference Genomes

PubMed Central

Xie, Gary; Lo, Chien-Chi; Scholz, Matthew; Chain, Patrick S. G.

2014-01-01

The human body consists of innumerable multifaceted environments that predispose colonization by a number of distinct microbial communities, which play fundamental roles in human health and disease. In addition to community surveys and shotgun metagenomes that seek to explore the composition and diversity of these microbiomes, there are significant efforts to sequence reference microbial genomes from many body sites of healthy adults. To illustrate the utility of reference genomes when studying more complex metagenomes, we present a reference-based analysis of sequence reads generated from 55 shotgun metagenomes, selected from 5 major body sites, including 16 sub-sites. Interestingly, between 13% and 92% (62.3% average) of these shotgun reads were aligned to a then-complete list of 2780 reference genomes, including 1583 references for the human microbiome. However, no reference genome was universally found in all body sites. For any given metagenome, the body site-specific reference genomes, derived from the same body site as the sample, accounted for an average of 58.8% of the mapped reads. While different body sites did differ in abundant genera, proximal or symmetrical body sites were found to be most similar to one another. The extent of variation observed, both between individuals sampled within the same microenvironment, or at the same site within the same individual over time, calls into question comparative studies across individuals even if sampled at the same body site. This study illustrates the high utility of reference genomes and the need for further site-specific reference microbial genome sequencing, even within the already well-sampled human microbiome. PMID:24454771

Unipolar Endocardial Voltage Mapping in the Right Ventricle: Optimal Cutoff Values Correcting for Computed Tomography-Derived Epicardial Fat Thickness and Their Clinical Value for Substrate Delineation.

PubMed

Venlet, Jeroen; Piers, Sebastiaan R D; Kapel, Gijsbert F L; de Riva, Marta; Pauli, Philippe F G; van der Geest, Rob J; Zeppenfeld, Katja

2017-08-01

Low endocardial unipolar voltage (UV) at sites with normal bipolar voltage (BV) may indicate epicardial scar. Currently applied UV cutoff values are based on studies that lacked epicardial fat information. This study aimed to define endocardial UV cutoff values using computed tomography-derived fat information and to analyze their clinical value for right ventricular substrate delineation. Thirty-three patients (50±14 years; 79% men) underwent combined endocardial-epicardial right ventricular electroanatomical mapping and ablation of right ventricular scar-related ventricular tachycardia with computed tomographic image integration, including computed tomography-derived fat thickness. Of 6889 endocardial-epicardial mapping point pairs, 547 (8%) pairs with distance <10 mm and fat thickness <1.0 mm were analyzed for voltage and abnormal (fragmented/late potential) electrogram characteristics. At sites with endocardial BV >1.50 mV, the optimal endocardial UV cutoff for identification of epicardial BV <1.50 mV was 3.9 mV (area under the curve, 0.75; sensitivity, 60%; specificity, 79%) and cutoff for identification of abnormal epicardial electrogram was 3.7 mV (area under the curve, 0.88; sensitivity, 100%; specificity, 67%). The majority of abnormal electrograms (130 of 151) were associated with transmural scar. Eighty-six percent of abnormal epicardial electrograms had corresponding endocardial sites with BV <1.50 mV, and the remaining could be identified by corresponding low endocardial UV <3.7 mV. For identification of epicardial right ventricular scar, an endocardial UV cutoff value of 3.9 mV is more accurate than previously reported cutoff values. Although the majority of epicardial abnormal electrograms are associated with transmural scar with low endocardial BV, the additional use of endocardial UV at normal BV sites improves the diagnostic accuracy resulting in identification of all epicardial abnormal electrograms at sites with <1.0 mm fat. © 2017 American Heart Association, Inc.

Hierarchical porous carbon materials derived from petroleum pitch for high-performance supercapacitors

NASA Astrophysics Data System (ADS)

Abudu, Patiman; Wang, Luxiang; Xu, Mengjiao; Jia, Dianzeng; Wang, Xingchao; Jia, Lixia

2018-06-01

In this work, a honeycomb-like carbon material derived from petroleum pitch was synthesized by a simple one-step carbonization/activation method using silica nanospheres as the hard templates. The obtained hierarchical porous carbon materials (HPCs) with a large specific surface area and uniform macropore distribution provide abundant active sites and sufficient ion migration channels. When used as an electrode material for supercapacitors, the HPCs exhibit a high specific capacitance of 341.0 F g-1 at 1 A g-1, excellent rate capability with a capacitance retention of 55.6% at 50 A g-1 (189.5 F g-1), and outstanding cycling performance in the three-electrode system.

Amino acid derivatives of 5-ASA as novel prodrugs for intestinal drug delivery.

PubMed

Clerici, C; Gentili, G; Boschetti, E; Santucci, C; Aburbeh, A G; Natalini, B; Pellicciari, R; Morelli, A

1994-12-01

In an attempt to obtain site-specific delivery of 5-ASA in the intestinal tract, we have determined the extent of absorption and metabolism of a number of novel 5-ASA derivatives, namely, (N-L-glutamyl)-amino-2-salicylic acid (1), (N-L-aspartyl)-amino-2-salicylic-acid (2), 5-aminosalicyl-L-proline-L-leucine (3), and 5-(N-L-glutamyl)-aminosalicyl-L-proline-L-leucine (4), which are selectively cleaved by intestinal brush border aminopeptidase A and carboxypeptidases. These novel prodrugs, 5-ASA, and sulfasalazine were administered to adult Fisher rats (N = 30) and to animals that had undergone prior colostomy (N = 30). Urine and feces were collected at timed intervals for 48 hr and the metabolites, 5-ASA, and N-acetyl-5-ASA were measured by high-performance liquid chromatography. The absorption and metabolism of all compounds were essentially identical in colostomized and normal animals. 5-ASA exhibited a rapid proximal intestinal absorption as evidenced by the high cumulative urinary excretion (> 65%) and low fecal excretion. Sulfasalazine, as expected, exhibited a lower urinary recovery (< 35%) and higher fecal excretion of 5-ASA and its metabolite. The novel glutamate and aspartate derivatives (1 and 2) behaved similarly to sulfasalazine, while administration of the proline-leucine derivative (3) resulted in urinary and fecal recovery values intermediate with respect to those observed with 5-ASA and sulfasalazine. 5-(N-L-Glutamyl)-aminosalicyl-L-proline-L-leucine yielded the highest fecal recovery of 5-ASA and its N-acetyl derivative, indicating a more efficient delivery to the distal bowel. Amino acid derivatives of 5-ASA appear to be potentially useful prodrugs for the site-specific delivery of 5-ASA to different regions of the intestinal tract.

DESIGN OF AQUIFER REMEDIATION SYSTEMS: (2) Estimating site-specific performance and benefits of partial source removal

EPA Science Inventory

A Lagrangian stochastic model is proposed as a tool that can be utilized in forecasting remedial performance and estimating the benefits (in terms of flux and mass reduction) derived from a source zone remedial effort. The stochastic functional relationships that describe the hyd...

ACCURACY OF THE 1992 NATIONAL LAND COVER DATASET AREA ESTIMATES: AN ANALYSIS AT MULTIPLE SPATIAL EXTENTS

EPA Science Inventory

Abstract for poster presentation:

Site-specific accuracy assessments evaluate fine-scale accuracy of land-use/land-cover(LULC) datasets but provide little insight into accuracy of area estimates of LULC

classes derived from sampling units of varying size. Additiona...

pH-sensitive inulin-based nanomicelles for intestinal site-specific and controlled release of celecoxib.

PubMed

Mandracchia, Delia; Trapani, Adriana; Perteghella, Sara; Sorrenti, Milena; Catenacci, Laura; Torre, Maria Luisa; Trapani, Giuseppe; Tripodo, Giuseppe

2018-02-01

Aiming at a site-specific drug release in the lower intestinal tract, this paper deals with the synthesis and physicochemical/biological characterization of pH-sensitive nanomicelles from an inulin (INU) amphiphilic derivative. To allow an intestinal site specific release of the payload, INU-Vitamin E (INVITE) bioconjugates were functionalized with succinic anhydride to provide the system with pH-sensitive groups preventing a premature release of the payload into the stomach. The obtained INVITESA micelles resulted nanosized, with a low critical aggregation concentration and the release studies showed a marked pH-dependent release. The drug loading stabilized the micelles against the acidic hydrolysis. From transport studies on Caco-2 cells, resulted that INVITESA nanomicelles cross the cellular monolayer but are actively re-transported in the secretory (basolateral-apical) direction when loaded in apical side. It suggests that the entrapped drug could not be absorbed before the release from the micelles, enabling so a local release of the active. Copyright © 2017 Elsevier Ltd. All rights reserved.

Prefusion F-specific antibodies determine the magnitude of RSV neutralizing activity in human sera.

PubMed

Ngwuta, Joan O; Chen, Man; Modjarrad, Kayvon; Joyce, M Gordon; Kanekiyo, Masaru; Kumar, Azad; Yassine, Hadi M; Moin, Syed M; Killikelly, April M; Chuang, Gwo-Yu; Druz, Aliaksandr; Georgiev, Ivelin S; Rundlet, Emily J; Sastry, Mallika; Stewart-Jones, Guillaume B E; Yang, Yongping; Zhang, Baoshan; Nason, Martha C; Capella, Cristina; Peeples, Mark E; Ledgerwood, Julie E; McLellan, Jason S; Kwong, Peter D; Graham, Barney S

2015-10-14

Respiratory syncytial virus (RSV) is estimated to claim more lives among infants <1 year old than any other single pathogen, except malaria, and poses a substantial global health burden. Viral entry is mediated by a type I fusion glycoprotein (F) that transitions from a metastable prefusion (pre-F) to a stable postfusion (post-F) trimer. A highly neutralization-sensitive epitope, antigenic site Ø, is found only on pre-F. We determined what fraction of neutralizing (NT) activity in human sera is dependent on antibodies specific for antigenic site Ø or other antigenic sites on F in healthy subjects from ages 7 to 93 years. Adsorption of individual sera with stabilized pre-F protein removed >90% of NT activity and depleted binding antibodies to both F conformations. In contrast, adsorption with post-F removed ~30% of NT activity, and binding antibodies to pre-F were retained. These findings were consistent across all age groups. Protein competition neutralization assays with pre-F mutants in which sites Ø or II were altered to knock out binding of antibodies to the corresponding sites showed that these sites accounted for ~35 and <10% of NT activity, respectively. Binding competition assays with monoclonal antibodies (mAbs) indicated that the amount of site Ø-specific antibodies correlated with NT activity, whereas the magnitude of binding competed by site II mAbs did not correlate with neutralization. Our results indicate that RSV NT activity in human sera is primarily derived from pre-F-specific antibodies, and therefore, inducing or boosting NT activity by vaccination will be facilitated by using pre-F antigens that preserve site Ø. Copyright © 2015, American Association for the Advancement of Science.

FunShift: a database of function shift analysis on protein subfamilies

PubMed Central

Abhiman, Saraswathi; Sonnhammer, Erik L. L.

2005-01-01

Members of a protein family normally have a general biochemical function in common, but frequently one or more subgroups have evolved a slightly different function, such as different substrate specificity. It is important to detect such function shifts for a more accurate functional annotation. The FunShift database described here is a compilation of function shift analysis performed between subfamilies in protein families. It consists of two main components: (i) subfamilies derived from protein domain families and (ii) pairwise subfamily comparisons analyzed for function shift. The present release, FunShift 12, was derived from Pfam 12 and consists of 151 934 subfamilies derived from 7300 families. We carried out function shift analysis by two complementary methods on families with up to 500 members. From a total of 179 210 subfamily pairs, 62 384 were predicted to be functionally shifted in 2881 families. Each subfamily pair is provided with a markup of probable functional specificity-determining sites. Tools for searching and exploring the data are provided to make this database a valuable resource for protein function annotation. Knowledge of these functionally important sites will be useful for experimental biologists performing functional mutation studies. FunShift is available at http://FunShift.cgb.ki.se. PMID:15608176

Investigation of the mechanism of meiotic DNA cleavage by VMA1-derived endonuclease uncovers a meiotic alteration in chromatin structure around the target site.

PubMed

Fukuda, Tomoyuki; Ohta, Kunihiro; Ohya, Yoshikazu

2006-06-01

VMA1-derived endonuclease (VDE), a homing endonuclease in Saccharomyces cerevisiae, is encoded by the mobile intein-coding sequence within the nuclear VMA1 gene. VDE recognizes and cleaves DNA at the 31-bp VDE recognition sequence (VRS) in the VMA1 gene lacking the intein-coding sequence during meiosis to insert a copy of the intein-coding sequence at the cleaved site. The mechanism underlying the meiosis specificity of VMA1 intein-coding sequence homing remains unclear. We studied various factors that might influence the cleavage activity in vivo and found that VDE binding to the VRS can be detected only when DNA cleavage by VDE takes place, implying that meiosis-specific DNA cleavage is regulated by the accessibility of VDE to its target site. As a possible candidate for the determinant of this accessibility, we analyzed chromatin structure around the VRS and revealed that local chromatin structure near the VRS is altered during meiosis. Although the meiotic chromatin alteration exhibits correlations with DNA binding and cleavage by VDE at the VMA1 locus, such a chromatin alteration is not necessarily observed when the VRS is embedded in ectopic gene loci. This suggests that nucleosome positioning or occupancy around the VRS by itself is not the sole mechanism for the regulation of meiosis-specific DNA cleavage by VDE and that other mechanisms are involved in the regulation.

Investigation of the Mechanism of Meiotic DNA Cleavage by VMA1-Derived Endonuclease Uncovers a Meiotic Alteration in Chromatin Structure around the Target Site

PubMed Central

Fukuda, Tomoyuki; Ohta, Kunihiro; Ohya, Yoshikazu

2006-01-01

VMA1-derived endonuclease (VDE), a homing endonuclease in Saccharomyces cerevisiae, is encoded by the mobile intein-coding sequence within the nuclear VMA1 gene. VDE recognizes and cleaves DNA at the 31-bp VDE recognition sequence (VRS) in the VMA1 gene lacking the intein-coding sequence during meiosis to insert a copy of the intein-coding sequence at the cleaved site. The mechanism underlying the meiosis specificity of VMA1 intein-coding sequence homing remains unclear. We studied various factors that might influence the cleavage activity in vivo and found that VDE binding to the VRS can be detected only when DNA cleavage by VDE takes place, implying that meiosis-specific DNA cleavage is regulated by the accessibility of VDE to its target site. As a possible candidate for the determinant of this accessibility, we analyzed chromatin structure around the VRS and revealed that local chromatin structure near the VRS is altered during meiosis. Although the meiotic chromatin alteration exhibits correlations with DNA binding and cleavage by VDE at the VMA1 locus, such a chromatin alteration is not necessarily observed when the VRS is embedded in ectopic gene loci. This suggests that nucleosome positioning or occupancy around the VRS by itself is not the sole mechanism for the regulation of meiosis-specific DNA cleavage by VDE and that other mechanisms are involved in the regulation. PMID:16757746

VITAL NMR: Using Chemical Shift Derived Secondary Structure Information for a Limited Set of Amino Acids to Assess Homology Model Accuracy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Brothers, Michael C; Nesbitt, Anna E; Hallock, Michael J

2011-01-01

Homology modeling is a powerful tool for predicting protein structures, whose success depends on obtaining a reasonable alignment between a given structural template and the protein sequence being analyzed. In order to leverage greater predictive power for proteins with few structural templates, we have developed a method to rank homology models based upon their compliance to secondary structure derived from experimental solid-state NMR (SSNMR) data. Such data is obtainable in a rapid manner by simple SSNMR experiments (e.g., (13)C-(13)C 2D correlation spectra). To test our homology model scoring procedure for various amino acid labeling schemes, we generated a library ofmore » 7,474 homology models for 22 protein targets culled from the TALOS+/SPARTA+ training set of protein structures. Using subsets of amino acids that are plausibly assigned by SSNMR, we discovered that pairs of the residues Val, Ile, Thr, Ala and Leu (VITAL) emulate an ideal dataset where all residues are site specifically assigned. Scoring the models with a predicted VITAL site-specific dataset and calculating secondary structure with the Chemical Shift Index resulted in a Pearson correlation coefficient (-0.75) commensurate to the control (-0.77), where secondary structure was scored site specifically for all amino acids (ALL 20) using STRIDE. This method promises to accelerate structure procurement by SSNMR for proteins with unknown folds through guiding the selection of remotely homologous protein templates and assessing model quality.« less

Epigenetic control of alternative mRNA processing at the imprinted Herc3/Nap1l5 locus

PubMed Central

Cowley, Michael; Wood, Andrew J.; Böhm, Sabrina; Schulz, Reiner; Oakey, Rebecca J.

2012-01-01

Alternative polyadenylation increases transcriptome diversity by generating multiple transcript isoforms from a single gene. It is thought that this process can be subject to epigenetic regulation, but few specific examples of this have been reported. We previously showed that the Mcts2/H13 locus is subject to genomic imprinting and that alternative polyadenylation of H13 transcripts occurs in an allele-specific manner, regulated by epigenetic mechanisms. Here, we demonstrate that allele-specific polyadenylation occurs at another imprinted locus with similar features. Nap1l5 is a retrogene expressed from the paternally inherited allele, is situated within an intron of a ‘host’ gene Herc3, and overlaps a CpG island that is differentially methylated between the parental alleles. In mouse brain, internal Herc3 polyadenylation sites upstream of Nap1l5 are used on the paternally derived chromosome, from which Nap1l5 is expressed, whereas a downstream site is used more frequently on the maternally derived chromosome. Ablating DNA methylation on the maternal allele at the Nap1l5 promoter increases the use of an internal Herc3 polyadenylation site and alters exon splicing. These changes demonstrate the influence of epigenetic mechanisms in regulating Herc3 alternative mRNA processing. Internal Herc3 polyadenylation correlates with expression levels of Nap1l5, suggesting a possible role for transcriptional interference. Similar mechanisms may regulate alternative polyadenylation elsewhere in the genome. PMID:22790983

Activity-Based Probes for Isoenzyme- and Site-Specific Functional Characterization of Glutathione S -Transferases

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stoddard, Ethan G.; Killinger, Bryan J.; Nair, Reji N.

Glutathione S-transferases (GSTs) comprise a highly diverse family of phase II drug metabolizing enzymes whose shared function is the conjugation of reduced glutathione to various endo- and xenobiotics. Although the conglomerate activity of these enzymes can be measured by colorimetric assays, measurement of the individual contribution from specific isoforms and their contribution to the detoxification of xenobiotics in complex biological samples has not been possible. For this reason, we have developed two activity-based probes that characterize active glutathione transferases in mammalian tissues. The GST active site is comprised of a glutathione binding “G site” and a distinct substrate binding “Hmore » site”. Therefore, we developed (1) a glutathione-based photoaffinity probe (GSH-ABP) to target the “G site”, and (2) a probe designed to mimic a substrate molecule and show “H site” activity (GST-ABP). The GSH-ABP features a photoreactive moiety for UV-induced covalent binding to GSTs and glutathione-binding enzymes. The GST-ABP is a derivative of a known mechanism-based GST inhibitor that binds within the active site and inhibits GST activity. Validation of probe targets and “G” and “H” site specificity was carried out using a series of competitors in liver homogenates. Herein, we present robust tools for the novel characterization of enzyme- and active site-specific GST activity in mammalian model systems.« less

Predicting risk of trace element pollution from municipal roads using site-specific soil samples and remotely sensed data.

PubMed

Reeves, Mari Kathryn; Perdue, Margaret; Munk, Lee Ann; Hagedorn, Birgit

2018-07-15

Studies of environmental processes exhibit spatial variation within data sets. The ability to derive predictions of risk from field data is a critical path forward in understanding the data and applying the information to land and resource management. Thanks to recent advances in predictive modeling, open source software, and computing, the power to do this is within grasp. This article provides an example of how we predicted relative trace element pollution risk from roads across a region by combining site specific trace element data in soils with regional land cover and planning information in a predictive model framework. In the Kenai Peninsula of Alaska, we sampled 36 sites (191 soil samples) adjacent to roads for trace elements. We then combined this site specific data with freely-available land cover and urban planning data to derive a predictive model of landscape scale environmental risk. We used six different model algorithms to analyze the dataset, comparing these in terms of their predictive abilities and the variables identified as important. Based on comparable predictive abilities (mean R 2 from 30 to 35% and mean root mean square error from 65 to 68%), we averaged all six model outputs to predict relative levels of trace element deposition in soils-given the road surface, traffic volume, sample distance from the road, land cover category, and impervious surface percentage. Mapped predictions of environmental risk from toxic trace element pollution can show land managers and transportation planners where to prioritize road renewal or maintenance by each road segment's relative environmental and human health risk. Published by Elsevier B.V.

Site-occupation embedding theory using Bethe ansatz local density approximations

NASA Astrophysics Data System (ADS)

Senjean, Bruno; Nakatani, Naoki; Tsuchiizu, Masahisa; Fromager, Emmanuel

2018-06-01

Site-occupation embedding theory (SOET) is an alternative formulation of density functional theory (DFT) for model Hamiltonians where the fully interacting Hubbard problem is mapped, in principle exactly, onto an impurity-interacting (rather than a noninteracting) one. It provides a rigorous framework for combining wave-function (or Green function)-based methods with DFT. In this work, exact expressions for the per-site energy and double occupation of the uniform Hubbard model are derived in the context of SOET. As readily seen from these derivations, the so-called bath contribution to the per-site correlation energy is, in addition to the latter, the key density functional quantity to model in SOET. Various approximations based on Bethe ansatz and perturbative solutions to the Hubbard and single-impurity Anderson models are constructed and tested on a one-dimensional ring. The self-consistent calculation of the embedded impurity wave function has been performed with the density-matrix renormalization group method. It has been shown that promising results are obtained in specific regimes of correlation and density. Possible further developments have been proposed in order to provide reliable embedding functionals and potentials.

The uncertainty of biodegradation rate constants of emerging organic compounds in soil and groundwater - A compilation of literature values for 82 substances.

PubMed

Greskowiak, Janek; Hamann, Enrico; Burke, Victoria; Massmann, Gudrun

2017-12-01

The present study reports on biodegradation rate constants of emerging organic compounds (EOCs) in soil and groundwater available in the literature. The major aim of this compilation was to provide an assessment of the uncertainty of hydrological models with respect to the fate of EOCs. The literature search identified a total number of 82 EOCs for which 1st-order rate constants could be derived. It was found that for the majority of compounds degradation rate constants vary over more than three orders of magnitude. Correlation to factors that are well known to affect the degradation rate, such as temperature or redox condition was weak. No correlation at all was found with results from available quantitative structure-activity relationship models. This suggests that many unknown site specific or experimentally specific factors influence the degradation behavior of EOCs in the environment. Thus, local and catchment scale predictive models to estimate EOC concentration at receptors, e.g., receiving waters or drinking water wells, need to consider the large uncertainty in 1st-order rate constants. As a consequence, applying rate constants that were derived from one experiment or field site investigation to other experiments or field sites should be done with extreme caution. Copyright © 2017 Elsevier Ltd. All rights reserved.

Transcript Isoform Variation Associated with Cytosine Modification in Human Lymphoblastoid Cell Lines.

PubMed

Zhang, Xu; Zhang, Wei

2016-06-01

Cytosine modification on DNA is variable among individuals, which could correlate with gene expression variation. The effect of cytosine modification on interindividual transcript isoform variation (TIV), however, remains unclear. In this study, we assessed the extent of cytosine modification-specific TIV in lymphoblastoid cell lines (LCLs) derived from unrelated individuals of European and African descent. Our study detected cytosine modification-specific TIVs for 17% of the analyzed genes at a 5% false discovery rate. Forty-five percent of the TIV-associated cytosine modifications correlated with the overall gene expression levels as well, with the corresponding CpG sites overrepresented in transcript initiation sites, transcription factor binding sites, and distinct histone modification peaks, suggesting that alternative isoform transcription underlies the TIVs. Our analysis also revealed 33% of the TIV-associated cytosine modifications that affected specific exons, with the corresponding CpG sites overrepresented in exon/intron junctions, splicing branching points, and transcript termination sites, implying that the TIVs are attributable to alternative splicing or transcription termination. Genetic and epigenetic regulation of TIV shared target preference but exerted independent effects on 61% of the common exon targets. Cytosine modification-specific TIVs detected from LCLs were differentially enriched in those detected from various tissues in The Cancer Genome Atlas, indicating their developmental dependency. Genes containing cytosine modification-specific TIVs were enriched in pathways of cancers and metabolic disorders. Our study demonstrated a prominent effect of cytosine modification variation on the transcript isoform spectrum over gross transcript abundance and revealed epigenetic contributions to diseases that were mediated through cytosine modification-specific TIV. Copyright © 2016 by the Genetics Society of America.

Exploring molecular insights into the interaction mechanism of cholesterol derivatives with the Mce4A: A combined spectroscopic and molecular dynamic simulation studies.

PubMed

Khan, Shagufta; Khan, Faez Iqbal; Mohammad, Taj; Khan, Parvez; Hasan, Gulam Mustafa; Lobb, Kevin A; Islam, Asimul; Ahmad, Faizan; Imtaiyaz Hassan, Md

2018-05-01

Mammalian cell entry protein (Mce4A) is a member of MCE-family, and is being considered as a potential drug target of Mycobacterium tuberculosis infection because it is required for invasion and latent survival of pathogen by utilizing host's cholesterol. In the present study, we performed molecular docking followed by 100 ns MD simulation studies to understand the mechanism of interaction of Mce4A to the cholesterol derivatives and probucol. The selected ligands, cholesterol, 25-hydroxycholesterol, 5-cholesten-3β-ol-7-one and probucol bind to the predicted active site cavity of Mce4A, and complexes remain stable during entire simulation of 100 ns. In silico studies were further validated by fluorescence-binding studies to calculate actual binding affinity and number of binding site(s). The non-toxicity of all ligands was confirmed on human monocytic cell (THP1) by MTT assay. This work provides a deeper insight into the mechanism of interaction of Mce4A to cholesterol derivatives, which may be further exploited to design potential and specific inhibitors to ameliorate the Mycobacterium pathogenesis. Copyright © 2018 Elsevier B.V. All rights reserved.

SITE SPECIFIC REFERENCE PERSON PARAMETERS AND DERIVED CONCENTRATION STANDARDS FOR THE SAVANNAH RIVER SITE

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jannik, T.

2013-03-14

The purpose of this report is twofold. The first is to develop a set of behavioral parameters for a reference person specific for the Savannah River Site (SRS) such that the parameters can be used to determine dose to members of the public in compliance with Department of Energy (DOE) Order 458.1 “Radiation Protection of the Public and the Environment.” A reference person is a hypothetical, gender and age aggregation of human physical and physiological characteristics arrived at by international consensus for the purpose of standardizing radiation dose calculations. DOE O 458.1 states that compliance with the annual dose limitmore » of 100 mrem (1 mSv) to a member of the public may be demonstrated by calculating the dose to the maximally exposed individual (MEI) or to a representative person. Historically, for dose compliance, SRS has used the MEI concept, which uses adult dose coefficients and adult male usage parameters. Beginning with the 2012 annual site environmental report, SRS will be using the representative person concept for dose compliance. The dose to a representative person will be based on 1) the SRS-specific reference person usage parameters at the 95th percentile of appropriate national or regional data, which are documented in this report, 2) the reference person (gender and age averaged) ingestion and inhalation dose coefficients provided in DOE Derived Concentration Technical Standard (DOE-STD-1196-2011), and 3) the external dose coefficients provided in the DC_PAK3 toolbox. The second purpose of this report is to develop SRS-specific derived concentration standards (DCSs) for all applicable food ingestion pathways, ground shine, and water submersion. The DCS is the concentration of a particular radionuclide in water, in air, or on the ground that results in a member of the public receiving 100 mrem (1 mSv) effective dose following continuous exposure for one year. In DOE-STD-1196-2011, DCSs were developed for the ingestion of water, inhalation of air and submersion in air pathways, only. These DCSs are required by DOE O 458.1 to be used at all DOE sites in the design and conduct of radiological environmental protection programs. In this report, DCSs for the following additional pathways were considered and documented: ingestion of meat, dairy, grains, produce (fruits and vegetables), seafood, submersion in water and ground shine. These additional DCSs were developed using the same methods as in DOE-STD-1196-2011 and will be used at SRS, where appropriate, as screening and reference values.« less

Site-specific chemical modification of antibody fragments using traceless cleavable linkers.

PubMed

Bernardes, Gonçalo J L; Steiner, Martina; Hartmann, Isabelle; Neri, Dario; Casi, Giulio

2013-11-01

Antibody-drug conjugates (ADCs) are promising agents for the selective delivery of cytotoxic drugs to specific cells (for example, tumors). In this protocol, we describe two strategies for the precise modification at engineered C- or N-terminal cysteines of antibodies in IgG, diabody and small immunoprotein (SIP) formats that yield homogenous ADCs. In this protocol, cemadotin derivatives are used as model drugs, as these agents have a potent cytotoxic activity and are easy to synthesize. However, other drugs with similar functional groups could be considered. In the first approach, a cemadotin derivative containing a sulfhydryl group results in a mixed disulfide linkage. In the second approach, a cemadotin derivative containing an aldehyde group is joined via a thiazolidine linkage. The procedures outlined are robust, enabling the preparation of ADCs with a defined number of drugs per antibody in a time frame between 7 and 24 h.

Evaluation of toxicity to the amphipod, Hyalella azteca, and to the midge, Chironomus dilutus; and bioaccumulation by the oligochaete, Lumbriculus variegatus, with exposure to PCB-contaminated sediments from Anniston, Alabama

USGS Publications Warehouse

Ingersoll, Christopher G.; Steevens, Jeffery A.; MacDonald, Donald D.; Brumbaugh, William G.; Coady, Matthew R.; Farrar, J. Daniel; Lotufo, Guilherme R.; Kemble, Nile E.; Kunz, James L.; Stanley, Jacob K.; Sinclair, Jesse A.; Ingersoll, Christopher G.; Steevens, Jeffery A.; MacDonald, Donald D.

2014-01-01

The U.S. Environmental Protection Agency (USEPA) requested that as part of the remedial investigation for the Anniston, Alabama Polychlorinated Biphenyl (PCB) Site (Anniston PCB Site), that Pharmacia Corporation and Solutia Inc. (P/S) perform long-term reproduction toxicity tests with the amphipod, Hyalella azteca, and the midge, Chironomus dilutus, and bioaccumulation tests with the oligochaete, Lumbriculus variegatus, using sediment samples collected from reference locations and from Operable Unit 4 of the Anniston PCB Site. The sediment toxicity testing and sediment bioaccumulation results will be used by ARCADIS U.S., Inc. (ARCADIS) as part of a weight-of-evidence assessment to evaluate risks and establish sediment remediation goals for contaminants to sediment-dwelling organisms inhabiting the Anniston PCB Site. The goal of this study was to characterize relations between sediment chemistry and sediment toxicity and relations between sediment chemistry and sediment bioaccumulation in samples of sediments collected from the Anniston PCB Site. A total of 32 samples were evaluated from six test sites and one reference site to provide a wide range in concentrations of chemicals of potential concern (COPCs) including PCBs in samples of whole sediment. The goal of this study was not to determine the extent of sediment contamination across the Anniston PCB Site. Hence, the test sites or samples collected from within a test site were not selected to represent the spatial extent of sediment contamination across the Anniston PCB Site. Sediment chemistry, pore-water chemistry, and sediment toxicity data were generated for 26 sediment samples from the Anniston PCB Site. All of the samples were evaluated to determine if they qualified as reference sediment samples. Those samples that met the chemical selection criteria and biological selection criteria were identified as reference samples and used to develop the reference envelope for each toxicity test endpoint. Physical characterization of samples of whole sediment included analyses of grain size, TOC, and nutrients. Organic chemical characterization of samples of whole sediment included PCB homologs and select (13) PCB congeners, parent and alkylated polycyclic aromatic hydrocarbons (PAHs), organochlorine pesticides, and polychlorinated dibenzo-p-dioxins; and dibenzofurans. The PCB aroclors analyzed included 1016, 1221, 1232, 1242, 1248, 1254, 1260, 1262 and 1268. Analyses of whole sediment also included total metals, simultaneously extracted metals, and acid volatile sulfide. Chemical characterization of samples of pore water isolated from samples of whole sediment at the start of the sediment toxicity exposures or at the start of the sediment bioaccumulation exposures included metals, major cations, major anions, dissolved organic carbon, and additional water-quality characteristics. Concentrations of metals or PCBs in pore water during the sediment toxicity exposures or during sediment bioaccumulation exposures also were determined using peeper samples (for metals) or solid-phase microextraction (SPME) samplers (for PCBs). The bioavailability and bioaccumulation of PCBs in 14 sediment samples were investigated using SPME passive samplers and the 28-d L. variegatus whole-sediment bioaccumulation exposures In general the accumulation of PCBs consistently was predicted through the use of organic carbon normalization and equilibrium partitioning. In these sediments, PCB homologs were accumulated differently based on bioavailability and potential to accumulate in oligochaetes. As part of this assessment homolog specific biota sediment accumulation factor values were developed that could be applied across the larger site to predict tissue levels of PCBs. The whole-sediment toxicity tests done with H. azteca and C. dilutus met the established ASTM and USEPA test acceptability criteria. The most responsive H. azteca endpoints were day 42 survival normalized young per female and day 28 biomass and that the most responsive C. dilutus endpoints were adult biomass and percent adult emergence. Overall, between the two species, the most responsive endpoint assessed for these two species was H. azteca survival-normalized young per female (67 percent of the samples classified as toxic). Concentration-response models (CRMs) and site-specific sediment toxicity thresholds (TTs) were generated with matching sediment chemistry and sediment toxicity data. Sediment chemistry, pore-water chemistry, and sediment toxicity data were evaluated for as many as 26 sediment samples from the Anniston PCB Site. The reference-envelope approach was used to identify the sediment samples that were toxic to benthic invertebrates. This procedure involved identification of reference sediment samples, normalizing the toxicity data to reflect control responses, developing a reference envelope for each toxicity test endpoint, and designating each sediment sample as toxic or not toxic for each toxicity test endpoint, for each species, and for all species combined. These results demonstrated percent emergence of adult C. dilutus, biomass of adult C. dilutus, and reproduction of H. azteca normalized to percent survival were among the most responsive endpoints that were evaluated. Therefore, these endpoints were selected for CRM development. The site-specific TTs for whole sediment provide a reliable basis for identifying toxic and not toxic sediment samples in the Anniston PCB Site (that is, for correctly classifying the sediment samples used to derive the TTs as toxic or not toxic, for the endpoint used to derive the TTs). Among the 69 TTs for sediment, the TTLRs for total PCB homologs [499 to 1,870 micrograms per kilogram dry weight (μg/kg DW)] and for lead [(9.48 to 10.3 milligrams per kilogram (mg/kg) DW] based on reproduction of H. azteca or based on emergence or biomass of adult C. dilutus, were the most reliable. Such TTs had low rates of false negative errors (that is, only 0 to 11 percent of the samples below the TT were toxic to benthic invertebrates), low rates of false positive errors (only 0 to 6 percent of the samples greater than the TT were not toxic to benthic invertebrates), and high rates of correct classification (that is, 92 to 96 percent). The site-specific TTs for PCBs and other COPCs derived in this study also were compared to empirically based sediment quality guidelines (SQGs), to equilibrium-partitioning based SQGs, and to the results of spiked-sediment toxicity tests. The results of this evaluation indicated that the site-specific sediment TTs for PCBs were comparable to the consensus-based SQGs that were derived for PCBs. In addition, the site-specific sediment TTs for PCBs are well within the range of SQGs derived using the equilibrium partitioning approach. The site-specific sediment TTs for PCBs also are consistent with the results of chronic TTs that have been estimated for benthic invertebrates using the results of spiked-sediment toxicity tests. As the site-specific sediment TTs for PCBs are consistent with empirically based SQGs, equilibrium-partitioning based SQGs, and results of sediment-spiking studies, these site- specific sediment TTs likely represent the concentrations of PCBs that are sufficient to cause toxicity to benthic invertebrates (as opposed to simply being correlated with adverse effects on the survival, weight, or reproduction of benthic invertebrates). Importantly, such site-specific sediment TTs have been demonstrated to accurately classify sediment samples as toxic or not toxic to benthic invertebrates at the Anniston PCB Site. In contrast, the TTs for metals, PAHs, and organochlorine pesticides were generally lower than consensus-based SQGs (that is, probable effect concentrations), and LC50s (median lethal effect concentrations) generated in spiked-sediment toxicity tests, indicating that these COPCs are likely not the main contributors to the observed toxicity of the site sediments evaluated in this study. The reproduction endpoint for H. azteca provided lower TTs compared to the day 28 biomass endpoint for H. azteca and the emergence or biomass endpoints for adult C. dilutus provided lower TTs compared to the day 13 biomass endpoint for C. dilutus.

Binding affinities of vascular endothelial growth factor (VEGF) for heparin-derived oligosaccharides

PubMed Central

Zhao, Wenjing; McCallum, Scott A.; Xiao, Zhongping; Zhang, Fuming; Linhardt, Robert J.

2011-01-01

Heparin and heparan sulphate (HS) exert their wide range of biological activities by interacting with extracellular protein ligands. Among these important protein ligands are various angiogenic growth factors and cytokines. HS-binding to vascular endothelial growth factor (VEGF) regulates multiple aspects of vascular development and function through its specific interaction with HS. Many studies have focused on HS-derived or HS-mimicking structures for the characterization of VEGF165 interaction with HS. Using a heparinase 1-prepared small library of heparin-derived oligosaccharides ranging from hexasaccharide to octadecasaccharide, we systematically investigated the heparin-specific structural features required for VEGF binding. We report the apparent affinities for the association between the heparin-derived oligosaccharides with both VEGF165 and VEGF55, a peptide construct encompassing exclusively the heparin-binding domain of VEGF165. An octasaccharide was the minimum size of oligosaccharide within the library to efficiently bind to both forms of VEGF and that a tetradecasaccharide displayed an effective binding affinity to VEGF165 comparable to unfractionated heparin. The range of relative apparent binding affinities among VEGF and the panel of heparin-derived oligosaccharides demonstrate that VEGF binding affinity likely depends on the specific structural features of these oligosaccharides including their degree of sulphation and sugar ring stereochemistry and conformation. Notably, the unique 3-O-sulpho group found within the specific antithrombin binding site of heparin is not required for VEGF165 binding. These findings afford new insight into the inherent kinetics and affinities for VEGF association with heparin and heparin-derived oligosaccharides with key residue specific modifications and may potentially benefit the future design of oligosaccharide-based anti-angiogenesis drugs. PMID:21658003

Antigenic Evolution of Vaccine-Derived Polioviruses: Changes in Individual Epitopes and Relative Stability of the Overall Immunological Properties

PubMed Central

Yakovenko, Maria L.; Cherkasova, Elena A.; Rezapkin, Gennady V.; Ivanova, Olga E.; Ivanov, Alexander P.; Eremeeva, Tatyana P.; Baykova, Olga Y.; Chumakov, Konstantin M.; Agol, Vadim I.

2006-01-01

The Sabin oral poliovirus vaccine (OPV) readily undergoes changes in antigenic sites upon replication in humans. Here, a set of antigenically altered descendants of the three OPV serotypes (76 isolates) was characterized to determine the driving forces behind these changes and their biological implications. The amino acid residues of OPV derivatives that lie within or close to the known antigenic sites exhibited a marked tendency to be replaced by residues characteristic of homotypic wild polioviruses, and these changes may occur very early in OPV evolution. The specific amino acid alterations nicely correlated with serotype-specific changes in the reactivity of certain individual antigenic sites, as revealed by the recently devised monoclonal antibody-based enzyme-linked immunosorbent assay. In comparison to the original vaccine, small changes, if any, in the neutralizing capacity of human or rabbit sera were observed in highly diverged vaccine polioviruses of three serotypes, in spite of strong alterations of certain epitopes. We propose that the common antigenic alterations in evolving OPV strains largely reflect attempts to eliminate fitness-decreasing mutations acquired either during the original selection of the vaccine or already present in the parental strains. Variability of individual epitopes does not appear to be primarily caused by, or lead to, a significant immune evasion, enhancing only slightly, if at all, the capacity of OPV derivatives to overcome immunity in human populations. This study reveals some important patterns of poliovirus evolution and has obvious implications for the rational design of live viral vaccines. PMID:16501074

Mapping replication dynamics in Trypanosoma brucei reveals a link with telomere transcription and antigenic variation

PubMed Central

Devlin, Rebecca; Marques, Catarina A; Paape, Daniel; Prorocic, Marko; Zurita-Leal, Andrea C; Campbell, Samantha J; Lapsley, Craig; Dickens, Nicholas; McCulloch, Richard

2016-01-01

Survival of Trypanosoma brucei depends upon switches in its protective Variant Surface Glycoprotein (VSG) coat by antigenic variation. VSG switching occurs by frequent homologous recombination, which is thought to require locus-specific initiation. Here, we show that a RecQ helicase, RECQ2, acts to repair DNA breaks, including in the telomeric site of VSG expression. Despite this, RECQ2 loss does not impair antigenic variation, but causes increased VSG switching by recombination, arguing against models for VSG switch initiation through direct generation of a DNA double strand break (DSB). Indeed, we show DSBs inefficiently direct recombination in the VSG expression site. By mapping genome replication dynamics, we reveal that the transcribed VSG expression site is the only telomeric site that is early replicating – a differential timing only seen in mammal-infective parasites. Specific association between VSG transcription and replication timing reveals a model for antigenic variation based on replication-derived DNA fragility. DOI: http://dx.doi.org/10.7554/eLife.12765.001 PMID:27228154

Mutations altering the cleavage specificity of a homing endonuclease

PubMed Central

Seligman, Lenny M.; Chisholm, Karen M.; Chevalier, Brett S.; Chadsey, Meggen S.; Edwards, Samuel T.; Savage, Jeremiah H.; Veillet, Adeline L.

2002-01-01

The homing endonuclease I-CreI recognizes and cleaves a particular 22 bp DNA sequence. The crystal structure of I-CreI bound to homing site DNA has previously been determined, leading to a number of predictions about specific protein–DNA contacts. We test these predictions by analyzing a set of endonuclease mutants and a complementary set of homing site mutants. We find evidence that all structurally predicted I-CreI/DNA contacts contribute to DNA recognition and show that these contacts differ greatly in terms of their relative importance. We also describe the isolation of a collection of altered specificity I-CreI derivatives. The in vitro DNA-binding and cleavage properties of two such endonucleases demonstrate that our genetic approach is effective in identifying homing endonucleases that recognize and cleave novel target sequences. PMID:12202772

49 CFR Appendix B to Part 222 - Alternative Safety Measures

Code of Federal Regulations, 2012 CFR

2012-10-01

... field data derived from the crossing sites. The specific crossing and applied mitigation measure will be... of a Public Highway-Rail Grade Crossing, (2) Four-Quadrant Gate System, (3) Gates With Medians or... as provided by 49 U.S.C. 20107. 3. Photo Enforcement: This ASM entails automated means of gathering...

Errors in Representing Regional Acid Deposition with Spatially Sparse Monitoring: Case Studies of the Eastern US Using Model Predictions

EPA Science Inventory

The current study uses case studies of model-estimated regional precipitation and wet ion deposition to estimate errors in corresponding regional values derived from the means of site-specific values within regions of interest located in the eastern US. The mean of model-estimate...

Australia’s first national level quantitative environmental justice assessment of industrial air pollution

NASA Astrophysics Data System (ADS)

Chakraborty, Jayajit; Green, Donna

2014-04-01

This study presents the first national level quantitative environmental justice assessment of industrial air pollution in Australia. Specifically, our analysis links the spatial distribution of sites and emissions associated with industrial pollution sources derived from the National Pollution Inventory, to Indigenous status and social disadvantage characteristics of communities derived from Australian Bureau of Statistics indicators. Our results reveal a clear national pattern of environmental injustice based on the locations of industrial pollution sources, as well as volume, and toxicity of air pollution released at these locations. Communities with the highest number of polluting sites, emission volume, and toxicity-weighted air emissions indicate significantly greater proportions of Indigenous population and higher levels of socio-economic disadvantage. The quantities and toxicities of industrial air pollution are particularly higher in communities with the lowest levels of educational attainment and occupational status. These findings emphasize the need for more detailed analysis in specific regions and communities where socially disadvantaged groups are disproportionately impacted by industrial air pollution. Our empirical findings also underscore the growing necessity to incorporate environmental justice considerations in environmental planning and policy-making in Australia.

Bridging disulfides for stable and defined antibody drug conjugates.

PubMed

Badescu, George; Bryant, Penny; Bird, Matthew; Henseleit, Korinna; Swierkosz, Julia; Parekh, Vimal; Tommasi, Rita; Pawlisz, Estera; Jurlewicz, Kosma; Farys, Monika; Camper, Nicolas; Sheng, XiaoBo; Fisher, Martin; Grygorash, Ruslan; Kyle, Andrew; Abhilash, Amrita; Frigerio, Mark; Edwards, Jeff; Godwin, Antony

2014-06-18

To improve both the homogeneity and the stability of ADCs, we have developed site-specific drug-conjugating reagents that covalently rebridge reduced disulfide bonds. The new reagents comprise a drug, a linker, and a bis-reactive conjugating moiety that is capable of undergoing reaction with both sulfur atoms derived from a reduced disulfide bond in antibodies and antibody fragments. A disulfide rebridging reagent comprising monomethyl auristatin E (MMAE) was prepared and conjugated to trastuzumab (TRA). A 78% conversion of antibody to ADC with a drug to antibody ratio (DAR) of 4 was achieved with no unconjugated antibody remaining. The MMAE rebridging reagent was also conjugated to the interchain disulfide of a Fab derived from proteolytic digestion of TRA, to give a homogeneous single drug conjugated product. The resulting conjugates retained antigen-binding, were stable in serum, and demonstrated potent and antigen-selective cell killing in in vitro and in vivo cancer models. Disulfide rebridging conjugation is a general approach to prepare stable ADCs, which does not require the antibody to be recombinantly re-engineered for site-specific conjugation.

Peak flood estimation using gene expression programming

NASA Astrophysics Data System (ADS)

Zorn, Conrad R.; Shamseldin, Asaad Y.

2015-12-01

As a case study for the Auckland Region of New Zealand, this paper investigates the potential use of gene-expression programming (GEP) in predicting specific return period events in comparison to the established and widely used Regional Flood Estimation (RFE) method. Initially calibrated to 14 gauged sites, the GEP derived model was further validated to 10 and 100 year flood events with a relative errors of 29% and 18%, respectively. This is compared to the RFE method providing 48% and 44% errors for the same flood events. While the effectiveness of GEP in predicting specific return period events is made apparent, it is argued that the derived equations should be used in conjunction with those existing methodologies rather than as a replacement.

A survey of Canadian websites providing information about female urinary incontinence.

PubMed

Farrell, Karen D; Robinson, Lynne M; Baydock, Sandra A; Farrell, Scott A; Irving, Linda E; O'Connell, Colleen M

2006-08-01

Urinary incontinence (UI) is a prevalent health issue that has significant detrimental effects on quality of life. The Internet offers a unique vehicle for incontinent women to access information that could facilitate conservative self-help therapy. An evaluation of Canadian websites offering female UI information was conducted to determine their quality and readability. We evaluated websites using published general quality criteria for health sites and a quality assessment tool compiled by the authors for specific UI information derived from published, peer-reviewed clinical practice guidelines. Three health care professionals reviewed sites for quality, Canadian content, and interactivity. The readability of health information was also evaluated. Fifty-six Canadian sites (18 professional, 22 organizational, 16 commercial) were evaluated. Significant agreement was found among the raters' evaluations on all measures. For all sites, the mean scores were general quality, 9/14; specific UI quality, 30/122; reading ease, 37/100; grade level, 10.9. The median score for Canadian content was high, but for interactivity it was low. The only significant difference between site types was for general quality (F [2,165]=3.38, P=0.036). Post hoc Tukey's tests showed a significant difference between organizational and commercial sites, with organizational sites having higher general quality. Canadian websites providing female UI information have moderately high general quality, low specific UI information quality, minimal interactivity, and more than minimal Canadian content. The reading level of most sites is too high for average consumers. A webliography of the best sites has been developed to guide patients.

A comparative study of matrix metalloproteinase and aggrecanase mediated release of latent cytokines at arthritic joints.

PubMed

Mullen, Lisa; Adams, Gill; Foster, Julie; Vessillier, Sandrine; Köster, Mario; Hauser, Hansjörg; Layward, Lorna; Gould, David; Chernajovsky, Yuti

2014-09-01

Latent cytokines are engineered by fusing the latency associated peptide (LAP) derived from transforming growth factor-β (TGF-β) with the therapeutic cytokine, in this case interferon-β (IFN-β), via an inflammation-specific matrix metalloproteinase (MMP) cleavage site. To demonstrate latency and specific delivery in vivo and to compare therapeutic efficacy of aggrecanase-mediated release of latent IFN-β in arthritic joints to the original MMP-specific release. Recombinant fusion proteins with MMP, aggrecanase or devoid of cleavage site were expressed in CHO cells, purified and characterised in vitro by Western blotting and anti-viral protection assays. Therapeutic efficacy and half-life were assessed in vivo using the mouse collagen-induced arthritis model (CIA) of rheumatoid arthritis and a model of acute paw inflammation, respectively. Transgenic mice with an IFN-regulated luciferase gene were used to assess latency in vivo and targeted delivery to sites of disease. Efficient localised delivery of IFN-β to inflamed paws, with low levels of systemic delivery, was demonstrated in transgenic mice using latent IFN-β. Engineering of latent IFN-β with an aggrecanase-sensitive cleavage site resulted in efficient cleavage by ADAMTS-4, ADAMTS-5 and synovial fluid from arthritic patients, with an extended half-life similar to the MMP-specific molecule and greater therapeutic efficacy in the CIA model. Latent cytokines require cleavage in vivo for therapeutic efficacy, and they are delivered in a dose dependent fashion only to arthritic joints. The aggrecanase-specific cleavage site is a viable alternative to the MMP cleavage site for the targeting of latent cytokines to arthritic joints. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

Delta ribozyme has the ability to cleave in transan mRNA.

PubMed Central

Roy, G; Ananvoranich, S; Perreault, J P

1999-01-01

We report here the first demonstration of the cleavage of an mRNA in trans by delta ribozyme derived from the antigenomic version of the human hepatitis delta virus (HDV). We characterized potential delta ribozyme cleavage sites within HDV mRNA sequence (i.e. C/UGN6), using oligonucleotide binding shift assays and ribonuclease H hydrolysis. Ribozymes were synthesized based on the structural data and then tested for their ability to cleave the mRNA. Of the nine ribozymes examined, three specifically cleaved a derivative HDV mRNA. All three active ribozymes gave consistent indications that they cleaved single-stranded regions. Kinetic characterization of the ability of ribozymes to cleave both the full-length mRNA and either wild-type or mutant small model substrate suggests: (i) delta ribozyme has turnovers, that is to say, several mRNA molecules can be successively cleaved by one ribozyme molecule; and (ii) the substrate specificity of delta ribozyme cleavage is not restricted to C/UGN6. Specifically, substrates with a higher guanosine residue content upstream of the cleavage site (i.e. positions -4 to -2) were always cleaved more efficiently than wild-type substrate. This work shows that delta ribozyme constitutes a potential catalytic RNA for further gene-inactivation therapy. PMID:9927724

Genome Integration and Excision by a New Streptomyces Bacteriophage, ϕJoe.

PubMed

Fogg, Paul C M; Haley, Joshua A; Stark, W Marshall; Smith, Margaret C M

2017-03-01

Bacteriophages are the source of many valuable tools for molecular biology and genetic manipulation. In Streptomyces , most DNA cloning vectors are based on serine integrase site-specific DNA recombination systems derived from phage. Because of their efficiency and simplicity, serine integrases are also used for diverse synthetic biology applications. Here, we present the genome of a new Streptomyces phage, ϕJoe, and investigate the conditions for integration and excision of the ϕJoe genome. ϕJoe belongs to the largest Streptomyces phage cluster (R4-like) and encodes a serine integrase. The attB site from Streptomyces venezuelae was used efficiently by an integrating plasmid, pCMF92, constructed using the ϕJoe int-attP locus. The attB site for ϕJoe integrase was occupied in several Streptomyces genomes, including that of S. coelicolor , by a mobile element that varies in gene content and size between host species. Serine integrases require a phage-encoded recombination directionality factor (RDF) to activate the excision reaction. The ϕJoe RDF was identified, and its function was confirmed in vivo Both the integrase and RDF were active in in vitro recombination assays. The ϕJoe site-specific recombination system is likely to be an important addition to the synthetic biology and genome engineering toolbox. IMPORTANCE Streptomyces spp. are prolific producers of secondary metabolites, including many clinically useful antibiotics. Bacteriophage-derived integrases are important tools for genetic engineering, as they enable integration of heterologous DNA into the Streptomyces chromosome with ease and high efficiency. Recently, researchers have been applying phage integrases for a variety of applications in synthetic biology, including rapid assembly of novel combinations of genes, biosensors, and biocomputing. An important requirement for optimal experimental design and predictability when using integrases, however, is the need for multiple enzymes with different specificities for their integration sites. In order to provide a broad platform of integrases, we identified and validated the integrase from a newly isolated Streptomyces phage, ϕJoe. ϕJoe integrase is active in vitro and in vivo The specific recognition site for integration is present in a wide range of different actinobacteria, including Streptomyces venezuelae , an emerging model bacterium in Streptomyces research. Copyright © 2017 Fogg et al.

Evaluating Variability and Uncertainty of Geological Strength Index at a Specific Site

NASA Astrophysics Data System (ADS)

Wang, Yu; Aladejare, Adeyemi Emman

2016-09-01

Geological Strength Index (GSI) is an important parameter for estimating rock mass properties. GSI can be estimated from quantitative GSI chart, as an alternative to the direct observational method which requires vast geological experience of rock. GSI chart was developed from past observations and engineering experience, with either empiricism or some theoretical simplifications. The GSI chart thereby contains model uncertainty which arises from its development. The presence of such model uncertainty affects the GSI estimated from GSI chart at a specific site; it is, therefore, imperative to quantify and incorporate the model uncertainty during GSI estimation from the GSI chart. A major challenge for quantifying the GSI chart model uncertainty is a lack of the original datasets that have been used to develop the GSI chart, since the GSI chart was developed from past experience without referring to specific datasets. This paper intends to tackle this problem by developing a Bayesian approach for quantifying the model uncertainty in GSI chart when using it to estimate GSI at a specific site. The model uncertainty in the GSI chart and the inherent spatial variability in GSI are modeled explicitly in the Bayesian approach. The Bayesian approach generates equivalent samples of GSI from the integrated knowledge of GSI chart, prior knowledge and observation data available from site investigation. Equations are derived for the Bayesian approach, and the proposed approach is illustrated using data from a drill and blast tunnel project. The proposed approach effectively tackles the problem of how to quantify the model uncertainty that arises from using GSI chart for characterization of site-specific GSI in a transparent manner.

Comparison of screening-level and Monte Carlo approaches for wildlife food web exposure modeling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pastorok, R.; Butcher, M.; LaTier, A.

1995-12-31

The implications of using quantitative uncertainty analysis (e.g., Monte Carlo) and site-specific tissue residue data for wildlife exposure modeling were examined with data on trace elements at the Clark Fork River Superfund Site. Exposure of white-tailed deer, red fox, and American kestrel was evaluated using three approaches. First, a screening-level exposure model was based on conservative estimates of exposure parameters, including estimates of dietary residues derived from bioconcentration factors (BCFs) and soil chemistry. A second model without Monte Carlo was based on site-specific data for tissue residues of trace elements (As, Cd, Cu, Pb, Zn) in key dietary species andmore » plausible assumptions for habitat spatial segmentation and other exposure parameters. Dietary species sampled included dominant grasses (tufted hairgrass and redtop), willows, alfalfa, barley, invertebrates (grasshoppers, spiders, and beetles), and deer mice. Third, the Monte Carlo analysis was based on the site-specific residue data and assumed or estimated distributions for exposure parameters. Substantial uncertainties are associated with several exposure parameters, especially BCFS, such that exposure and risk may be greatly overestimated in screening-level approaches. The results of the three approaches are compared with respect to realism, practicality, and data gaps. Collection of site-specific data on trace elements concentrations in plants and animals eaten by the target wildlife receptors is a cost-effective way to obtain realistic estimates of exposure. Implications of the results for exposure and risk estimates are discussed relative to use of wildlife exposure modeling and evaluation of remedial actions at Superfund sites.« less

Insights into the functionality of the putative residues involved in enterocin AS-48 maturation.

PubMed

Cebrián, Rubén; Maqueda, Mercedes; Neira, José Luis; Valdivia, Eva; Martínez-Bueno, Manuel; Montalbán-López, Manuel

2010-11-01

AS-48 is a 70-residue, α-helical, cationic bacteriocin produced by Enterococcus faecalis and is very singular in its circular structure and its broad antibacterial spectrum. The AS-48 preprotein consists of an N-terminal signal peptide (SP) (35 residues) followed by a proprotein moiety that undergoes posttranslational modifications to yield the mature and active circular protein. For the study of the specificity of the region of AS-48 that is responsible for maturation, three single mutants have been generated by site-directed mutagenesis in the as-48A structural gene. The substitutions were made just in the residues that are thought to constitute a recognition site for the SP cleavage enzyme (His-1, Met1) and in those involved in circularization (Met1, Trp70). Each derivative was expressed in the enterococcal JH2-2 strain containing the necessary native biosynthetic machinery for enterocin production. The importance of these derivatives in AS-48 processing has been evaluated on the basis of the production and structural characterization of the corresponding derivatives. Notably, only two of them (Trp70Ala and Met1Ala derivatives) could be purified in different forms and amounts and are characterized for their bactericidal activity and secondary structure. We could not detect any production of AS-48 in JH2-2(pAM401-81(His-1Ile)) by using the conventional chromatographic techniques, despite the high efficiency of the culture conditions applied to produce this enterocin. Our results underline the different important roles of the mutated residues in (i) the elimination of the SP, (ii) the production levels and antibacterial activity of the mature proteins, and (iii) protein circularization. Moreover, our findings suggest that His-1 is critically involved in cleavage site recognition, its substitution being responsible for the blockage of processing, thereby hampering the production of the specific protein in the cellular culture supernatant.

Positional differences in the wound transcriptome of skin and oral mucosa

PubMed Central

2010-01-01

Background When compared to skin, oral mucosal wounds heal rapidly and with reduced scar formation. Recent studies suggest that intrinsic differences in inflammation, growth factor production, levels of stem cells, and cellular proliferation capacity may underlie the exceptional healing that occurs in oral mucosa. The current study was designed to compare the transcriptomes of oral mucosal and skin wounds in order to identify critical differences in the healing response at these two sites using an unbiased approach. Results Using microarray analysis, we explored the differences in gene expression in skin and oral mucosal wound healing in a murine model of paired equivalent sized wounds. Samples were examined from days 0 to 10 and spanned all stages of the wound healing process. Using unwounded matched tissue as a control, filtering identified 1,479 probe sets in skin wounds yet only 502 probe sets in mucosal wounds that were significantly differentially expressed over time. Clusters of genes that showed similar patterns of expression were also identified in each wound type. Analysis of functionally related gene expression demonstrated dramatically different reactions to injury between skin and mucosal wounds. To explore whether site-specific differences might be derived from intrinsic differences in cellular responses at each site, we compared the response of isolated epithelial cells from skin and oral mucosa to a defined in vitro stimulus. When cytokine levels were measured, epithelial cells from skin produced significantly higher amounts of proinflammatory cytokines than cells from oral mucosa. Conclusions The results provide the first detailed molecular profile of the site-specific differences in the genetic response to injury in mucosa and skin, and suggest the divergent reactions to injury may derive from intrinsic differences in the cellular responses at each site. PMID:20704739

Positional differences in the wound transcriptome of skin and oral mucosa.

PubMed

Chen, Lin; Arbieva, Zarema H; Guo, Shujuan; Marucha, Phillip T; Mustoe, Thomas A; DiPietro, Luisa A

2010-08-12

When compared to skin, oral mucosal wounds heal rapidly and with reduced scar formation. Recent studies suggest that intrinsic differences in inflammation, growth factor production, levels of stem cells, and cellular proliferation capacity may underlie the exceptional healing that occurs in oral mucosa. The current study was designed to compare the transcriptomes of oral mucosal and skin wounds in order to identify critical differences in the healing response at these two sites using an unbiased approach. Using microarray analysis, we explored the differences in gene expression in skin and oral mucosal wound healing in a murine model of paired equivalent sized wounds. Samples were examined from days 0 to 10 and spanned all stages of the wound healing process. Using unwounded matched tissue as a control, filtering identified 1,479 probe sets in skin wounds yet only 502 probe sets in mucosal wounds that were significantly differentially expressed over time. Clusters of genes that showed similar patterns of expression were also identified in each wound type. Analysis of functionally related gene expression demonstrated dramatically different reactions to injury between skin and mucosal wounds. To explore whether site-specific differences might be derived from intrinsic differences in cellular responses at each site, we compared the response of isolated epithelial cells from skin and oral mucosa to a defined in vitro stimulus. When cytokine levels were measured, epithelial cells from skin produced significantly higher amounts of proinflammatory cytokines than cells from oral mucosa. The results provide the first detailed molecular profile of the site-specific differences in the genetic response to injury in mucosa and skin, and suggest the divergent reactions to injury may derive from intrinsic differences in the cellular responses at each site.

An MRM-based workflow for quantifying cardiac mitochondrial protein phosphorylation in murine and human tissue.

PubMed

Lam, Maggie P Y; Scruggs, Sarah B; Kim, Tae-Young; Zong, Chenggong; Lau, Edward; Wang, Ding; Ryan, Christopher M; Faull, Kym F; Ping, Peipei

2012-08-03

The regulation of mitochondrial function is essential for cardiomyocyte adaptation to cellular stress. While it has long been understood that phosphorylation regulates flux through metabolic pathways, novel phosphorylation sites are continually being discovered in all functionally distinct areas of the mitochondrial proteome. Extracting biologically meaningful information from these phosphorylation sites requires an adaptable, sensitive, specific and robust method for their quantification. Here we report a multiple reaction monitoring-based mass spectrometric workflow for quantifying site-specific phosphorylation of mitochondrial proteins. Specifically, chromatographic and mass spectrometric conditions for 68 transitions derived from 23 murine and human phosphopeptides, and their corresponding unmodified peptides, were optimized. These methods enabled the quantification of endogenous phosphopeptides from the outer mitochondrial membrane protein VDAC, and the inner membrane proteins ANT and ETC complexes I, III and V. The development of this quantitative workflow is a pivotal step for advancing our knowledge and understanding of the regulatory effects of mitochondrial protein phosphorylation in cardiac physiology and pathophysiology. This article is part of a Special Issue entitled: Translational Proteomics. Copyright © 2012 Elsevier B.V. All rights reserved.

Interaction of SR 33557 with skeletal muscle calcium channel blocker receptors in the baboon: characterization of its binding sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sol-Rolland, J.; Joseph, M.; Rinaldi-Carmona, M.

1991-05-01

A procedure for the isolation of primate skeletal microsomal membranes was initiated. Membranes exhibited specific enzymatic markers such as 5'-nucleotidase, Ca{sup 2}{sup +},Mg({sup 2}{sup +})-adenosine triphosphatase and an ATP-dependent calcium uptake. Baboon skeletal microsomes bound specifically with high-affinity potent Ca{sup 2}{sup +} channel blockers such as dihydropyridine, phenylalkylamine and benzothiazepine derivatives. Scatchard analysis of equilibrium binding assays with ({sup 3}H)(+)-PN 200-110, ({sup 3}H)(-)-desmethoxyverapamil (( {sup 3}H)(-)-D888) and ({sup 3}H)-d-cis-dilitiazem were consistent with a single class of binding sites for the three radioligands. The pharmacological profile of SR 33557, an original compound with calcium antagonist properties, was investigated using radioligand bindingmore » studies. SR 33557 totally inhibited the specific binding of the three main classes of Ca{sup 2}{sup +} channel effectors and interacted allosterically with them. In addition, SR 33557 bound with high affinity to a homogeneous population of binding sites in baboon skeletal muscle.« less

Developing and Testing Locally Derived Mental Health Scales: Examples from North India and Haiti

ERIC Educational Resources Information Center

Weaver, Lesley Jo; Kaiser, Bonnie N.

2015-01-01

Cross-cultural studies of mental health and illness generally adhere to one of two agendas: the comparison of mental health between sites using standard measurement tools, or the identification of locally specific ways of discussing mental illness. Here, we illustrate a methodological approach to measuring mental health that unites these two…

21 CFR 528.1070 - Bc6 recombinant deoxyribonucleic acid construct.

Code of Federal Regulations, 2010 CFR

2010-04-01

... 155-92 site in a specific hemizygous diploid line of dairy breeds of domestic goats (Capra aegagrus... of humans) in the mammary gland of goats derived from lineage progenitor 155-92. (b) Sponsor. See No. 042976 in § 510.600 of this chapter. (c) Limitations. Food or feed from GTC-155-92 goats is not permitted...

21 CFR 528.1070 - Bc6 recombinant deoxyribonucleic acid construct.

Code of Federal Regulations, 2011 CFR

2011-04-01

... 155-92 site in a specific hemizygous diploid line of dairy breeds of domestic goats (Capra aegagrus... of humans) in the mammary gland of goats derived from lineage progenitor 155-92. (b) Sponsor. See No. 042976 in § 510.600 of this chapter. (c) Limitations. Food or feed from GTC-155-92 goats is not permitted...

76 FR 31379 - Notice of Availability of Environmental Assessment and Finding of No Significant Impact for...

Federal Register 2010, 2011, 2012, 2013, 2014

2011-05-31

... for approval as an addendum to the DP, Revision 2 (Derivation of the Site-Specific Soil DCGLs Addendum Soil DCGLs for Thorium and Radium). The revised DP does not change any previously approved remediation... ensures safety and protection of the public and the environment. Environmental Impacts of the Proposed...

Hypochlorite and superoxide radicals can act synergistically to induce fragmentation of hyaluronan and chondroitin sulphates

PubMed Central

2004-01-01

Activated phagocytes release the haem enzyme MPO (myeloperoxidase) and also generate superoxide radicals (O2•−), and hence H2O2, via an oxidative burst. Reaction of MPO with H2O2 in the presence of chloride ions generates HOCl (the physiological mixture of hypochlorous acid and its anion present at pH 7.4). Exposure of glycosaminoglycans to a MPO–H2O2–Cl− system or reagent HOCl generates long-lived chloramides [R-NCl-C(O)-R′] derived from the glycosamine N-acetyl functions. Decomposition of these species by transition metal ions gives polymer-derived amidyl (nitrogen-centred) radicals [R-N•-C(O)-R′], polymer-derived carbon-centred radicals and site-specific strand scission. In the present study, we have shown that exposure of glycosaminoglycan chloramides to O2•− also promotes chloramide decomposition and glycosaminoglycan fragmentation. These processes are inhibited by superoxide dismutase, metal ion chelators and the metal ion-binding protein BSA, consistent with chloramide decomposition and polymer fragmentation occurring via O2•−-dependent one-electron reduction, possibly catalysed by trace metal ions. Polymer fragmentation induced by O2•− [generated by the superoxide thermal source 1, di-(4-carboxybenzyl)hyponitrite] was demonstrated to be entirely chloramide dependent as no fragmentation occurred with the native polymers or when the chloramides were quenched by prior treatment with methionine. EPR spin-trapping experiments using 5,5-dimethyl1-pyrroline-N-oxide and 2-methyl-2-nitrosopropane have provided evidence for both O2•− and polymer-derived carbon-centred radicals as intermediates. The results obtained are consistent with a mechanism involving one-electron reduction of the chloramides to yield polymer-derived amidyl radicals, which subsequently undergo intramolecular hydrogen atom abstraction reactions to give carbon-centred radicals. The latter undergo fragmentation reactions in a site-specific manner. This synergistic damage to glycosaminoglycans induced by HOCl and O2•− may be of significance at sites of inflammation where both oxidants are generated concurrently. PMID:15078224

Hydraulic-Geometry Relations for Rivers in Coastal and Central Maine

USGS Publications Warehouse

Dudley, Robert W.

2004-01-01

Hydraulic-geometry relations (curves) were derived for 15 sites on 12 rivers in coastal and central Maine on the basis of site-specific (at-a-station) hydraulic-geometry relations and hydraulic models. At-a-station hydraulic-geometry curves, expressed as well-established power functions, describe the relations between channel geometry, velocity, and flow at a given point on a river. The derived at-a-station hydraulic-geometry curves indicate that, on average, a given increase in flow at a given river cross section in the study area will be nearly equally conveyed by increases in velocity and channel cross-sectional area. Regional curves describing the bankfull streamflow and associated channel geometry as functions of drainage area were derived for use in stream-channel assessment and restoration projects specific to coastal and central Maine. Regional hydraulic-geometry curves were derived by combining hydraulic-geometry information for 15 river cross sections using bankfull flow as the common reference streamflow. The exponents of the derived regional hydraulic-geometry relations indicate that, in the downstream direction, most of the conveyance of increasing contribution of flow is accommodated by an increase in cross-sectional area?with about 50 percent of the increase in flow accommodated by an increase in channel width, and 32 percent by an increase in depth. The remaining 18 percent is accommodated by an increase in streamflow velocity. On an annual-peak-series basis, results of this study indicate that the occurrence of bankfull streamflow for rivers in Maine is more frequent than the 1.5-year streamflow. On a flow-duration basis, bankfull streamflow for rivers in coastal and central Maine is equaled or exceeded approximately 8.1 percent of the time on mean?or about 30 days a year. Bankfull streamflow is roughly three times that of the mean annual streamflow for the sites investigated in this study. Regional climate, snowmelt hydrology, and glacial geology may play important roles in dictating the magnitude and frequency of occurrence of bankfull streamflows observed for rivers in coastal and central Maine.

TESS: a geometric hashing algorithm for deriving 3D coordinate templates for searching structural databases. Application to enzyme active sites.

PubMed Central

Wallace, A. C.; Borkakoti, N.; Thornton, J. M.

1997-01-01

It is well established that sequence templates such as those in the PROSITE and PRINTS databases are powerful tools for predicting the biological function and tertiary structure for newly derived protein sequences. The number of X-ray and NMR protein structures is increasing rapidly and it is apparent that a 3D equivalent of the sequence templates is needed. Here, we describe an algorithm called TESS that automatically derives 3D templates from structures deposited in the Brookhaven Protein Data Bank. While a new sequence can be searched for sequence patterns, a new structure can be scanned against these 3D templates to identify functional sites. As examples, 3D templates are derived for enzymes with an O-His-O "catalytic triad" and for the ribonucleases and lysozymes. When these 3D templates are applied to a large data set of nonidentical proteins, several interesting hits are located. This suggests that the development of a 3D template database may help to identify the function of new protein structures, if unknown, as well as to design proteins with specific functions. PMID:9385633

Generic Assessment Criteria for human health risk assessment of potentially contaminated land in China.

PubMed

Cheng, Yuanyuan; Nathanail, Paul C

2009-12-20

Generic Assessment Criteria (GAC) are derived using widely applicable assumptions about the characteristics and behaviour of contaminant sources, pathways and receptors. GAC provide nationally consistent guidance, thereby saving money and time. Currently, there are no human health based Generic Assessment Criteria (GAC) for contaminated sites in China. Protection of human health is therefore difficult to ensure and demonstrate; and the lack of GAC makes it difficult to tell if there is potential significant risk to human health unless site-specific criteria are derived. This paper derived Chinese GAC (GAC) for five inorganic and eight organic substances for three regions in China for three land uses: urban residential without plant uptake, Chinese cultivated land, and commercial/industrial using the SNIFFER model. The SNIFFER model has been further implemented with a dermal absorption algorithm and the model default input values have been changed to reflect the Chinese exposure scenarios. It is envisaged that the modified SNIFFER model could be used to derive GAC for more contaminants, more Regions, and more land uses. Further research to enhance the reliability and acceptability of the GAC is needed in regional/national surveys in diet and working patterns.

Probabilistic analysis of tsunami hazards

USGS Publications Warehouse

Geist, E.L.; Parsons, T.

2006-01-01

Determining the likelihood of a disaster is a key component of any comprehensive hazard assessment. This is particularly true for tsunamis, even though most tsunami hazard assessments have in the past relied on scenario or deterministic type models. We discuss probabilistic tsunami hazard analysis (PTHA) from the standpoint of integrating computational methods with empirical analysis of past tsunami runup. PTHA is derived from probabilistic seismic hazard analysis (PSHA), with the main difference being that PTHA must account for far-field sources. The computational methods rely on numerical tsunami propagation models rather than empirical attenuation relationships as in PSHA in determining ground motions. Because a number of source parameters affect local tsunami runup height, PTHA can become complex and computationally intensive. Empirical analysis can function in one of two ways, depending on the length and completeness of the tsunami catalog. For site-specific studies where there is sufficient tsunami runup data available, hazard curves can primarily be derived from empirical analysis, with computational methods used to highlight deficiencies in the tsunami catalog. For region-wide analyses and sites where there are little to no tsunami data, a computationally based method such as Monte Carlo simulation is the primary method to establish tsunami hazards. Two case studies that describe how computational and empirical methods can be integrated are presented for Acapulco, Mexico (site-specific) and the U.S. Pacific Northwest coastline (region-wide analysis).

Dynamics Govern Specificity of a Protein-Protein Interface: Substrate Recognition by Thrombin.

PubMed

Fuchs, Julian E; Huber, Roland G; Waldner, Birgit J; Kahler, Ursula; von Grafenstein, Susanne; Kramer, Christian; Liedl, Klaus R

2015-01-01

Biomolecular recognition is crucial in cellular signal transduction. Signaling is mediated through molecular interactions at protein-protein interfaces. Still, specificity and promiscuity of protein-protein interfaces cannot be explained using simplistic static binding models. Our study rationalizes specificity of the prototypic protein-protein interface between thrombin and its peptide substrates relying solely on binding site dynamics derived from molecular dynamics simulations. We find conformational selection and thus dynamic contributions to be a key player in biomolecular recognition. Arising entropic contributions complement chemical intuition primarily reflecting enthalpic interaction patterns. The paradigm "dynamics govern specificity" might provide direct guidance for the identification of specific anchor points in biomolecular recognition processes and structure-based drug design.

Immobilization of metals in contaminated soil from E-waste recycling site by dairy-manure-derived biochar.

PubMed

Chen, Zhiliang; Zhang, Jianqiang; Liu, Minchao; Wu, Yingxin; Yuan, Zhihui

2017-08-24

E-waste is a growing concern around the world and varieties of abandoned E-waste recycling sites, especially in urban area, need to remediate immediately. The impacts of dairy-manure-derived biochars (BCs) on the amelioration of soil properties, the changes in the morphologies as well as the mobility of metals were studied to test their efficacy in immobilization of metals for a potential restoration of vegetation landscape in abandoned E-waste recycling site. The amendment with BCs produced positive effects on bioavailability and mobility reduction for Pb, Cd, Zn and Cu depending on BC ratio and incubation time. The BCs promoted the transformation of species of heavy metals to a more stable fraction, and the metals concentrations in Toxicity Characteristic Leaching Procedure extract declined significantly, especially Pb and Cu. Besides, the BCs ameliorated the substrate with increasing the soil pH, cations exchangeable capacity and available phosphorous, which suggested BC as a potential amendment material for abandoned E-waste recycling sites before restoration of vegetation landscape. Generally, the BC modified by alkaline treatment has a higher efficacy, probably due to increase of specific surface area and porosity as well as the functional groups after alkaline treatment.

Does Osmotic Stress Affect Natural Product Expression in Fungi?

PubMed

Overy, David; Correa, Hebelin; Roullier, Catherine; Chi, Wei-Chiung; Pang, Ka-Lai; Rateb, Mostafa; Ebel, Rainer; Shang, Zhuo; Capon, Rob; Bills, Gerald; Kerr, Russell

2017-08-13

The discovery of new natural products from fungi isolated from the marine environment has increased dramatically over the last few decades, leading to the identification of over 1000 new metabolites. However, most of the reported marine-derived species appear to be terrestrial in origin yet at the same time, facultatively halo- or osmotolerant. An unanswered question regarding the apparent chemical productivity of marine-derived fungi is whether the common practice of fermenting strains in seawater contributes to enhanced secondary metabolism? To answer this question, a terrestrial isolate of Aspergillus aculeatus was fermented in osmotic and saline stress conditions in parallel across multiple sites. The ex-type strain of A. aculeatus was obtained from three different culture collections. Site-to-site variations in metabolite expression were observed, suggesting that subculturing of the same strain and subtle variations in experimental protocols can have pronounced effects upon metabolite expression. Replicated experiments at individual sites indicated that secondary metabolite production was divergent between osmotic and saline treatments. Titers of some metabolites increased or decreased in response to increasing osmolite (salt or glycerol) concentrations. Furthermore, in some cases, the expression of some secondary metabolites in relation to osmotic and saline stress was attributed to specific sources of the ex-type strains.

Linear alkylbenzenes as tracers of sewage-sludge-derived inputs of organic matter, PCBs, and PAHs to sediments at the 106-mile deep water disposal site

USGS Publications Warehouse

Lamoureux, E.M.; Brownawell, Bruce J.; Bothner, Michael H.

1996-01-01

Linear alkylbenzenes (LABs) are sensitive source-specific tracers of sewage inputs to the marine environment. Because they are highly particle reactive and nonspecifically sorbed to organic matter, LABs are potential tracers of the transport of both sludge-derived organic matter and other low solubility hydrophobic contaminants (e.g., PCBs and PAHs); sediment trap studies at the 106-Mile Site have shown LABs to be valuable in testing models of sludge deposition to the sea floor. In this study we report on the distributions of LABs, PCBs, PAHs, and Ag in surface sediments collected within a month of the complete cessation of dumping (July, 1992) in the vicinity of the dump site. Total LAB concentrations were lower than those measured by Takada and coworkers in samples from nearby sites collected in 1989. LABs from both studies appear to be significantly depleted (6 to 25-fold) in surface sediments relative to excess Ag (another sludge tracer) when compared to sewage sludge and sediment trap compositions. Comparison of LAB sediment inventories to model predictions of sludge particle fluxes supports the contention that LABs have been lost from the bed. The use of LABs to examine the short-or long-term fate of sludge derived materials in deep-sea sediments should be questioned. The causes of this LAB depletion are unclear at this point, and we discuss several hypotheses. The concentrations of total PCBs and PAHs are both correlated with sludge tracers, suggesting that there may be a measurable contribution of sludge-derived inputs on top of other nonpoint sources of these contaminant classes. This possibility is consistent with the composition of these contaminants determined in recent and historical analyses of sewage sludge.

Photoaffinity-labeling and fluorescence-distribution studies of gonadotropin-releasing hormone receptors in ovarian granulosa cells.

PubMed Central

Hazum, E; Nimrod, A

1982-01-01

Photoaffinity labeling of rat ovarian granulosa cells and membrane preparations with a bioactive photoaffinity derivative of gonadotropin-releasing hormone resulted in identification of two specific components with apparent molecular weights of 60,000 and 54,000. Fluorescent visualization of gonadotropin-releasing hormone receptors in these cells, by using a bioactive rhodamine derivative of the hormone, indicated that the fluorescently labeled receptors were initially distributed uniformly on the cell surface and then formed patches that subsequently internalized (at 37 degrees C) into endocytic vesicles. These processes were dependent on specific binding sites for the rhodamine-labeled peptide on the granulosa cells. These studies may provide an experimental basis for understanding the molecular events involved in the action of the hormone in the ovary. Images PMID:6281784

IFI44L promoter methylation as a blood biomarker for systemic lupus erythematosus

PubMed Central

Zhao, Ming; Zhou, Yin; Zhu, Bochen; Wan, Mengjie; Jiang, Tingting; Tan, Qiqun; Liu, Yan; Jiang, Juqing; Luo, Shuaihantian; Tan, Yixin; Wu, Haijing; Renauer, Paul; Gutiérrez, Maria del Mar Ayala; Palma, Maria Jesús Castillo; Castro, Rafaela Ortega; Fernández-Roldán, Concepción; Raya, Enrique; Faria, Raquel; Carvalho, Claudia; Alarcón-Riquelme, Marta E; Xiang, Zhongyuan; Chen, Jinwei; Li, Fen; Ling, Guanghui; Zhao, Hongjun; Liao, Xiangping; Lin, Youkun; Sawalha, Amr H; Lu, Qianjin

2016-01-01

Objective Systemic lupus erythematosus (SLE) is a clinically heterogeneous disease with limited reliable diagnostic biomarkers. We investigated whether gene methylation could meet sensitivity and specificity criteria for a robust biomarker. Methods IFI44L promoter methylation was examined using DNA samples from a discovery set including 377 patients with SLE, 358 healthy controls (HCs) and 353 patients with rheumatoid arthritis (RA). Two independent sets including 1144 patients with SLE, 1350 HCs, 429 patients with RA and 199 patients with primary Sjögren’s syndrome (pSS) were used for validation. Results Significant hypomethylation of two CpG sites within IFI44L promoter, Site1 (Chr1: 79 085 222) and Site2 (Chr1: 79 085 250; cg06872964), was identified in patients with SLE compared with HCs, patients with RA and patients with pSS. In a comparison between patients with SLE and HCs included in the first validation cohort, Site1 methylation had a sensitivity of 93.6% and a specificity of 96.8% at a cut-off methylation level of 75.5% and Site2 methylation had a sensitivity of 94.1% and a specificity of 98.2% at a cut-off methylation level of 25.5%. The IFI44L promoter methylation marker was also validated in an European-derived cohort. In addition, the methylation levels of Site1 and Site2 within IFI44L promoter were significantly lower in patients with SLE with renal damage than those without renal damage. Patients with SLE showed significantly increased methylation levels of Site1 and Site2 during remission compared with active stage. Conclusions The methylation level of IFI44L promoter can distinguish patients with SLE from healthy persons and other autoimmune diseases, and is a highly sensitive and specific diagnostic marker for SLE. PMID:26787370

IFI44L promoter methylation as a blood biomarker for systemic lupus erythematosus.

PubMed

Zhao, Ming; Zhou, Yin; Zhu, Bochen; Wan, Mengjie; Jiang, Tingting; Tan, Qiqun; Liu, Yan; Jiang, Juqing; Luo, Shuaihantian; Tan, Yixin; Wu, Haijing; Renauer, Paul; Del Mar Ayala Gutiérrez, Maria; Castillo Palma, Maria Jesús; Ortega Castro, Rafaela; Fernández-Roldán, Concepción; Raya, Enrique; Faria, Raquel; Carvalho, Claudia; Alarcón-Riquelme, Marta E; Xiang, Zhongyuan; Chen, Jinwei; Li, Fen; Ling, Guanghui; Zhao, Hongjun; Liao, Xiangping; Lin, Youkun; Sawalha, Amr H; Lu, Qianjin

2016-11-01

Systemic lupus erythematosus (SLE) is a clinically heterogeneous disease with limited reliable diagnostic biomarkers. We investigated whether gene methylation could meet sensitivity and specificity criteria for a robust biomarker. IFI44L promoter methylation was examined using DNA samples from a discovery set including 377 patients with SLE, 358 healthy controls (HCs) and 353 patients with rheumatoid arthritis (RA). Two independent sets including 1144 patients with SLE, 1350 HCs, 429 patients with RA and 199 patients with primary Sjögren's syndrome (pSS) were used for validation. Significant hypomethylation of two CpG sites within IFI44L promoter, Site1 (Chr1: 79 085 222) and Site2 (Chr1: 79 085 250; cg06872964), was identified in patients with SLE compared with HCs, patients with RA and patients with pSS. In a comparison between patients with SLE and HCs included in the first validation cohort, Site1 methylation had a sensitivity of 93.6% and a specificity of 96.8% at a cut-off methylation level of 75.5% and Site2 methylation had a sensitivity of 94.1% and a specificity of 98.2% at a cut-off methylation level of 25.5%. The IFI44L promoter methylation marker was also validated in an European-derived cohort. In addition, the methylation levels of Site1 and Site2 within IFI44L promoter were significantly lower in patients with SLE with renal damage than those without renal damage. Patients with SLE showed significantly increased methylation levels of Site1 and Site2 during remission compared with active stage. The methylation level of IFI44L promoter can distinguish patients with SLE from healthy persons and other autoimmune diseases, and is a highly sensitive and specific diagnostic marker for SLE. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

On the relationship between ecosystem-scale hyperspectral reflectance and CO2 exchange in European mountain grasslands

NASA Astrophysics Data System (ADS)

Balzarolo, M.; Vescovo, L.; Hammerle, A.; Gianelle, D.; Papale, D.; Tomelleri, E.; Wohlfahrt, G.

2015-05-01

In this paper we explore the skill of hyperspectral reflectance measurements and vegetation indices (VIs) derived from these in estimating carbon dioxide (CO2) fluxes of grasslands. Hyperspectral reflectance data, CO2 fluxes and biophysical parameters were measured at three grassland sites located in European mountain regions using standardized protocols. The relationships between CO2 fluxes, ecophysiological variables, traditional VIs and VIs derived using all two-band combinations of wavelengths available from the whole hyperspectral data space were analysed. We found that VIs derived from hyperspectral data generally explained a large fraction of the variability in the investigated dependent variables but differed in their ability to estimate midday and daily average CO2 fluxes and various derived ecophysiological parameters. Relationships between VIs and CO2 fluxes and ecophysiological parameters were site-specific, likely due to differences in soils, vegetation parameters and environmental conditions. Chlorophyll and water-content-related VIs explained the largest fraction of variability in most of the dependent variables. Band selection based on a combination of a genetic algorithm with random forests (GA-rF) confirmed that it is difficult to select a universal band region suitable across the investigated ecosystems. Our findings have major implications for upscaling terrestrial CO2 fluxes to larger regions and for remote- and proximal-sensing sampling and analysis strategies and call for more cross-site synthesis studies linking ground-based spectral reflectance with ecosystem-scale CO2 fluxes.

Summary and Preliminary Interpretation of Tritium and Dissolved Noble Gas Data from Site 300

DOE Office of Scientific and Technical Information (OSTI.GOV)

Visser, A.; Singleton, M.; Madrid, V.

2014-01-29

In October 2013, groundwater samples were collected from 10 wells from Site 300 and analyzed by the Environmental Radiochemistry Laboratory at Lawrence Livermore National Laboratory (LLNL). Groundwater samples were analyzed for groundwater age tracers: tritium, the helium isotope ratio of dissolved helium and the concentrations of dissolved noble gases (Helium, Neon, Argon, Krypton, and Xenon). A subset of the samples was also analyzed for excess nitrogen due to saturated zone denitrification. The age-dating data were used to evaluate the degree to which groundwater at a particular monitoring well was derived from pre-modern and/or modern sources. More specifically, the analyses canmore » be used to determine whether the recharge age of the groundwater beneath the site pre-dates anthropogenic activities at the site.« less

Docking analysis targeted to the whole enzyme: an application to the prediction of inhibition of PTP1B by thiomorpholine and thiazolyl derivatives.

PubMed

Ganou, C A; Eleftheriou, P Th; Theodosis-Nobelos, P; Fesatidou, M; Geronikaki, A A; Lialiaris, T; Rekka, E A

2018-02-01

PTP1b is a protein tyrosine phosphatase involved in the inactivation of insulin receptor. Since inhibition of PTP1b may prolong the action of the receptor, PTP1b has become a drug target for the treatment of type II diabetes. In the present study, prediction of inhibition using docking analysis targeted specifically to the active or allosteric site was performed on 87 compounds structurally belonging to 10 different groups. Two groups, consisting of 15 thiomorpholine and 10 thiazolyl derivatives exhibiting the best prediction results, were selected for in vitro evaluation. All thiomorpholines showed inhibitory action (with IC 50 = 4-45 μΜ, Ki = 2-23 μM), while only three thiazolyl derivatives showed low inhibition (best IC 50 = 18 μΜ, Ki = 9 μΜ). However, free binding energy (E) was in accordance with the IC 50 values only for some compounds. Docking analysis targeted to the whole enzyme revealed that the compounds exhibiting IC 50 values higher than expected could bind to other peripheral sites with lower free energy, E o , than when bound to the active/allosteric site. A prediction factor, E- (Σ Eo × 0.16), which takes into account lower energy binding to peripheral sites, was proposed and was found to correlate well with the IC 50 values following an asymmetrical sigmoidal equation with r 2 = 0.9692.

Prediction of proprotein convertase cleavage sites.

PubMed

Duckert, Peter; Brunak, Søren; Blom, Nikolaj

2004-01-01

Many secretory proteins and peptides are synthesized as inactive precursors that in addition to signal peptide cleavage undergo post-translational processing to become biologically active polypeptides. Precursors are usually cleaved at sites composed of single or paired basic amino acid residues by members of the subtilisin/kexin-like proprotein convertase (PC) family. In mammals, seven members have been identified, with furin being the one first discovered and best characterized. Recently, the involvement of furin in diseases ranging from Alzheimer's disease and cancer to anthrax and Ebola fever has created additional focus on proprotein processing. We have developed a method for prediction of cleavage sites for PCs based on artificial neural networks. Two different types of neural networks have been constructed: a furin-specific network based on experimental results derived from the literature, and a general PC-specific network trained on data from the Swiss-Prot protein database. The method predicts cleavage sites in independent sequences with a sensitivity of 95% for the furin neural network and 62% for the general PC network. The ProP method is made publicly available at http://www.cbs.dtu.dk/services/ProP.

In vivo modification of a maize engineered minichromosome.

PubMed

Gaeta, Robert T; Masonbrink, Rick E; Zhao, Changzeng; Sanyal, Abhijit; Krishnaswamy, Lakshminarasimhan; Birchler, James A

2013-06-01

Engineered minichromosomes provide efficient platforms for stacking transgenes in crop plants. Methods for modifying these chromosomes in vivo are essential for the development of customizable systems for the removal of selection genes or other sequences and for the addition of new genes. Previous studies have demonstrated that Cre, a site-specific recombinase, could be used to modify lox sites on transgenes on maize minichromosomes; however, these studies demonstrated somatic recombination only, and modified minichromosomes could not be recovered. We describe the recovery of an engineered chromosome composed of little more than a centromere plus transgene that was derived by telomere-mediated truncation. We used the fiber fluorescence in situ hybridization technique and detected a transgene on the minichromosome inserted among stretches of CentC centromere repeats, and this insertion was large enough to suggest a tandem insertion. By crossing the minichromosome to a plant expressing Cre-recombinase, the Bar selection gene was removed, leaving behind a single loxP site. This study demonstrates that engineered chromosomes can be modified in vivo using site-specific recombinases, a demonstration essential to the development of amendable chromosome platforms in plants.

Active site electrostatics protect genome integrity by blocking abortive hydrolysis during DNA recombination

PubMed Central

Ma, Chien-Hui; Rowley, Paul A; Macieszak, Anna; Guga, Piotr; Jayaram, Makkuni

2009-01-01

Water, acting as a rogue nucleophile, can disrupt transesterification steps of important phosphoryl transfer reactions in DNA and RNA. We have unveiled this risk, and identified safeguards instituted against it, during strand cleavage and joining by the tyrosine site-specific recombinase Flp. Strand joining is threatened by a latent Flp endonuclease activity (type I) towards the 3′-phosphotyrosyl intermediate resulting from strand cleavage. This risk is not alleviated by phosphate electrostatics; neutralizing the negative charge on the scissile phosphate through methylphosphonate (MeP) substitution does not stimulate type I endonuclease. Rather, protection derives from the architecture of the recombination synapse and conformational dynamics within it. Strand cleavage is protected against water by active site electrostatics. Replacement of the catalytic Arg-308 of Flp by alanine, along with MeP substitution, elicits a second Flp endonuclease activity (type II) that directly targets the scissile phosphodiester bond in DNA. MeP substitution, combined with appropriate active site mutations, will be useful in revealing anti-hydrolytic mechanisms engendered by systems that mediate DNA relaxation, DNA transposition, site-specific recombination, telomere resolution, RNA splicing and retrohoming of mobile introns. PMID:19440204

Thermodynamic Characterization of Hydration Sites from Integral Equation-Derived Free Energy Densities: Application to Protein Binding Sites and Ligand Series.

PubMed

Güssregen, Stefan; Matter, Hans; Hessler, Gerhard; Lionta, Evanthia; Heil, Jochen; Kast, Stefan M

2017-07-24

Water molecules play an essential role for mediating interactions between ligands and protein binding sites. Displacement of specific water molecules can favorably modulate the free energy of binding of protein-ligand complexes. Here, the nature of water interactions in protein binding sites is investigated by 3D RISM (three-dimensional reference interaction site model) integral equation theory to understand and exploit local thermodynamic features of water molecules by ranking their possible displacement in structure-based design. Unlike molecular dynamics-based approaches, 3D RISM theory allows for fast and noise-free calculations using the same detailed level of solute-solvent interaction description. Here we correlate molecular water entities instead of mere site density maxima with local contributions to the solvation free energy using novel algorithms. Distinct water molecules and hydration sites are investigated in multiple protein-ligand X-ray structures, namely streptavidin, factor Xa, and factor VIIa, based on 3D RISM-derived free energy density fields. Our approach allows the semiquantitative assessment of whether a given structural water molecule can potentially be targeted for replacement in structure-based design. Finally, PLS-based regression models from free energy density fields used within a 3D-QSAR approach (CARMa - comparative analysis of 3D RISM Maps) are shown to be able to extract relevant information for the interpretation of structure-activity relationship (SAR) trends, as demonstrated for a series of serine protease inhibitors.

Psychotomimetic opiate receptors labeled and visualized with (+)-(/sup 3/H)3-(3-hydroxyphenyl)-N-(1-propyl)piperidine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Largent, B.L.; Gundlach, A.L.; Snyder, S.H.

1984-08-01

3-(3-Hydroxyphenyl)-N-(1-propyl)piperidine (3-PPP) has been proposed as a selective dopamine autoreceptor agonist in the central nervous system. This report describes the pharmacology and localization of specific high-affinity binding sites for (+)-(/sup 3/H)3-PPP in brain. The drug specificity of (+)-(/sup 3/H)3-PPP binding is identical to that of sigma receptors, which may mediate psychotomimetic effects of some opiates. Haloperidol and the opioid derivatives, pentazocine, cyclazocine, and SKF 10,047 are potent inhibitors of (+)-(/sup 3/H)3-PPP binding. Stereoselectivity is exhibited for the (+) isomers of cyclazocine and SKF 10.047 at the sigma site, opposite to the stereoselectivity seen at ..mu.., sigma, and k opiate receptors.more » (+)-(/sup 3/H)3-PPP does not label dopamine receptors, as potent dopamine agonists and antagonists are weak inhibitors of binding and the localization of specific (+)-(/sup 3/H)3-PPP binding sites does not parallel that of dopamine neurons. Discrete localizations of (+)-(/sup 3/H)3-PPP binding sites in many brain areas including limbic, midbrain, brainstem, and cerebellar regions may explain psychotomimetic actions of opiates and behavior effects of 3-PPP. 41 references, 2 figures, 1 table.« less

Multi-element compound specific stable isotope analysis of chlorinated aliphatic contaminants derived from chlorinated pitches.

PubMed

Filippini, Maria; Nijenhuis, Ivonne; Kümmel, Steffen; Chiarini, Veronica; Crosta, Giovanni; Richnow, Hans H; Gargini, Alessandro

2018-05-30

Tetrachloroethene and trichloroethene are typical by-products of the industrial production of chloromethanes. These by-products are known as "chlorinated pitches" and were often dumped in un-contained waste disposal sites causing groundwater contaminations. Previous research showed that a strongly depleted stable carbon isotope signature characterizes chlorinated compounds associated with chlorinated pitches whereas manufactured commercial compounds have more enriched carbon isotope ratios. The findings were restricted to a single case study and one element (i.e. carbon). This paper presents a multi-element Compound-Specific Stable Isotope Analysis (CSIA, including carbon, chlorine and hydrogen) of chlorinated aliphatic contaminants originated from chlorinated pitches at two sites with different hydrogeology and different producers of chloromethanes. The results show strongly depleted carbon signatures at both sites whereas the chlorine and the hydrogen signatures are comparable to those presented in the literature for manufactured commercial compounds. Multi-element CSIA allowed the identification of sources and site-specific processes affecting chloroethene transformation in groundwater as a result of emergency remediation measures. CSIA turned out to be an effective forensic tool to address the liability for the contamination, leading to a conviction for the crimes of unintentional aggravated public water supply poisoning and environmental disaster. Copyright © 2018 Elsevier B.V. All rights reserved.

Can camera traps monitor Komodo dragons a large ectothermic predator?

PubMed

Ariefiandy, Achmad; Purwandana, Deni; Seno, Aganto; Ciofi, Claudio; Jessop, Tim S

2013-01-01

Camera trapping has greatly enhanced population monitoring of often cryptic and low abundance apex carnivores. Effectiveness of passive infrared camera trapping, and ultimately population monitoring, relies on temperature mediated differences between the animal and its ambient environment to ensure good camera detection. In ectothermic predators such as large varanid lizards, this criterion is presumed less certain. Here we evaluated the effectiveness of camera trapping to potentially monitor the population status of the Komodo dragon (Varanus komodoensis), an apex predator, using site occupancy approaches. We compared site-specific estimates of site occupancy and detection derived using camera traps and cage traps at 181 trapping locations established across six sites on four islands within Komodo National Park, Eastern Indonesia. Detection and site occupancy at each site were estimated using eight competing models that considered site-specific variation in occupancy (ψ)and varied detection probabilities (p) according to detection method, site and survey number using a single season site occupancy modelling approach. The most parsimonious model [ψ (site), p (site survey); ω = 0.74] suggested that site occupancy estimates differed among sites. Detection probability varied as an interaction between site and survey number. Our results indicate that overall camera traps produced similar estimates of detection and site occupancy to cage traps, irrespective of being paired, or unpaired, with cage traps. Whilst one site showed some evidence detection was affected by trapping method detection was too low to produce an accurate occupancy estimate. Overall, as camera trapping is logistically more feasible it may provide, with further validation, an alternative method for evaluating long-term site occupancy patterns in Komodo dragons, and potentially other large reptiles, aiding conservation of this species.

Can Camera Traps Monitor Komodo Dragons a Large Ectothermic Predator?

PubMed Central

Ariefiandy, Achmad; Purwandana, Deni; Seno, Aganto; Ciofi, Claudio; Jessop, Tim S.

2013-01-01

Camera trapping has greatly enhanced population monitoring of often cryptic and low abundance apex carnivores. Effectiveness of passive infrared camera trapping, and ultimately population monitoring, relies on temperature mediated differences between the animal and its ambient environment to ensure good camera detection. In ectothermic predators such as large varanid lizards, this criterion is presumed less certain. Here we evaluated the effectiveness of camera trapping to potentially monitor the population status of the Komodo dragon (Varanus komodoensis), an apex predator, using site occupancy approaches. We compared site-specific estimates of site occupancy and detection derived using camera traps and cage traps at 181 trapping locations established across six sites on four islands within Komodo National Park, Eastern Indonesia. Detection and site occupancy at each site were estimated using eight competing models that considered site-specific variation in occupancy (ψ)and varied detection probabilities (p) according to detection method, site and survey number using a single season site occupancy modelling approach. The most parsimonious model [ψ (site), p (site*survey); ω = 0.74] suggested that site occupancy estimates differed among sites. Detection probability varied as an interaction between site and survey number. Our results indicate that overall camera traps produced similar estimates of detection and site occupancy to cage traps, irrespective of being paired, or unpaired, with cage traps. Whilst one site showed some evidence detection was affected by trapping method detection was too low to produce an accurate occupancy estimate. Overall, as camera trapping is logistically more feasible it may provide, with further validation, an alternative method for evaluating long-term site occupancy patterns in Komodo dragons, and potentially other large reptiles, aiding conservation of this species. PMID:23527027

Array-Based Rational Design of Short Peptide Probe-Derived from an Anti-TNT Monoclonal Antibody.

PubMed

Okochi, Mina; Muto, Masaki; Yanai, Kentaro; Tanaka, Masayoshi; Onodera, Takeshi; Wang, Jin; Ueda, Hiroshi; Toko, Kiyoshi

2017-10-09

Complementarity-determining regions (CDRs) are sites on the variable chains of antibodies responsible for binding to specific antigens. In this study, a short peptide probe for recognition of 2,4,6-trinitrotoluene (TNT), was identified by testing sequences derived from the CDRs of an anti-TNT monoclonal antibody. The major TNT-binding site in this antibody was identified in the heavy chain CDR3 by antigen docking simulation and confirmed by an immunoassay using a spot-synthesis based peptide array comprising amino acid sequences of six CDRs in the variable region. A peptide derived from heavy chain CDR3 (RGYSSFIYWF) bound to TNT with a dissociation constant of 1.3 μM measured by surface plasmon resonance. Substitution of selected amino acids with basic residues increased TNT binding while substitution with acidic amino acids decreased affinity, an isoleucine to arginine change showed the greatest improvement of 1.8-fold. The ability to create simple peptide binders of volatile organic compounds from sequence information provided by the immune system in the creation of an immune response will be beneficial for sensor developments in the future.

Oxime amides as a novel zinc binding group in histone deacetylase inhibitors: synthesis, biological activity, and computational evaluation.

PubMed

Botta, Cinzia B; Cabri, Walter; Cini, Elena; De Cesare, Lucia; Fattorusso, Caterina; Giannini, Giuseppe; Persico, Marco; Petrella, Antonello; Rondinelli, Francesca; Rodriquez, Manuela; Russo, Adele; Taddei, Maurizio

2011-04-14

Several oxime containing molecules, characterized by a SAHA-like structure, were explored to select a potentially new biasing binding element for the zinc in HDAC catalytic site. All compounds were evaluated for their in vitro inhibitory activity against the 11 human HDACs isoforms. After identification of a "hit" molecule, a programmed variation at the cap group and at the linker was carried out in order to increase HDAC inhibition and/or paralogue selectivity. Some of the new derivatives showed increased activity against a number of HDAC isoforms, even if their overall activity range is still far from the inhibition values reported for SAHA. Moreover, different from what was reported for their hydroxamic acid analogues the new α-oxime amide derivatives do not select between class I and class II HDACs; rather they target specific isoforms in each class. These somehow contradictory results were finally rationalized by a computational assisted SAR, which gave us the chance to understand how the oxime derivatives interact with the catalytic site and justify the observed activity profile.

Characterization of IntA, a Bidirectional Site-Specific Recombinase Required for Conjugative Transfer of the Symbiotic Plasmid of Rhizobium etli CFN42

PubMed Central

Hernández-Tamayo, Rogelio; Sohlenkamp, Christian; Puente, José Luis; Brom, Susana

2013-01-01

Site-specific recombination occurs at short specific sequences, mediated by the cognate recombinases. IntA is a recombinase from Rhizobium etli CFN42 and belongs to the tyrosine recombinase family. It allows cointegration of plasmid p42a and the symbiotic plasmid via site-specific recombination between attachment regions (attA and attD) located in each replicon. Cointegration is needed for conjugative transfer of the symbiotic plasmid. To characterize this system, two plasmids harboring the corresponding attachment sites and intA were constructed. Introduction of these plasmids into R. etli revealed IntA-dependent recombination events occurring at high frequency. Interestingly, IntA promotes not only integration, but also excision events, albeit at a lower frequency. Thus, R. etli IntA appears to be a bidirectional recombinase. IntA was purified and used to set up electrophoretic mobility shift assays with linear fragments containing attA and attD. IntA-dependent retarded complexes were observed only with fragments containing either attA or attD. Specific retarded complexes, as well as normal in vivo recombination abilities, were seen even in derivatives harboring only a minimal attachment region (comprising the 5-bp central region flanked by 9- to 11-bp inverted repeats). DNase I-footprinting assays with IntA revealed specific protection of these zones. Mutations that disrupt the integrity of the 9- to 11-bp inverted repeats abolish both specific binding and recombination ability, while mutations in the 5-bp central region severely reduce both binding and recombination. These results show that IntA is a bidirectional recombinase that binds to att regions without requiring neighboring sequences as enhancers of recombination. PMID:23935046

Efficient DNA binding and nuclear uptake by distamycin derivatives conjugated to octa-arginine sequences.

PubMed

Vázquez, Olalla; Blanco-Canosa, Juan B; Vázquez, M Eugenio; Martínez-Costas, Jose; Castedo, Luis; Mascareñas, José L

2008-11-24

Efficient targeting of DNA by designed molecules requires not only careful fine-tuning of their DNA-recognition properties, but also appropriate cell internalization of the compounds so that they can reach the cell nucleus in a short period of time. Previous observations in our group on the relatively high affinity displayed by conjugates between distamycin derivatives and bZIP basic regions for A-rich DNA sites, led us to investigate whether the covalent attachment of a positively charged cell-penetrating peptide to a distamycin-like tripyrrole might yield high affinity DNA binders with improved cell internalization properties. Our work has led to the discovery of synthetic tripyrrole-octa-arginine conjugates that are capable of targeting specific DNA sites that contain A-rich tracts with low nanomolar affinity; they simultaneously exhibit excellent membrane and nuclear translocation properties in living HeLa cells.

Design of novel HIV-1 protease inhibitors incorporating isophthalamide-derived P2-P3 ligands: Synthesis, biological evaluation and X-ray structural studies of inhibitor-HIV-1 protease complex

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ghosh, Arun K.; Brindisi, Margherita; Nyalapatla, Prasanth R.

Based upon molecular insights from the X-ray structures of inhibitor-bound HIV-1 protease complexes, we have designed a series of isophthalamide-derived inhibitors incorporating substituted pyrrolidines, piperidines and thiazolidines as P2-P3 ligands for specific interactions in the S2-S3 extended site. Compound 4b has shown an enzyme Ki of 0.025 nM and antiviral IC50 of 69 nM. An X-ray crystal structure of inhibitor 4b-HIV-1 protease complex was determined at 1.33 Å resolution. We have also determined X-ray structure of 3b-bound HIV-1 protease at 1.27 Å resolution. These structures revealed important molecular insight into the inhibitor–HIV-1 protease interactions in the active site.

Trigger values for investigation of hormonal activity in drinking water and its sources using CALUX bioassays.

PubMed

Brand, Walter; de Jongh, Cindy M; van der Linden, Sander C; Mennes, Wim; Puijker, Leo M; van Leeuwen, Cornelis J; van Wezel, Annemarie P; Schriks, Merijn; Heringa, Minne B

2013-05-01

To screen for hormonal activity in water samples, highly sensitive in vitro CALUX bioassays are available which allow detection of estrogenic (ERα), androgenic (AR), progestagenic (PR), and glucocorticoid (GR) activities. This paper presents trigger values for the ERα, AR, PR, and GR CALUX bioassays for agonistic hormonal activities in (drinking) water, which define a level above which human health risk cannot be waived a priori and additional examination of specific endocrine activity may be warranted. The trigger values are based on 1) acceptable or tolerable daily intake (ADI/TDI) values of specific compounds, 2) pharmacokinetic factors defining their bioavailability, 3) estimations of the bioavailability of unknown compounds with equivalent hormonal activity, 4) relative endocrine potencies, and 5) physiological, and drinking water allocation factors. As a result, trigger values of 3.8ng 17β-estradiol (E2)-equivalents (eq)/L, 11ng dihydrotestosterone (DHT)-eq/L, 21ng dexamethasone (DEX)-eq/L, and 333ng Org2058-eq/L were derived. Benchmark Quotient (BQ) values were derived by dividing hormonal activity in water samples by the derived trigger using the highest concentrations detected in a recent, limited screening of Dutch water samples, and were in the order of (value) AR (0.41)>ERα (0.13)>GR (0.06)>PR (0.04). The application of trigger values derived in the present study can help to judge measured agonistic hormonal activities in water samples using the CALUX bioassays and help to decide whether further examination of specific endocrine activity followed by a subsequent safety evaluation may be warranted, or whether concentrations of such activity are of low priority with respect to health concerns in the human population. For instance, at one specific drinking water production site ERα and AR (but no GR and PR) activities were detected in drinking water, however, these levels are at least a factor 83 smaller than the respective trigger values, and therefore no human health risks are to be expected from hormonal activity in Dutch drinking water from this site. Copyright © 2013 Elsevier Ltd. All rights reserved.

Protein 3-Nitrotyrosine in Complex Biological Samples: Quantification by High-Pressure Liquid Chromatography/Electrochemical Detection and Emergence of Proteomic Approaches for Unbiased Identification of Modification Sites

PubMed Central

Nuriel, Tal; Deeb, Ruba S.; Hajjar, David P.; Gross, Steven S.

2008-01-01

Nitration of tyrosine residues by nitric oxide (NO)-derived species results in the accumulation of 3-nitrotyrosine in proteins, a hallmark of nitrosative stress in cells and tissues. Tyrosine nitration is recognized as one of the multiple signaling modalities used by NO-derived species for the regulation of protein structure and function in health and disease. Various methods have been described for the quantification of protein 3-nitrotyrosine residues, and several strategies have been presented toward the goal of proteome-wide identification of protein tyrosine modification sites. This chapter details a useful protocol for the quantification of 3-nitrotyrosine in cells and tissues using high-pressure liquid chromatography with electrochemical detection. Additionally, this chapter describes a novel biotin-tagging strategy for specific enrichment of 3-nitrotyrosine-containing peptides. Application of this strategy, in conjunction with high-throughput MS/MS-based peptide sequencing, is anticipated to fuel efforts in developing comprehensive inventories of nitrosative stress-induced protein-tyrosine modification sites in cells and tissues. PMID:18554526

Proposal for novel curcumin derivatives as potent inhibitors against Alzheimer's disease: Ab initio molecular simulations on the specific interactions between amyloid-beta peptide and curcumin

NASA Astrophysics Data System (ADS)

Ota, Shintaro; Fujimori, Mitsuki; Ishimura, Hiromi; Shulga, Sergiy; Kurita, Noriyuki

2017-10-01

Accumulation of amyloid-β (Aβ) peptides in a brain is closely related with the pathogenesis of Alzheimer's disease. To suppress the production of Aβ peptides, we propose novel curcumin derivatives and investigate their binding properties with the amyloid precursor protein (APP), using protein-ligand docking as well as ab initio molecular simulations. Our proposed derivative (curcumin XIV) is found to have a large binding energy with APP and interacts strongly with the cleavage site Ala19 by secretase. It is thus expected that curcumin XIV can protect APP from the secretase attack and be a potent inhibitor against the production of Aβ peptides.

Redundancy of primary RNA-binding functions of the bacterial transcription terminator Rho

PubMed Central

Shashni, Rajesh; Qayyum, M. Zuhaib; Vishalini, V.; Dey, Debashish; Sen, Ranjan

2014-01-01

The bacterial transcription terminator, Rho, terminates transcription at half of the operons. According to the classical model derived from in vitro assays on a few terminators, Rho is recruited to the transcription elongation complex (EC) by recognizing specific sites (rut) on the nascent RNA. Here, we explored the mode of in vivo recruitment process of Rho. We show that sequence specific recognition of the rut site, in majority of the Rho-dependent terminators, can be compromised to a great extent without seriously affecting the genome-wide termination function as well as the viability of Escherichia coli. These terminators function optimally only through a NusG-assisted recruitment and activation of Rho. Our data also indicate that at these terminators, Rho-EC-bound NusG interaction facilitates the isomerization of Rho into a translocase-competent form by stabilizing the interactions of mRNA with the secondary RNA binding site, thereby overcoming the defects of the primary RNA binding functions. PMID:25081210

Transposable Elements and DNA Methylation Create in Embryonic Stem Cells Human-Specific Regulatory Sequences Associated with Distal Enhancers and Noncoding RNAs

PubMed Central

Glinsky, Gennadi V.

2015-01-01

Despite significant progress in the structural and functional characterization of the human genome, understanding of the mechanisms underlying the genetic basis of human phenotypic uniqueness remains limited. Here, I report that transposable element-derived sequences, most notably LTR7/HERV-H, LTR5_Hs, and L1HS, harbor 99.8% of the candidate human-specific regulatory loci (HSRL) with putative transcription factor-binding sites in the genome of human embryonic stem cells (hESC). A total of 4,094 candidate HSRL display selective and site-specific binding of critical regulators (NANOG [Nanog homeobox], POU5F1 [POU class 5 homeobox 1], CCCTC-binding factor [CTCF], Lamin B1), and are preferentially located within the matrix of transcriptionally active DNA segments that are hypermethylated in hESC. hESC-specific NANOG-binding sites are enriched near the protein-coding genes regulating brain size, pluripotency long noncoding RNAs, hESC enhancers, and 5-hydroxymethylcytosine-harboring regions immediately adjacent to binding sites. Sequences of only 4.3% of hESC-specific NANOG-binding sites are present in Neanderthals’ genome, suggesting that a majority of these regulatory elements emerged in Modern Humans. Comparisons of estimated creation rates of novel TF-binding sites revealed that there was 49.7-fold acceleration of creation rates of NANOG-binding sites in genomes of Chimpanzees compared with the mouse genomes and further 5.7-fold acceleration in genomes of Modern Humans compared with the Chimpanzees genomes. Preliminary estimates suggest that emergence of one novel NANOG-binding site detectable in hESC required 466 years of evolution. Pathway analysis of coding genes that have hESC-specific NANOG-binding sites within gene bodies or near gene boundaries revealed their association with physiological development and functions of nervous and cardiovascular systems, embryonic development, behavior, as well as development of a diverse spectrum of pathological conditions such as cancer, diseases of cardiovascular and reproductive systems, metabolic diseases, multiple neurological and psychological disorders. A proximity placement model is proposed explaining how a 33–47% excess of NANOG, CTCF, and POU5F1 proteins immobilized on a DNA scaffold may play a functional role at distal regulatory elements. PMID:25956794

Long-range electrostatic complementarity governs substrate recognition by human chymotrypsin C, a key regulator of digestive enzyme activation.

PubMed

Batra, Jyotica; Szabó, András; Caulfield, Thomas R; Soares, Alexei S; Sahin-Tóth, Miklós; Radisky, Evette S

2013-04-05

Human chymotrypsin C (CTRC) is a pancreatic serine protease that regulates activation and degradation of trypsinogens and procarboxypeptidases by targeting specific cleavage sites within their zymogen precursors. In cleaving these regulatory sites, which are characterized by multiple flanking acidic residues, CTRC shows substrate specificity that is distinct from that of other isoforms of chymotrypsin and elastase. Here, we report the first crystal structure of active CTRC, determined at 1.9-Å resolution, revealing the structural basis for binding specificity. The structure shows human CTRC bound to the small protein protease inhibitor eglin c, which binds in a substrate-like manner filling the S6-S5' subsites of the substrate binding cleft. Significant binding affinity derives from burial of preferred hydrophobic residues at the P1, P4, and P2' positions of CTRC, although acidic P2' residues can also be accommodated by formation of an interfacial salt bridge. Acidic residues may also be specifically accommodated in the P6 position. The most unique structural feature of CTRC is a ring of intense positive electrostatic surface potential surrounding the primarily hydrophobic substrate binding site. Our results indicate that long-range electrostatic attraction toward substrates of concentrated negative charge governs substrate discrimination, which explains CTRC selectivity in regulating active digestive enzyme levels.

A reference skeletal dosimetry model for an adult male radionuclide therapy patient based on three-dimensional imaging and paired-image radiation transport

NASA Astrophysics Data System (ADS)

Shah, Amish P.

The need for improved patient-specificity of skeletal dose estimates is widely recognized in radionuclide therapy. Current clinical models for marrow dose are based on skeletal mass estimates from a variety of sources and linear chord-length distributions that do not account for particle escape into cortical bone. To predict marrow dose, these clinical models use a scheme that requires separate calculations of cumulated activity and radionuclide S values. Selection of an appropriate S value is generally limited to one of only three sources, all of which use as input the trabecular microstructure of an individual measured 25 years ago, and the tissue mass derived from different individuals measured 75 years ago. Our study proposed a new modeling approach to marrow dosimetry---the Paired Image Radiation Transport (PIRT) model---that properly accounts for both the trabecular microstructure and the cortical macrostructure of each skeletal site in a reference male radionuclide patient. The PIRT model, as applied within EGSnrc, requires two sets of input geometry: (1) an infinite voxel array of segmented microimages of the spongiosa acquired via microCT; and (2) a segmented ex-vivo CT image of the bone site macrostructure defining both the spongiosa (marrow, endosteum, and trabeculae) and the cortical bone cortex. Our study also proposed revising reference skeletal dosimetry models for the adult male cancer patient. Skeletal site-specific radionuclide S values were obtained for a 66-year-old male reference patient. The derivation for total skeletal S values were unique in that the necessary skeletal mass and electron dosimetry calculations were formulated from the same source bone site over the entire skeleton. We conclude that paired-image radiation-transport techniques provide an adoptable method by which the intricate, anisotropic trabecular microstructure of the skeletal site; and the physical size and shape of the bone can be handled together, for improved compilation of reference radionuclide S values. We also conclude that this comprehensive model for the adult male cancer patient should be implemented for use in patient-specific calculations for radionuclide dosimetry of the skeleton.

Multi-site precipitation downscaling using a stochastic weather generator

NASA Astrophysics Data System (ADS)

Chen, Jie; Chen, Hua; Guo, Shenglian

2018-03-01

Statistical downscaling is an efficient way to solve the spatiotemporal mismatch between climate model outputs and the data requirements of hydrological models. However, the most commonly-used downscaling method only produces climate change scenarios for a specific site or watershed average, which is unable to drive distributed hydrological models to study the spatial variability of climate change impacts. By coupling a single-site downscaling method and a multi-site weather generator, this study proposes a multi-site downscaling approach for hydrological climate change impact studies. Multi-site downscaling is done in two stages. The first stage involves spatially downscaling climate model-simulated monthly precipitation from grid scale to a specific site using a quantile mapping method, and the second stage involves the temporal disaggregating of monthly precipitation to daily values by adjusting the parameters of a multi-site weather generator. The inter-station correlation is specifically considered using a distribution-free approach along with an iterative algorithm. The performance of the downscaling approach is illustrated using a 10-station watershed as an example. The precipitation time series derived from the National Centers for Environment Prediction (NCEP) reanalysis dataset is used as the climate model simulation. The precipitation time series of each station is divided into 30 odd years for calibration and 29 even years for validation. Several metrics, including the frequencies of wet and dry spells and statistics of the daily, monthly and annual precipitation are used as criteria to evaluate the multi-site downscaling approach. The results show that the frequencies of wet and dry spells are well reproduced for all stations. In addition, the multi-site downscaling approach performs well with respect to reproducing precipitation statistics, especially at monthly and annual timescales. The remaining biases mainly result from the non-stationarity of NCEP precipitation. Overall, the proposed approach is efficient for generating multi-site climate change scenarios that can be used to investigate the spatial variability of climate change impacts on hydrology.

Exploring Covalent Allosteric Inhibition of Antigen 85C from Mycobacterium tuberculosis by Ebselen Derivatives

DOE Office of Scientific and Technical Information (OSTI.GOV)

Goins, Christopher M.; Dajnowicz, Steven; Thanna, Sandeep

Previous studies identified ebselen as a potent in vitro and in vivo inhibitor of the Mycobacterium tuberculosis ( Mtb) antigen 85 (Ag85) complex, comprising three homologous enzymes required for the biosynthesis of the mycobacterial cell wall. In this study, the Mtb Ag85C enzyme was cocrystallized with azido and adamantyl ebselen derivatives, resulting in two crystallographic structures of 2.01 and 1.30 Å resolution, respectively. Both structures displayed the anticipated covalent modification of the solvent accessible, noncatalytic Cys209 residue forming a selenenylsulfide bond. Continuous difference density for both thiol modifiers allowed for the assessment of interactions that influence ebselen binding and inhibitormore » orientation that were unobserved in previous Ag85C ebselen structures. The k inact/ K I values for ebselen, adamantyl ebselen, and azido ebselen support the importance of observed constructive chemical interactions with Arg239 for increased in vitro efficacy toward Ag85C. To better understand the in vitro kinetic properties of these ebselen derivatives, the energetics of specific protein–inhibitor interactions and relative reaction free energies were calculated for ebselen and both derivatives using density functional theory. These studies further support the different in vitro properties of ebselen and two select ebselen derivatives from our previously published ebselen library with respect to kinetics and protein–inhibitor interactions. In both structures, the α9 helix was displaced farther from the enzyme active site than the previous Ag85C ebselen structure, resulting in the restructuring of a connecting loop and imparting a conformational change to residues believed to play a role in substrate binding specific to Ag85C. These notable structural changes directly affect protein stability, reducing the overall melting temperature by up to 14.5 °C, resulting in the unfolding of protein at physiological temperatures. Additionally, this structural rearrangement due to covalent allosteric modification creates a sizable solvent network that encompasses the active site and extends to the modified Cys209 residue. In all, this study outlines factors that influence enzyme inhibition by ebselen and its derivatives while further highlighting the effects of the covalent modification of Cys209 by said inhibitors on the structure and stability of Ag85C. Moreover, the results suggest a strategy for developing new classes of Ag85 inhibitors with increased specificity and potency.« less

Exploring Covalent Allosteric Inhibition of Antigen 85C from Mycobacterium tuberculosis by Ebselen Derivatives.

PubMed

Goins, Christopher M; Dajnowicz, Steven; Thanna, Sandeep; Sucheck, Steven J; Parks, Jerry M; Ronning, Donald R

2017-05-12

Previous studies identified ebselen as a potent in vitro and in vivo inhibitor of the Mycobacterium tuberculosis (Mtb) antigen 85 (Ag85) complex, comprising three homologous enzymes required for the biosynthesis of the mycobacterial cell wall. In this study, the Mtb Ag85C enzyme was cocrystallized with azido and adamantyl ebselen derivatives, resulting in two crystallographic structures of 2.01 and 1.30 Å resolution, respectively. Both structures displayed the anticipated covalent modification of the solvent accessible, noncatalytic Cys209 residue forming a selenenylsulfide bond. Continuous difference density for both thiol modifiers allowed for the assessment of interactions that influence ebselen binding and inhibitor orientation that were unobserved in previous Ag85C ebselen structures. The k inact /K I values for ebselen, adamantyl ebselen, and azido ebselen support the importance of observed constructive chemical interactions with Arg239 for increased in vitro efficacy toward Ag85C. To better understand the in vitro kinetic properties of these ebselen derivatives, the energetics of specific protein-inhibitor interactions and relative reaction free energies were calculated for ebselen and both derivatives using density functional theory. These studies further support the different in vitro properties of ebselen and two select ebselen derivatives from our previously published ebselen library with respect to kinetics and protein-inhibitor interactions. In both structures, the α9 helix was displaced farther from the enzyme active site than the previous Ag85C ebselen structure, resulting in the restructuring of a connecting loop and imparting a conformational change to residues believed to play a role in substrate binding specific to Ag85C. These notable structural changes directly affect protein stability, reducing the overall melting temperature by up to 14.5 °C, resulting in the unfolding of protein at physiological temperatures. Additionally, this structural rearrangement due to covalent allosteric modification creates a sizable solvent network that encompasses the active site and extends to the modified Cys209 residue. In all, this study outlines factors that influence enzyme inhibition by ebselen and its derivatives while further highlighting the effects of the covalent modification of Cys209 by said inhibitors on the structure and stability of Ag85C. Furthermore, the results suggest a strategy for developing new classes of Ag85 inhibitors with increased specificity and potency.

Exploring Covalent Allosteric Inhibition of Antigen 85C from Mycobacterium tuberculosis by Ebselen Derivatives

DOE PAGES

Goins, Christopher M.; Dajnowicz, Steven; Thanna, Sandeep; ...

2017-03-13

Previous studies identified ebselen as a potent in vitro and in vivo inhibitor of the Mycobacterium tuberculosis ( Mtb) antigen 85 (Ag85) complex, comprising three homologous enzymes required for the biosynthesis of the mycobacterial cell wall. In this study, the Mtb Ag85C enzyme was cocrystallized with azido and adamantyl ebselen derivatives, resulting in two crystallographic structures of 2.01 and 1.30 Å resolution, respectively. Both structures displayed the anticipated covalent modification of the solvent accessible, noncatalytic Cys209 residue forming a selenenylsulfide bond. Continuous difference density for both thiol modifiers allowed for the assessment of interactions that influence ebselen binding and inhibitormore » orientation that were unobserved in previous Ag85C ebselen structures. The k inact/ K I values for ebselen, adamantyl ebselen, and azido ebselen support the importance of observed constructive chemical interactions with Arg239 for increased in vitro efficacy toward Ag85C. To better understand the in vitro kinetic properties of these ebselen derivatives, the energetics of specific protein–inhibitor interactions and relative reaction free energies were calculated for ebselen and both derivatives using density functional theory. These studies further support the different in vitro properties of ebselen and two select ebselen derivatives from our previously published ebselen library with respect to kinetics and protein–inhibitor interactions. In both structures, the α9 helix was displaced farther from the enzyme active site than the previous Ag85C ebselen structure, resulting in the restructuring of a connecting loop and imparting a conformational change to residues believed to play a role in substrate binding specific to Ag85C. These notable structural changes directly affect protein stability, reducing the overall melting temperature by up to 14.5 °C, resulting in the unfolding of protein at physiological temperatures. Additionally, this structural rearrangement due to covalent allosteric modification creates a sizable solvent network that encompasses the active site and extends to the modified Cys209 residue. In all, this study outlines factors that influence enzyme inhibition by ebselen and its derivatives while further highlighting the effects of the covalent modification of Cys209 by said inhibitors on the structure and stability of Ag85C. Moreover, the results suggest a strategy for developing new classes of Ag85 inhibitors with increased specificity and potency.« less

Chimpanzee-Human Monoclonal Antibodies for Treatment of Chronic Poliovirus Excretors and Emergency Postexposure Prophylaxis▿‡

PubMed Central

Chen, Zhaochun; Chumakov, Konstantin; Dragunsky, Eugenia; Kouiavskaia, Diana; Makiya, Michelle; Neverov, Alexander; Rezapkin, Gennady; Sebrell, Andrew; Purcell, Robert

2011-01-01

Six poliovirus-neutralizing Fabs were recovered from a combinatorial Fab phage display library constructed from bone marrow-derived lymphocytes of immunized chimpanzees. The chimeric chimpanzee-human full-length IgGs (hereinafter called monoclonal antibodies [MAbs]) were generated by combining a chimpanzee IgG light chain and a variable domain of heavy chain with a human constant Fc region. The six MAbs neutralized vaccine strains and virulent strains of poliovirus. Five MAbs were serotype specific, while one MAb cross-neutralized serotypes 1 and 2. Epitope mapping performed by selecting and sequencing antibody-resistant viral variants indicated that the cross-neutralizing MAb bound between antigenic sites 1 and 2, thereby covering the canyon region containing the receptor-binding site. Another serotype 1-specific MAb recognized a region located between antigenic sites 2 and 3 that included parts of capsid proteins VP1 and VP3. Both serotype 2-specific antibodies recognized antigenic site 1. No escape mutants to serotype 3-specific MAbs could be generated. The administration of a serotype 1-specific MAb to transgenic mice susceptible to poliovirus at a dose of 5 μg/mouse completely protected them from paralysis after challenge with a lethal dose of wild-type poliovirus. Moreover, MAb injection 6 or 12 h after virus infection provided significant protection. The MAbs described here could be tested in clinical trials to determine whether they might be useful for treatment of immunocompromised chronic virus excretors and for emergency protection of contacts of a paralytic poliomyelitis case. PMID:21345966

Recombinant PrPSc shares structural features with brain-derived PrPSc: Insights from limited proteolysis.

PubMed

Sevillano, Alejandro M; Fernández-Borges, Natalia; Younas, Neelam; Wang, Fei; R Elezgarai, Saioa; Bravo, Susana; Vázquez-Fernández, Ester; Rosa, Isaac; Eraña, Hasier; Gil, David; Veiga, Sonia; Vidal, Enric; Erickson-Beltran, Melissa L; Guitián, Esteban; Silva, Christopher J; Nonno, Romolo; Ma, Jiyan; Castilla, Joaquín; R Requena, Jesús

2018-01-01

Very solid evidence suggests that the core of full length PrPSc is a 4-rung β-solenoid, and that individual PrPSc subunits stack to form amyloid fibers. We recently used limited proteolysis to map the β-strands and connecting loops that make up the PrPSc solenoid. Using high resolution SDS-PAGE followed by epitope analysis, and mass spectrometry, we identified positions ~116/118, 133-134, 141, 152-153, 162, 169 and 179 (murine numbering) as Proteinase K (PK) cleavage sites in PrPSc. Such sites likely define loops and/or borders of β-strands, helping us to predict the threading of the β-solenoid. We have now extended this approach to recombinant PrPSc (recPrPSc). The term recPrPSc refers to bona fide recombinant prions prepared by PMCA, exhibiting infectivity with attack rates of ~100%. Limited proteolysis of mouse and bank vole recPrPSc species yielded N-terminally truncated PK-resistant fragments similar to those seen in brain-derived PrPSc, albeit with varying relative yields. Along with these fragments, doubly N- and C-terminally truncated fragments, in particular ~89/97-152, were detected in some recPrPSc preparations; similar fragments are characteristic of atypical strains of brain-derived PrPSc. Our results suggest a shared architecture of recPrPSc and brain PrPSc prions. The observed differences, in particular the distinct yields of specific PK-resistant fragments, are likely due to differences in threading which result in the specific biochemical characteristics of recPrPSc. Furthermore, recombinant PrPSc offers exciting opportunities for structural studies unachievable with brain-derived PrPSc.

EPIC-Simulated and MODIS-Derived Leaf Area Index (LAI) ...

EPA Pesticide Factsheets

Leaf Area Index (LAI) is an important parameter in assessing vegetation structure for characterizing forest canopies over large areas at broad spatial scales using satellite remote sensing data. However, satellite-derived LAI products can be limited by obstructed atmospheric conditions yielding sub-optimal values, or complete non-returns. The United States Environmental Protection Agency’s Exposure Methods and Measurements and Computational Exposure Divisions are investigating the viability of supplemental modelled LAI inputs into satellite-derived data streams to support various regional and local scale air quality models for retrospective and future climate assessments. In this present study, one-year (2002) of plot level stand characteristics at four study sites located in Virginia and North Carolina are used to calibrate species-specific plant parameters in a semi-empirical biogeochemical model. The Environmental Policy Integrated Climate (EPIC) model was designed primarily for managed agricultural field crop ecosystems, but also includes managed woody species that span both xeric and mesic sites (e.g., mesquite, pine, oak, etc.). LAI was simulated using EPIC at a 4 km2 and 12 km2 grid coincident with the regional Community Multiscale Air Quality Model (CMAQ) grid. LAI comparisons were made between model-simulated and MODIS-derived LAI. Field/satellite-upscaled LAI was also compared to the corresponding MODIS LAI value. Preliminary results show field/satel

Habitat-specific foraging strategies in Australasian gannets

PubMed Central

Wells, Melanie R.; Arnould, John P. Y.

2016-01-01

ABSTRACT Knowledge of top predator foraging adaptability is imperative for predicting their biological response to environmental variability. While seabirds have developed highly specialised techniques to locate prey, little is known about intraspecific variation in foraging strategies with many studies deriving information from uniform oceanic environments. Australasian gannets (Morus serrator) typically forage in continental shelf regions on small schooling prey. The present study used GPS and video data loggers to compare habitat-specific foraging strategies at two sites of contrasting oceanographic regimes (deep water near the continental shelf edge, n=23; shallow inshore embayment, n=26), in south-eastern Australia. Individuals from the continental shelf site exhibited pelagic foraging behaviours typical of gannet species, using local enhancement to locate and feed on small schooling fish; in contrast only 50% of the individuals from the inshore site foraged offshore, displaying the typical pelagic foraging strategy. The remainder adopted a strategy of searching sand banks in shallow inshore waters in the absence of conspecifics and other predators for large, single prey items. Furthermore, of the individuals foraging inshore, 93% were male, indicating that the inshore strategy may be sex-specific. Large inter-colony differences in Australasian gannets suggest strong plasticity in foraging behaviours, essential for adapting to environmental change. PMID:27305927

A continuous map of near-surface S-wave attenuation in New Zealand

NASA Astrophysics Data System (ADS)

Van Houtte, Chris; Ktenidou, Olga-Joan; Larkin, Tam; Holden, Caroline

2018-04-01

Quantifying the near-surface attenuation of seismic waves at a given location can be important for seismic hazard analysis of high-frequency ground motion. This study calculates the site attenuation parameter, κ0, at 41 seismograph locations in New Zealand. Combined with results of a previous study, a total of 46 κ0 values are available across New Zealand. The results compare well with previous t* studies, revealing high attenuation in the volcanic arc and forearc ranges, and low attenuation in the South Island. However, for site-specific seismic hazard analyses, there is a need to calculate κ0 at locations away from a seismograph location. For these situations, it is common to infer κ0 from weak correlations with the shear wave velocity in the top 30 m, VS30, or to adopt an indicative regional value. This study attempts to improve on this practice. Geostatistical models of the station-specific κ0 data are developed, and continuous maps are derived using ordinary kriging. The obtained κ0 maps can provide a median κ0 and its uncertainty for any location in New Zealand, which may be useful for future site-specific seismic hazard analyses.

Site-Selective Regulation of Platelet-Derived Growth Factor β Receptor Tyrosine Phosphorylation by T-Cell Protein Tyrosine Phosphatase

PubMed Central

Persson, Camilla; Sävenhed, Catrine; Bourdeau, Annie; Tremblay, Michel L.; Markova, Boyka; Böhmer, Frank D.; Haj, Fawaz G.; Neel, Benjamin G.; Elson, Ari; Heldin, Carl-Henrik; Rönnstrand, Lars; Östman, Arne; Hellberg, Carina

2004-01-01

The platelet-derived growth factor (PDGF) β receptor mediates mitogenic and chemotactic signals. Like other tyrosine kinase receptors, the PDGF β receptor is negatively regulated by protein tyrosine phosphatases (PTPs). To explore whether T-cell PTP (TC-PTP) negatively regulates the PDGF β receptor, we compared PDGF β receptor tyrosine phosphorylation in wild-type and TC-PTP knockout (ko) mouse embryos. PDGF β receptors were hyperphosphorylated in TC-PTP ko embryos. Fivefold-higher ligand-induced receptor phosphorylation was observed in TC-PTP ko mouse embryo fibroblasts (MEFs) as well. Reexpression of TC-PTP partly abolished this difference. As determined with site-specific phosphotyrosine antibodies, the extent of hyperphosphorylation varied among different autophosphorylation sites. The phospholipase Cγ1 binding site Y1021, previously implicated in chemotaxis, displayed the largest increase in phosphorylation. The increase in Y1021 phosphorylation was accompanied by increased phospholipase Cγ1 activity and migratory hyperresponsiveness to PDGF. PDGF β receptor tyrosine phosphorylation in PTP-1B ko MEFs but not in PTPɛ ko MEFs was also higher than that in control cells. This increase occurred with a site distribution different from that seen after TC-PTP depletion. PDGF-induced migration was not increased in PTP-1B ko cells. In summary, our findings identify TC-PTP as a previously unrecognized negative regulator of PDGF β receptor signaling and support the general notion that PTPs display site selectivity in their action on tyrosine kinase receptors. PMID:14966296

Developing risk-based screening guidelines for dioxin management at a Melbourne sewage treatment plant.

PubMed

Gorman, J; Mival, K; Wright, J; Howell, M

2003-01-01

Dioxin is a generic term used to refer to the congeners of polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs). The principal source of dioxin production is generally thought to be from unintended by-products of waste incineration, but dioxins are also naturally formed from volcanic activity and forest fires (WHO, 1998). Estimates of dioxin emissions in Australia suggest that approximately 75% of the total PCDD and PCDF emissions derive from prescribed burning and wild bushfires. Currently, no screening guidelines for dioxins within soils are available in Australia. This paper presents the general approach and results of a human health risk-based assessment performed by URS Australia in 2001 to develop site specific reference criteria for remediation of a former sewage plant in Melbourne. Risk-based soil remediation concentrations for dioxins at the sewage treatment plant site were developed using tolerable daily intake values of 4, 2 and 1 pg/kg/day. The potentially significant exposure pathways and processes for exposure to dioxins were identified and risk-based soil concentrations derived in accordance with the general method framework presented in the National Environmental Protection Measure (Assessment of Site Contamination). The derived dioxin reference criteria were used to develop an effective risk management program focussed on those conditions that present the greatest contribution to overall risk to human health.

Analysis Techniques, Materials, and Methods for Treatment of Thermal Bridges in Building Envelopes

DTIC Science & Technology

2013-08-01

effects of the R-value for given increment of time ............................................. 89 64 Crystals on a post-conditioned Aspen Aerogel ... aerogel on specific sites compared to conventional polyurethane foam insulation. Figures 55 and 56 show two examples of preliminary parametric... Aerogel , and (4) Honeywell’s polyurethane. Table 14 lists the four tested insulation ma- terials, their experimental thermal properties (derived

The application of automatic recognition techniques in the Apollo 9 SO-65 experiment

NASA Technical Reports Server (NTRS)

Macdonald, R. B.

1970-01-01

A synoptic feature analysis is reported on Apollo 9 remote earth surface photographs that uses the methods of statistical pattern recognition to classify density points and clusterings in digital conversion of optical data. A computer derived geological map of a geological test site indicates that geological features of the range are separable, but that specific rock types are not identifiable.

Performance of Pallet Nails and Staples in 22 Southern Hardwoods

Treesearch

E.G. Stern

1976-01-01

The effectiveness of pointless, helically threaded, hardened-steel, 3 by O.120-inch, pallet nails with umbrella heads and 21/2-inch, 15-gauge, 7/16-inch crown, plastic-coated, pallet staples in 22 hardwoods grown on southern pine sites was determined on the basis of tests on 522 joints. Specific average performance values were derived from the test data for the tested...

Cell type-specific termination of transcription by transposable element sequences.

PubMed

Conley, Andrew B; Jordan, I King

2012-09-30

Transposable elements (TEs) encode sequences necessary for their own transposition, including signals required for the termination of transcription. TE sequences within the introns of human genes show an antisense orientation bias, which has been proposed to reflect selection against TE sequences in the sense orientation owing to their ability to terminate the transcription of host gene transcripts. While there is evidence in support of this model for some elements, the extent to which TE sequences actually terminate transcription of human gene across the genome remains an open question. Using high-throughput sequencing data, we have characterized over 9,000 distinct TE-derived sequences that provide transcription termination sites for 5,747 human genes across eight different cell types. Rarefaction curve analysis suggests that there may be twice as many TE-derived termination sites (TE-TTS) genome-wide among all human cell types. The local chromatin environment for these TE-TTS is similar to that seen for 3' UTR canonical TTS and distinct from the chromatin environment of other intragenic TE sequences. However, those TE-TTS located within the introns of human genes were found to be far more cell type-specific than the canonical TTS. TE-TTS were much more likely to be found in the sense orientation than other intragenic TE sequences of the same TE family and TE-TTS in the sense orientation terminate transcription more efficiently than those found in the antisense orientation. Alu sequences were found to provide a large number of relatively weak TTS, whereas LTR elements provided a smaller number of much stronger TTS. TE sequences provide numerous termination sites to human genes, and TE-derived TTS are particularly cell type-specific. Thus, TE sequences provide a powerful mechanism for the diversification of transcriptional profiles between cell types and among evolutionary lineages, since most TE-TTS are evolutionarily young. The extent of transcription termination by TEs seen here, along with the preference for sense-oriented TE insertions to provide TTS, is consistent with the observed antisense orientation bias of human TEs.

The Effects of Urbanization and Other Environmental Gradients on Algal Assemblages in Nine Metropolitan Areas across the United States

USGS Publications Warehouse

Coles, James F.; Bell, Amanda H.; Scudder, Barbara C.; Carpenter, Kurt D.

2009-01-01

The U.S. Geological Survey conducted studies from 2000 to 2004 to determine the effects of urbanization on stream ecosystems in nine major metropolitan study areas across the United States. Biological, chemical, and physical components of streams were assessed at 28 to 30 sites in each study area. Benthic algae were sampled to compare the degree to which algal assemblages correlated to urbanization, as characterized by an urban intensity index (UII), relative to other environmental gradients that function at either the watershed or reach scales. Ordination site scores were derived from principal components analyses of the environmental data to define environmental gradients at two spatial scales: (1) watershed-scale gradients that summarized (a) landscape modifications and (b) socioeconomic factors, and (2) reach-scale gradients that characterized (a) physical habitat and (b) water chemistry. Algal response was initially quantified by site scores derived from nonmetric multi-dimensional scaling ordinations of the algal assemblage data. The site scores were then correlated with a set of algal metrics of structure and function to help select specific indicators that would best represent changes in the algal assemblages and would infer ecological condition. The selected metrics were correlated to the UII and other environmental gradients. The results indicated that diatom-taxa in the assemblages were distinctly different across the nine study areas, likely due to physiographic differences across the country, but nevertheless, some algal metrics were applicable to all areas. Overall, the study results indicated that although the UII represented various landscape changes associated with urbanization across the country, the algal response was more strongly related to more specific factors generally associated with water quality measured within the stream reach.

Precipitation Depth-Duration-Frequency Analysis for the Nevada National Security Site and Surrounding Areas

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Li; Miller, Julianne J.

Accurate precipitation frequency data are important for Environmental Management Soils Activities on the Nevada National Security Site (NNSS). These data are important for environmental assessments performed for regulatory closure of Soils Corrective Action Unit (CAU) Sites, as well as engineering mitigation designs and post-closure monitoring strategies to assess and minimize potential contaminant migration from Soils CAU Sites. Although the National Oceanic and Atmospheric Administration (NOAA) Atlas 14 (Bonnin et al., 2011) provides precipitation frequency data for the NNSS area, the NNSS-specific observed precipitation data were not consistent with the NOAA Atlas 14 predicted data. This is primarily due to themore » NOAA Atlas 14 products being produced from analyses without including the approximately 30 NNSS precipitation gage records, several of which approach or exceed 50 year of record. Therefore, a study of precipitation frequency that incorporated the NNSS precipitation gage records into the NOAA Atlas 14 dataset, was performed specifically for the NNSS to derive more accurate site-specific precipitation data products. Precipitation frequency information, such as the depth-duration-frequency (DDF) relationships, are required to generate synthetic standard design storm hydrographs and assess actual precipitation events. In this study, the actual long-term NNSS precipitation gage records, some of which are the longest gage records in southern and central Nevada, were analyzed to allow for more accurate precipitation DDF estimates to be developed for the NNSS. Gridded maps of precipitation frequency for the NNSS and surrounding areas were then produced.« less

Does Osmotic Stress Affect Natural Product Expression in Fungi?

PubMed Central

Overy, David; Chi, Wei-Chiung; Pang, Ka-Lai; Rateb, Mostafa; Shang, Zhuo; Capon, Rob; Bills, Gerald; Kerr, Russell

2017-01-01

The discovery of new natural products from fungi isolated from the marine environment has increased dramatically over the last few decades, leading to the identification of over 1000 new metabolites. However, most of the reported marine-derived species appear to be terrestrial in origin yet at the same time, facultatively halo- or osmotolerant. An unanswered question regarding the apparent chemical productivity of marine-derived fungi is whether the common practice of fermenting strains in seawater contributes to enhanced secondary metabolism? To answer this question, a terrestrial isolate of Aspergillus aculeatus was fermented in osmotic and saline stress conditions in parallel across multiple sites. The ex-type strain of A. aculeatus was obtained from three different culture collections. Site-to-site variations in metabolite expression were observed, suggesting that subculturing of the same strain and subtle variations in experimental protocols can have pronounced effects upon metabolite expression. Replicated experiments at individual sites indicated that secondary metabolite production was divergent between osmotic and saline treatments. Titers of some metabolites increased or decreased in response to increasing osmolite (salt or glycerol) concentrations. Furthermore, in some cases, the expression of some secondary metabolites in relation to osmotic and saline stress was attributed to specific sources of the ex-type strains. PMID:28805714

Regionally Adaptable Ground Motion Prediction Equation (GMPE) from Empirical Models of Fourier and Duration of Ground Motion

NASA Astrophysics Data System (ADS)

Bora, Sanjay; Scherbaum, Frank; Kuehn, Nicolas; Stafford, Peter; Edwards, Benjamin

2016-04-01

The current practice of deriving empirical ground motion prediction equations (GMPEs) involves using ground motions recorded at multiple sites. However, in applications like site-specific (e.g., critical facility) hazard ground motions obtained from the GMPEs are need to be adjusted/corrected to a particular site/site-condition under investigation. This study presents a complete framework for developing a response spectral GMPE, within which the issue of adjustment of ground motions is addressed in a manner consistent with the linear system framework. The present approach is a two-step process in which the first step consists of deriving two separate empirical models, one for Fourier amplitude spectra (FAS) and the other for a random vibration theory (RVT) optimized duration (Drvto) of ground motion. In the second step the two models are combined within the RVT framework to obtain full response spectral amplitudes. Additionally, the framework also involves a stochastic model based extrapolation of individual Fourier spectra to extend the useable frequency limit of the empirically derived FAS model. The stochastic model parameters were determined by inverting the Fourier spectral data using an approach similar to the one as described in Edwards and Faeh (2013). Comparison of median predicted response spectra from present approach with those from other regional GMPEs indicates that the present approach can also be used as a stand-alone model. The dataset used for the presented analysis is a subset of the recently compiled database RESORCE-2012 across Europe, the Middle East and the Mediterranean region.

Soil redistribution model for undisturbed and cultivated sites based on Chernobyl-derived cesium-137 fallout.

PubMed

Hrachowitz, Markus; Maringer, Franz-Josef; Steineder, Christian; Gerzabek, Martin H

2005-01-01

Measurements of 137Cs fallout have been used in combination with a range of conversion models for the investigation of soil relocation mechanisms and sediment budgets in many countries for more than 20 yr. The objective of this paper is to develop a conversion model for quantifying soil redistribution, based on Chernobyl-derived 137Cs. The model is applicable on uncultivated as well as on cultivated sites, taking into account temporal changes in the 137Cs depth distribution pattern as well as tillage-induced 137Cs dilution effects. The main idea of the new model is the combination of a modified exponential model describing uncultivated soil with a Chapman distribution based model describing cultivated soil. The compound model subsequently allows a dynamic description of the Chernobyl derived 137Cs situation in the soil and its change, specifically migration and soil transport processes over the course of time. Using the suggested model at the sampling site in Pettenbach, in the Austrian province of Oberösterreich 137Cs depth distributions were simulated with a correlation coefficient of 0.97 compared with the measured 137Cs depth profile. The simulated rates of soil distribution at different positions at the sampling site were found to be between 27 and 60 Mg ha(-1) yr(-1). It was shown that the model can be used to describe the temporal changes of 137Cs depth distributions in cultivated as well as uncultivated soils. Additionally, the model allows to quantify soil redistribution in good correspondence with already existing models.

Method for Implementing Subsurface Solid Derived Concentration Guideline Levels (DCGL) - 12331

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lively, J.W.

2012-07-01

The U.S. Nuclear Regulatory Commission (NRC) and other federal agencies currently approve the Multi-Agency Radiation Site Survey and Investigation Manual (MARSSIM) as guidance for licensees who are conducting final radiological status surveys in support of decommissioning. MARSSIM provides a method to demonstrate compliance with the applicable regulation by comparing residual radioactivity in surface soils with derived concentration guideline levels (DCGLs), but specifically discounts its applicability to subsurface soils. Many sites and facilities undergoing decommissioning contain subsurface soils that are potentially impacted by radiological constituents. In the absence of specific guidance designed to address the derivation of subsurface soil DCGLs andmore » compliance demonstration, decommissioning facilities have attempted to apply DCGLs and final status survey techniques designed specifically for surface soils to subsurface soils. The decision to apply surface soil limits and surface soil compliance metrics to subsurface soils typically results in significant over-excavation with associated cost escalation. MACTEC, Inc. has developed the overarching concepts and principles found in recent NRC decommissioning guidance in NUREG 1757 to establish a functional method to derive dose-based subsurface soil DCGLs. The subsurface soil method developed by MACTEC also establishes a rigorous set of criterion-based data evaluation metrics (with analogs to the MARSSIM methodology) that can be used to demonstrate compliance with the developed subsurface soil DCGLs. The method establishes a continuum of volume factors that relate the size and depth of a volume of subsurface soil having elevated concentrations of residual radioactivity with its ability to produce dose. The method integrates the subsurface soil sampling regime with the derivation of the subsurface soil DCGL such that a self-regulating optimization is naturally sought by both the responsible party and regulator. This paper describes the concepts and basis used by MACTEC to develop the dose-based subsurface soil DCGL method. The paper will show how MACTEC's method can be used to demonstrate that higher concentrations of residual radioactivity in subsurface soils (as compared with surface soils) can meet the NRC's dose-based regulations. MACTEC's method has been used successfully to obtain the NRC's radiological release at a site with known radiological impacts to subsurface soils exceeding the surface soil DCGL, saving both time and cost. Having considered the current NRC guidance for consideration of residual radioactivity in subsurface soils during decommissioning, MACTEC has developed a technically based approach to the derivation of and demonstration of compliance with subsurface soil DCGLs for radionuclides. In fact, the process uses the already accepted concepts and metrics approved for surface soils as the foundation for deriving scaling factors used to calculate subsurface soil DCGLs that are at least equally protective of the decommissioning annual dose standard. Each of the elements identified for consideration in the current NRC guidance is addressed in this proposed method. Additionally, there is considerable conservatism built into the assumptions and techniques used to arrive at subsurface soil scaling factors and DCGLs. The degree of conservatism embodied in the approach used is such that risk managers and decision makers approving and using subsurface soil DCGLs derived in accordance with this method can be confident that the future exposures will be well below permissible and safe levels. The technical basis for the method can be applied to a broad variety of sites with residual radioactivity in subsurface soils. Given the costly nature of soil surveys, excavation, and disposal of soils as low-level radioactive waste, MACTEC's method for deriving and demonstrating compliance with subsurface soil DCGLs offers the possibility of significant cost savings over the traditional approach of applying surface soil DCGLs to subsurface soils. Furthermore, while yet untested, MACTEC believes that the concepts and methods embodied in this approach could readily be applied to other types of contamination found in subsurface soils. (author)« less

Eucalyptus Energy Farm: feasibility study and demonstration. Phase 1: site and species selection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mariani, E.O.; Wood, W.A.; Kouchoukos, P.C.

1978-06-01

The results of investigations to determine the feasibility for establishing ''Eucalyptus Energy Farms'' as a potential source of biomass derived energy are presented. Included in this report are: the results of an extensive literature search and numerous field interviews (both domestically and internationally) to identify those Eucalyptus species most suitable for use within the United States; a description of the site selection criteria, site selection process, and identification of potential sites for use in testing the Eucalyptus Energy Farm concept; a description of the Eucalyptus species selection criteria, species selection process, and an identification of the specific species recommended formore » testing at specific sites; preliminary engineering designs for each of the proposed plantation sites; and environmental impact considerations related to Eucalyptus growing. It is concluded that although the Eucalyptus are extremely temperature sensitive and the geographic areas which appear suitable for their planting (within the United States) are principally confined to western and central California and the warmest regions of the southeastern United States, each of the areas contain large amounts of marginal land which could be converted into Eucalyptus based biomass plantations. Suitable sites, which are representative of the larger marginal land areas, are immediately available for use in establishing Eucalyptus screening test sites. Furthermore, there are not significant environmental conditions which would preclude the establishment of these initial test sites. It is therefore recommended that efforts be initiated to obtain selected provenances of Eucalyptus seeds (of the designated species) for further field testing and data collection within the United States, and that actual site selection be initiated.« less

Regression modeling of gas-particle partitioning of atmospheric oxidized mercury from temperature data

NASA Astrophysics Data System (ADS)

Cheng, Irene; Zhang, Leiming; Blanchard, Pierrette

2014-10-01

Models describing the partitioning of atmospheric oxidized mercury (Hg(II)) between the gas and fine particulate phases were developed as a function of temperature. The models were derived from regression analysis of the gas-particle partitioning parameters, defined by a partition coefficient (Kp) and Hg(II) fraction in fine particles (fPBM) and temperature data from 10 North American sites. The generalized model, log(1/Kp) = 12.69-3485.30(1/T) (R2 = 0.55; root-mean-square error (RMSE) of 1.06 m3/µg for Kp), predicted the observed average Kp at 7 of the 10 sites. Discrepancies between the predicted and observed average Kp were found at the sites impacted by large Hg sources because the model had not accounted for the different mercury speciation profile and aerosol compositions of different sources. Site-specific equations were also generated from average Kp and fPBM corresponding to temperature interval data. The site-specific models were more accurate than the generalized Kp model at predicting the observations at 9 of the 10 sites as indicated by RMSE of 0.22-0.5 m3/µg for Kp and 0.03-0.08 for fPBM. Both models reproduced the observed monthly average values, except for a peak in Hg(II) partitioning observed during summer at two locations. Weak correlations between the site-specific model Kp or fPBM and observations suggest the role of aerosol composition, aerosol water content, and relative humidity factors on Hg(II) partitioning. The use of local temperature data to parameterize Hg(II) partitioning in the proposed models potentially improves the estimation of mercury cycling in chemical transport models and elsewhere.

Small Molecule Interactome Mapping by Photoaffinity Labeling Reveals Binding Site Hotspots for the NSAIDs.

PubMed

Gao, Jinxu; Mfuh, Adelphe; Amako, Yuka; Woo, Christina M

2018-03-28

Many therapeutics elicit cell-type specific polypharmacology that is executed by a network of molecular recognition events between a small molecule and the whole proteome. However, measurement of the structures that underpin the molecular associations between the proteome and even common therapeutics, such as the nonsteroidal anti-inflammatory drugs (NSAIDs), is limited by the inability to map the small molecule interactome. To address this gap, we developed a platform termed small molecule interactome mapping by photoaffinity labeling (SIM-PAL) and applied it to the in cellulo direct characterization of specific NSAID binding sites. SIM-PAL uses (1) photochemical conjugation of NSAID derivatives in the whole proteome and (2) enrichment and isotope-recoding of the conjugated peptides for (3) targeted mass spectrometry-based assignment. Using SIM-PAL, we identified the NSAID interactome consisting of over 1000 significantly enriched proteins and directly characterized nearly 200 conjugated peptides representing direct binding sites of the photo-NSAIDs with proteins from Jurkat and K562 cells. The enriched proteins were often identified as parts of complexes, including known targets of NSAID activity (e.g., NF-κB) and novel interactions (e.g., AP-2, proteasome). The conjugated peptides revealed direct NSAID binding sites from the cell surface to the nucleus and a specific binding site hotspot for the three photo-NSAIDs on histones H2A and H2B. NSAID binding stabilized COX-2 and histone H2A by cellular thermal shift assay. Since small molecule stabilization of protein complexes is a gain of function regulatory mechanism, it is conceivable that NSAIDs affect biological processes through these broader proteomic interactions. SIM-PAL enabled characterization of NSAID binding site hotspots and is amenable to map global binding sites for virtually any molecule of interest.

An Overview on Measurement-While-Drilling Technique and its Scope in Excavation Industry

NASA Astrophysics Data System (ADS)

Rai, P.; Schunesson, H.; Lindqvist, P.-A.; Kumar, U.

2015-04-01

Measurement-while-drilling (MWD) aims at collecting accurate, speedy and high resolution information from the production blast hole drills with a target of characterization of highly variable rock masses encountered in sub-surface excavations. The essence of the technique rests on combining the physical drill variables in a manner to yield a fairly accurate description of the sub-surface rock mass much ahead of following downstream operations. In this light, the current paper presents an overview of the MWD by explaining the technique and its set-up, the existing drill-rock mass relationships and numerous on-going researches highlighting the real-time applications. Although the paper acknowledges the importance of concepts of specific energy, rock quality index and a couple of other indices and techniques for rock mass characterization, it must be distinctly borne in mind that the technique of MWD is highly site-specific, which entails derivation of site-specific calibration with utmost care.

Prediction of active sites of enzymes by maximum relevance minimum redundancy (mRMR) feature selection.

PubMed

Gao, Yu-Fei; Li, Bi-Qing; Cai, Yu-Dong; Feng, Kai-Yan; Li, Zhan-Dong; Jiang, Yang

2013-01-27

Identification of catalytic residues plays a key role in understanding how enzymes work. Although numerous computational methods have been developed to predict catalytic residues and active sites, the prediction accuracy remains relatively low with high false positives. In this work, we developed a novel predictor based on the Random Forest algorithm (RF) aided by the maximum relevance minimum redundancy (mRMR) method and incremental feature selection (IFS). We incorporated features of physicochemical/biochemical properties, sequence conservation, residual disorder, secondary structure and solvent accessibility to predict active sites of enzymes and achieved an overall accuracy of 0.885687 and MCC of 0.689226 on an independent test dataset. Feature analysis showed that every category of the features except disorder contributed to the identification of active sites. It was also shown via the site-specific feature analysis that the features derived from the active site itself contributed most to the active site determination. Our prediction method may become a useful tool for identifying the active sites and the key features identified by the paper may provide valuable insights into the mechanism of catalysis.

Urban Climate Station Site Selection Through Combined Digital Surface Model and Sun Angle Calculations

NASA Technical Reports Server (NTRS)

Kidd, Chris; Chapman, Lee

2012-01-01

Meteorological measurements within urban areas are becoming increasingly important due to the accentuating effects of climate change upon the Urban Heat Island (UHI). However, ensuring that such measurements are representative of the local area is often difficult due to the diversity of the urban environment. The evaluation of sites is important for both new sites and for the relocation of established sites to ensure that long term changes in the meteorological and climatological conditions continue to be faithfully recorded. Site selection is traditionally carried out in the field using both local knowledge and visual inspection. This paper exploits and assesses the use of lidar-derived digital surface models (DSMs) to quantitatively aid the site selection process. This is acheived by combining the DSM with a solar model, first to generate spatial maps of sky view factors and sun-hour potential and second, to generate site-specific views of the horizon. The results show that such a technique is a useful first-step approach to identify key sites that may be further evaluated for the location of meteorological stations within urban areas.

Deriving site-specific soil clean-up values for metals and metalloids: Rationale for including protection of soil microbial processes

PubMed Central

Kuperman, Roman G; Siciliano, Steven D; Römbke, Jörg; Oorts, Koen

2014-01-01

Although it is widely recognized that microorganisms are essential for sustaining soil fertility, structure, nutrient cycling, groundwater purification, and other soil functions, soil microbial toxicity data were excluded from the derivation of Ecological Soil Screening Levels (Eco-SSL) in the United States. Among the reasons for such exclusion were claims that microbial toxicity tests were too difficult to interpret because of the high variability of microbial responses, uncertainty regarding the relevance of the various endpoints, and functional redundancy. Since the release of the first draft of the Eco-SSL Guidance document by the US Environmental Protection Agency in 2003, soil microbial toxicity testing and its use in ecological risk assessments have substantially improved. A wide range of standardized and nonstandardized methods became available for testing chemical toxicity to microbial functions in soil. Regulatory frameworks in the European Union and Australia have successfully incorporated microbial toxicity data into the derivation of soil threshold concentrations for ecological risk assessments. This article provides the 3-part rationale for including soil microbial processes in the development of soil clean-up values (SCVs): 1) presenting a brief overview of relevant test methods for assessing microbial functions in soil, 2) examining data sets for Cu, Ni, Zn, and Mo that incorporated soil microbial toxicity data into regulatory frameworks, and 3) offering recommendations on how to integrate the best available science into the method development for deriving site-specific SCVs that account for bioavailability of metals and metalloids in soil. Although the primary focus of this article is on the development of the approach for deriving SCVs for metals and metalloids in the United States, the recommendations provided in this article may also be applicable in other jurisdictions that aim at developing ecological soil threshold values for protection of microbial processes in contaminated soils. PMID:24376192

Treatment of hypoxic-ischemic encephalopathy in mouse by transplantation of embryonic stem cell-derived cells.

PubMed

Ma, Jie; Wang, Yu; Yang, Jianhua; Yang, Min; Chang, Keun-A; Zhang, Linhua; Jiang, Feng; Li, Yi; Zhang, Zhonggong; Heo, Chaejeong; Suh, Yoo-Hun

2007-07-01

A 7-day-old hypoxic-ischemic encephalopathy (HIE) mouse model was used to study the effect of transplantation of embryonic stem (ES) cell-derived cells on the HIE. After the inducement in vitro, the ES cell-derived cells expressed Nestin and MAP-2, rather than GFAP mRNA. After transplantation, ES cell-derived cells can survive, migrate into the injury site, and specifically differentiate into neurons, showing improvement of the learning ability and memory of the HIE mouse at 8 months post-transplantation. The non-grafted HIE mouse brain showed typical pathological changes in the hippocampus and cerebral cortex, where the number of neurons was reduced, while in the cell graft group, number of the neurons increased in the same regions. Although further study is necessary to elucidate the precise mechanisms responsible for this functional recovery, we believe that ES cells have advantages for use as a donor source in HIE.

Evaluating multicenter DTI data in Huntington's disease on site specific effects: An ex post facto approach.

PubMed

Müller, Hans-Peter; Grön, Georg; Sprengelmeyer, Reiner; Kassubek, Jan; Ludolph, Albert C; Hobbs, Nicola; Cole, James; Roos, Raymund A C; Duerr, Alexandra; Tabrizi, Sarah J; Landwehrmeyer, G Bernhard; Süssmuth, Sigurd D

2013-01-01

Assessment of the feasibility to average diffusion tensor imaging (DTI) metrics of MRI data acquired in the course of a multicenter study. Sixty-one early stage Huntington's disease patients and forty healthy controls were studied using four different MR scanners at four European sites with acquisition protocols as close as possible to a given standard protocol. The potential and feasibility of averaging data acquired at different sites was evaluated quantitatively by region-of-interest (ROI) based statistical comparisons of coefficients of variation (CV) across centers, as well as by testing for significant group-by-center differences on averaged fractional anisotropy (FA) values between patients and controls. In addition, a whole-brain based statistical between-group comparison was performed using FA maps. The ex post facto statistical evaluation of CV and FA-values in a priori defined ROIs showed no differences between sites above chance indicating that data were not systematically biased by center specific factors. Averaging FA-maps from DTI data acquired at different study sites and different MR scanner types does not appear to be systematically biased. A suitable recipe for testing on the possibility to pool multicenter DTI data is provided to permit averaging of DTI-derived metrics to differentiate patients from healthy controls at a larger scale.

The Reduction of Risk Perception: Consensus-Making versus Truth-Seeking

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lawless, W.F.; Whitton, J.

2006-07-01

We concluded last year that the U.S. Department of Energy's (DOE's) policy of consensus-seeking (CR) for its Citizen Advisory Boards (CAB's or Boards) promoted risk mis-perception, a lack of education, and an anti-science bias about DOE's mission to cleanup its sites. Our conclusions countered an earlier study of the CAB's funded by DOE; using only subjective data, it had concluded that consensus-seeking was an improvement in American democracy. However, our conclusion was reached by comparing decision-making at the CAB's with results in the field at the DOE sites associated with the Boards. To extend our earlier findings, we looked atmore » recent meetings of the Board Chairs and preliminary results from the laboratory. We hypothesize that CR and the truth-seeking from majority rules (MR) reflect a tradeoff between a single world view derived from risk perceptions versus specific guidance from risk determinations. Based on both the field evidence and preliminary data from the experiment, we find that this tradeoff impacts site operations. At DOE's Hanford site, the risk perceptions of its Advisory Board (HAB) have contributed to 'gridlock'; at DOE's Savannah River Site (SRS), the specific recommendations by its Board (SAB) have contributed to accelerating cleanup. (authors)« less

Surface complexation modeling of Cu(II) adsorption on mixtures of hydrous ferric oxide and kaolinite

PubMed Central

Lund, Tracy J; Koretsky, Carla M; Landry, Christopher J; Schaller, Melinda S; Das, Soumya

2008-01-01

Background The application of surface complexation models (SCMs) to natural sediments and soils is hindered by a lack of consistent models and data for large suites of metals and minerals of interest. Furthermore, the surface complexation approach has mostly been developed and tested for single solid systems. Few studies have extended the SCM approach to systems containing multiple solids. Results Cu adsorption was measured on pure hydrous ferric oxide (HFO), pure kaolinite (from two sources) and in systems containing mixtures of HFO and kaolinite over a wide range of pH, ionic strength, sorbate/sorbent ratios and, for the mixed solid systems, using a range of kaolinite/HFO ratios. Cu adsorption data measured for the HFO and kaolinite systems was used to derive diffuse layer surface complexation models (DLMs) describing Cu adsorption. Cu adsorption on HFO is reasonably well described using a 1-site or 2-site DLM. Adsorption of Cu on kaolinite could be described using a simple 1-site DLM with formation of a monodentate Cu complex on a variable charge surface site. However, for consistency with models derived for weaker sorbing cations, a 2-site DLM with a variable charge and a permanent charge site was also developed. Conclusion Component additivity predictions of speciation in mixed mineral systems based on DLM parameters derived for the pure mineral systems were in good agreement with measured data. Discrepancies between the model predictions and measured data were similar to those observed for the calibrated pure mineral systems. The results suggest that quantifying specific interactions between HFO and kaolinite in speciation models may not be necessary. However, before the component additivity approach can be applied to natural sediments and soils, the effects of aging must be further studied and methods must be developed to estimate reactive surface areas of solid constituents in natural samples. PMID:18783619

Tumor infiltrating lymphocyte therapy for metastatic melanoma: analysis of tumors resected for TIL.

PubMed

Goff, Stephanie L; Smith, Franz O; Klapper, Jacob A; Sherry, Richard; Wunderlich, John R; Steinberg, Seth M; White, Donald; Rosenberg, Steven A; Dudley, Mark E; Yang, James C

2010-10-01

Adoptive cell transfer of tumor-infiltrating lymphocytes (TILs) can mediate objective tumor regression in 49% to 72% of patients with many long-term durable responses. To undergo treatment a patient must have (1) a resectable tumor from which (2) TIL can be generated that (3) exhibit tumor-specific reactivity. From July 2002 to July 2007, 787 tumors from 402 patients were processed for possible use in the generation of TIL, leading to the eventual treatment of 107 patients (27%). Viable TILs were generated in 376 patients (94%), and active, specific TILs were identified in 269 patients (67%). Patient demographics and tumor characteristics were analyzed for possible prognostic factors for growth and activity. Gastrointestinal-derived TIL grew less frequently, whereas lymph node and lung-derived TIL exhibited specific activity more often. TIL that grew and exhibited specific reactivity were from tumors that were larger in diameter and digests that had a higher percentage of lymphocytes. Despite these considerations, active, specific TIL could be generated from almost any site of metastasis. As more centers begin exploring the use of adoptive transfer with TIL, this compendium may provide a framework for therapeutic decision making and future investigation.

An alternative approach to probabilistic seismic hazard analysis in the Aegean region using Monte Carlo simulation

NASA Astrophysics Data System (ADS)

Weatherill, Graeme; Burton, Paul W.

2010-09-01

The Aegean is the most seismically active and tectonically complex region in Europe. Damaging earthquakes have occurred here throughout recorded history, often resulting in considerable loss of life. The Monte Carlo method of probabilistic seismic hazard analysis (PSHA) is used to determine the level of ground motion likely to be exceeded in a given time period. Multiple random simulations of seismicity are generated to calculate, directly, the ground motion for a given site. Within the seismic hazard analysis we explore the impact of different seismic source models, incorporating both uniform zones and distributed seismicity. A new, simplified, seismic source model, derived from seismotectonic interpretation, is presented for the Aegean region. This is combined into the epistemic uncertainty analysis alongside existing source models for the region, and models derived by a K-means cluster analysis approach. Seismic source models derived using the K-means approach offer a degree of objectivity and reproducibility into the otherwise subjective approach of delineating seismic sources using expert judgment. Similar review and analysis is undertaken for the selection of peak ground acceleration (PGA) attenuation models, incorporating into the epistemic analysis Greek-specific models, European models and a Next Generation Attenuation model. Hazard maps for PGA on a "rock" site with a 10% probability of being exceeded in 50 years are produced and different source and attenuation models are compared. These indicate that Greek-specific attenuation models, with their smaller aleatory variability terms, produce lower PGA hazard, whilst recent European models and Next Generation Attenuation (NGA) model produce similar results. The Monte Carlo method is extended further to assimilate epistemic uncertainty into the hazard calculation, thus integrating across several appropriate source and PGA attenuation models. Site condition and fault-type are also integrated into the hazard mapping calculations. These hazard maps are in general agreement with previous maps for the Aegean, recognising the highest hazard in the Ionian Islands, Gulf of Corinth and Hellenic Arc. Peak Ground Accelerations for some sites in these regions reach as high as 500-600 cm s -2 using European/NGA attenuation models, and 400-500 cm s -2 using Greek attenuation models.

Site specificity of adrenalectomy-induced brain growth.

PubMed

Thomas, T L; Devenport, L D

1988-12-01

Infant, juvenile, and adult brain growth is modulated by corticosterone. This study was designed to determine whether such modulation is confined to certain specific brain areas, and if the pattern of growth revealed is consistent across strains of rats. Young female Sprague-Dawley-derived rats were either adrenalectomized (ADX) or sham-operated (Sham) and allowed to mature 45 days before they were sacrificed for histological analysis. Fore brain sections were taken at several planes for display by projection microscope. Of the 21 sites examined, ADX exerted its greatest effect upon neocortical tissue and myelinated fiber tracts. The only other brain region affected was thalamus, which exhibited a significant widening as a result of ADX. In contrast, archicortical structures were notably unaffected by ADX. Neither the hippocampus, measured from a variety of planes, nor nuclei in the septal area were subject to increased growth by ADX. This general portrayal of ADX's site specificity held across strains of rats. However, there were local differences. Within the neopallium, the frontal region underwent the greatest thickening in one strain, while the occipital area was most strongly affected in the other. Parietal cortex was equally responsive in both strains. The pattern of sensitive vs insensitive sites bore a resemblance to the pattern of increased growth brought about by environmental enrichment as well as the fore brain distribution of Type 2 corticosterone receptors.

Evolutionary Diversifaction of Aminopeptidase N in Lepidoptera by Conserved Clade-specific Amino Acid Residues

PubMed Central

Hughes, Austin L.

2015-01-01

Members of the aminopepidase N (APN) gene family of the insect order Lepidoptera (moths and butterflies) bind the naturally insecticidal Cry toxins produced by the bacterium Bacillus thuringiensis. Phylogenetic analysis of amino acid sequences of seven lepidopteran APN classes provided strong support for the hypothesis that lepidopteran APN2 class arose by gene duplication prior to the most recent common ancestor of Lepidoptera and Diptera. The Cry toxin-binding region (BR) of lepidopteran and dipteran APNs was subject to stronger purifying selection within APN classes than was the remainder of the molecule, reflecting conservation of catalytic site and adjoining residues within the BR. Of lepidopteran APN classes, APN2, APN6, and APN8 showed the strongest evidence of functional specialization, both in expression patterns and in the occurrence of conserved derived amino acid residues. The latter three APN classes also shared a convergently evolved conserved residue close to the catalytic site. APN8 showed a particularly strong tendency towards class-specific conserved residues, including one of the catalytic site residues in the BR and ten others in close vicinity to the catalytic site residues. The occurrence of class-specific sequences along with the conservation of enzymatic function is consistent with the hypothesis that the presence of Cry toxins in the environment has been a factor shaping the evolution of this multi-gene family. PMID:24675701

Documenting the location of systematic transrectal ultrasound-guided prostate biopsies: correlation with multi-parametric MRI.

PubMed

Turkbey, Baris; Xu, Sheng; Kruecker, Jochen; Locklin, Julia; Pang, Yuxi; Shah, Vijay; Bernardo, Marcelino; Baccala, Angelo; Rastinehad, Ardeshir; Benjamin, Compton; Merino, Maria J; Wood, Bradford J; Choyke, Peter L; Pinto, Peter A

2011-03-29

During transrectal ultrasound (TRUS)-guided prostate biopsies, the actual location of the biopsy site is rarely documented. Here, we demonstrate the capability of TRUS-magnetic resonance imaging (MRI) image fusion to document the biopsy site and correlate biopsy results with multi-parametric MRI findings. Fifty consecutive patients (median age 61 years) with a median prostate-specific antigen (PSA) level of 5.8 ng/ml underwent 12-core TRUS-guided biopsy of the prostate. Pre-procedural T2-weighted magnetic resonance images were fused to TRUS. A disposable needle guide with miniature tracking sensors was attached to the TRUS probe to enable fusion with MRI. Real-time TRUS images during biopsy and the corresponding tracking information were recorded. Each biopsy site was superimposed onto the MRI. Each biopsy site was classified as positive or negative for cancer based on the results of each MRI sequence. Sensitivity, specificity, and receiver operating curve (ROC) area under the curve (AUC) values were calculated for multi-parametric MRI. Gleason scores for each multi-parametric MRI pattern were also evaluated. Six hundred and 5 systemic biopsy cores were analyzed in 50 patients, of whom 20 patients had 56 positive cores. MRI identified 34 of 56 positive cores. Overall, sensitivity, specificity, and ROC area values for multi-parametric MRI were 0.607, 0.727, 0.667, respectively. TRUS-MRI fusion after biopsy can be used to document the location of each biopsy site, which can then be correlated with MRI findings. Based on correlation with tracked biopsies, T2-weighted MRI and apparent diffusion coefficient maps derived from diffusion-weighted MRI are the most sensitive sequences, whereas the addition of delayed contrast enhancement MRI and three-dimensional magnetic resonance spectroscopy demonstrated higher specificity consistent with results obtained using radical prostatectomy specimens.

Impact of metals in surface matrices from formal and informal electronic-waste recycling around Metro Manila, the Philippines, and intra-Asian comparison.

PubMed

Fujimori, Takashi; Takigami, Hidetaka; Agusa, Tetsuro; Eguchi, Akifumi; Bekki, Kanae; Yoshida, Aya; Terazono, Atsushi; Ballesteros, Florencio C

2012-06-30

We report concentrations, enrichment factors, and hazard indicators of 11 metals (Ag, As, Cd, Co, Cu, Fe, In, Mn, Ni, Pb, and Zn) in soil and dust surface matrices from formal and informal electronic waste (e-waste) recycling sites around Metro Manila, the Philippines, referring to soil guidelines and previous data from various e-waste recycling sites in Asia. Surface dust from e-waste recycling sites had higher levels of metal contamination than surface soil. Comparison of formal and informal e-waste recycling sites (hereafter, "formal" and "informal") revealed differences in specific contaminants. Formal dust contained a mixture of serious pollutant metals (Ni, Cu, Pb, and Zn) and Cd (polluted modestly), quite high enrichment metals (Ag and In), and crust-derived metals (As, Co, Fe, and Mn). For informal soil, concentration levels of specific metals (Cd, Co, Cu, Mn, Ni, Pb, and Zn) were similar among Asian recycling sites. Formal dust had significantly higher hazardous risk than the other matrices (p<0.005), excluding informal dust (p=0.059, almost significant difference). Thus, workers exposed to formal dust should protect themselves from hazardous toxic metals (Pb and Cu). There is also a high health risk for children ingesting surface matrices from informal e-waste recycling sites. Copyright © 2012 Elsevier B.V. All rights reserved.

Discovery and validation of information theory-based transcription factor and cofactor binding site motifs.

PubMed

Lu, Ruipeng; Mucaki, Eliseos J; Rogan, Peter K

2017-03-17

Data from ChIP-seq experiments can derive the genome-wide binding specificities of transcription factors (TFs) and other regulatory proteins. We analyzed 765 ENCODE ChIP-seq peak datasets of 207 human TFs with a novel motif discovery pipeline based on recursive, thresholded entropy minimization. This approach, while obviating the need to compensate for skewed nucleotide composition, distinguishes true binding motifs from noise, quantifies the strengths of individual binding sites based on computed affinity and detects adjacent cofactor binding sites that coordinate with the targets of primary, immunoprecipitated TFs. We obtained contiguous and bipartite information theory-based position weight matrices (iPWMs) for 93 sequence-specific TFs, discovered 23 cofactor motifs for 127 TFs and revealed six high-confidence novel motifs. The reliability and accuracy of these iPWMs were determined via four independent validation methods, including the detection of experimentally proven binding sites, explanation of effects of characterized SNPs, comparison with previously published motifs and statistical analyses. We also predict previously unreported TF coregulatory interactions (e.g. TF complexes). These iPWMs constitute a powerful tool for predicting the effects of sequence variants in known binding sites, performing mutation analysis on regulatory SNPs and predicting previously unrecognized binding sites and target genes. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Dynamics Govern Specificity of a Protein-Protein Interface: Substrate Recognition by Thrombin

PubMed Central

Fuchs, Julian E.; Huber, Roland G.; Waldner, Birgit J.; Kahler, Ursula; von Grafenstein, Susanne; Kramer, Christian; Liedl, Klaus R.

2015-01-01

Biomolecular recognition is crucial in cellular signal transduction. Signaling is mediated through molecular interactions at protein-protein interfaces. Still, specificity and promiscuity of protein-protein interfaces cannot be explained using simplistic static binding models. Our study rationalizes specificity of the prototypic protein-protein interface between thrombin and its peptide substrates relying solely on binding site dynamics derived from molecular dynamics simulations. We find conformational selection and thus dynamic contributions to be a key player in biomolecular recognition. Arising entropic contributions complement chemical intuition primarily reflecting enthalpic interaction patterns. The paradigm “dynamics govern specificity” might provide direct guidance for the identification of specific anchor points in biomolecular recognition processes and structure-based drug design. PMID:26496636

Evaluating Protein Structure and Dynamics Using Co-Solvents, Photochemical Triggers, and Site-Specific Spectroscopic Probes

NASA Astrophysics Data System (ADS)

Abaskharon, Rachel M.

As ubiquitous and diverse biopolymers, proteins are dynamic molecules that are constantly engaging in inter- and intramolecular interactions responsible for their structure, fold, and function. Because of this, gaining a comprehensive understanding of the factors that control protein conformation and dynamics remains elusive as current experimental techniques often lack the ability to initiate and probe a specific interaction or conformational transition. For this reason, this thesis aims to develop methods to control and monitor protein conformations, conformational transitions, and dynamics in a site-specific manner, as well as to understand how specific and non-specific interactions affect the protein folding energy landscape. First, by using the co-solvent, trifluoroethanol (TFE), we show that the rate at which a peptide folds can be greatly impacted and thus controlled by the excluded volume effect. Secondly, we demonstrate the utility of several light-responsive molecules and reactions as methods to manipulate and investigate protein-folding processes. Using an azobenzene linker as a photo-initiator, we are able to increase the folding rate of a protein system by an order of magnitude by channeling a sub-population through a parallel, faster folding pathway. Additionally, we utilize a tryptophan-mediated electron transfer process to a nearby disulfide bond to strategically unfold a protein molecule with ultraviolet light. We also demonstrate the potential of two ruthenium polypyridyl complexes as ultrafast phototriggers of protein reactions. Finally, we develop several site-specific spectroscopic probes of protein structure and environment. Specifically, we demonstrate that a 13C-labeled aspartic acid residue constitutes a useful site-specific infrared probe for investigating salt-bridges and hydration dynamics of proteins, particularly in proteins containing several acidic amino acids. We also show that a proline-derivative, 4-oxoproline, possesses novel infrared properties that can be exploited to monitor the cis-trans isomerization process of individual proline residues in proteins.

Virtual screening and biological evaluation of piperazine derivatives as human acetylcholinesterase inhibitors.

PubMed

Varadaraju, Kavitha Raj; Kumar, Jajur Ramanna; Mallesha, Lingappa; Muruli, Archana; Mohana, Kikkeri Narasimha Shetty; Mukunda, Chethan Kumar; Sharanaiah, Umesha

2013-01-01

The piperazine derivatives have been shown to inhibit human acetylcholinesterase. Virtual screening by molecular docking of piperazine derivatives 1-(1,4-benzodioxane-2-carbonyl) piperazine (K), 4-(4-methyl)-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S1), and 4-(4-chloro)-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S3) has been shown to bind at peripheral anionic site and catalytic sites, whereas 4-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S4) and 4-(2,5-dichloro)-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S7) do not bind either to peripheral anionic site or catalytic site with hydrogen bond. All the derivatives have differed in number of H-bonds and hydrophobic interactions. The peripheral anionic site interacting molecules have proven to be potential therapeutics in inhibiting amyloid peptides aggregation in Alzheimer's disease. All the piperazine derivatives follow Lipinski's rule of five. Among all the derivatives 1-(1,4-benzodioxane-2-carbonyl) piperazine (K) was found to have the lowest TPSA value.

Virtual Screening and Biological Evaluation of Piperazine Derivatives as Human Acetylcholinesterase Inhibitors

PubMed Central

Varadaraju, Kavitha Raj; Kumar, Jajur Ramanna; Mallesha, Lingappa; Muruli, Archana; Mohana, Kikkeri Narasimha Shetty; Mukunda, Chethan Kumar; Sharanaiah, Umesha

2013-01-01

The piperazine derivatives have been shown to inhibit human acetylcholinesterase. Virtual screening by molecular docking of piperazine derivatives 1-(1,4-benzodioxane-2-carbonyl) piperazine (K), 4-(4-methyl)-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S1), and 4-(4-chloro)-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S3) has been shown to bind at peripheral anionic site and catalytic sites, whereas 4-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S4) and 4-(2,5-dichloro)-benzenesulfonyl-1-(1,4-benzodioxane-2-carbonyl) piperazine (S7) do not bind either to peripheral anionic site or catalytic site with hydrogen bond. All the derivatives have differed in number of H-bonds and hydrophobic interactions. The peripheral anionic site interacting molecules have proven to be potential therapeutics in inhibiting amyloid peptides aggregation in Alzheimer's disease. All the piperazine derivatives follow Lipinski's rule of five. Among all the derivatives 1-(1,4-benzodioxane-2-carbonyl) piperazine (K) was found to have the lowest TPSA value. PMID:24288651

Resveratrol derivatives as a pharmacological tool.

PubMed

Biasutto, Lucia; Mattarei, Andrea; Azzolini, Michele; La Spina, Martina; Sassi, Nicola; Romio, Matteo; Paradisi, Cristina; Zoratti, Mario

2017-09-01

Prodrugs of resveratrol are under development. Among the long-term goals, still largely elusive, are (1) modulating physical properties (e.g., water-soluble derivatives bearing polyethylene glycol chains), (2) changing distribution in the body (e.g., galactosyl derivatives restricted to the intestinal lumen), (3) increasing absorption from the gastrointestinal tract (e.g., derivatives imitating the natural substrates of endogenous transporters), and (4) hindering phase II metabolism (e.g., temporarily blocking the hydroxyls), all contributing to (5) increasing bioavailability. The chemical bonds that have been tested for functionalization include carboxyester, acetal, and carbamate groups. A second approach, which can be combined with the first, seeks to reinforce or modify the biochemical activities of resveratrol by concentrating the compound at specific subcellular sites. An example is provided by mitochondria-targeted derivatives. These proved to be pro-oxidant and cytotoxic in vitro, selectively killing fast-growing and tumor cells when supplied in the low micromolar range. This suggests the possibility of anticancer applications. © 2017 New York Academy of Sciences.

Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET

PubMed Central

Peinado, Héctor; Alečković, Maša; Lavotshkin, Simon; Matei, Irina; Costa-Silva, Bruno; Moreno-Bueno, Gema; Hergueta-Redondo, Marta; Williams, Caitlin; García-Santos, Guillermo; Nitadori-Hoshino, Ayuko; Hoffman, Caitlin; Badal, Karen; Garcia, Benjamin A.; Callahan, Margaret K.; Yuan, Jianda; Martins, Vilma R.; Skog, Johan; Kaplan, Rosandra N.; Brady, Mary S.; Wolchok, Jedd D.; Chapman, Paul B.; Kang, Yibin; Bromberg, Jacqueline; Lyden, David

2013-01-01

Tumor-derived exosomes are emerging mediators of tumorigenesis with tissue-specific addresses and messages. We explored the function of melanoma-derived exosomes in the formation of primary tumor and metastases in mouse and human subjects. Exosomes from highly metastatic melanoma increased the metastatic behavior of primary tumors by permanently “educating” bone marrow (BM) progenitors via the MET receptor. Melanoma-derived exosomes also induced vascular leakiness at pre-metastatic sites, and reprogrammed BM progenitors towards a c-Kit+Tie2+Met+ pro-vasculogenic phenotype. Reducing Met expression in exosomes diminished the pro-metastatic behavior of BM cells. Importantly, MET expression was elevated in circulating CD45−C-KITlow/+TIE2+ BM progenitors from metastatic melanoma subjects. RAB1a, RAB5b, RAB7, and RAB27a were highly expressed in melanoma cells and Rab27a RNA interference decreased exosome production, preventing BM education, tumor growth and metastasis. Finally, we identified an exosome-specific “melanoma signature” with prognostic and therapeutic potential, comprised of TYRP2, VLA-4, HSP70, an HSP90 isoform and the MET oncoprotein. PMID:22635005

Synovium-derived stem cells: a tissue-specific stem cell for cartilage engineering and regeneration.

PubMed

Jones, Brendan A; Pei, Ming

2012-08-01

Articular cartilage is difficult to heal once injury or disease occurs. Autologous chondrocyte transplantation is a biological treatment with good prognosis, but donor site morbidity and limited cell source are disadvantages. Currently, mesenchymal stem cells (MSCs) are a promising approach for cartilage regeneration. Despite there being various sources, the best candidate for cartilage regeneration is the one with the greatest chondrogenic potential and the least hypertrophic differentiation. These properties are able to insure that the regenerated tissue is hyaline cartilage of high quality. This review article will summarize relevant literature to justify synovium-derived stem cells (SDSCs) as a tissue-specific stem cell for chondrogenesis by comparing synovium and cartilage with respect to anatomical location and functional structure, comparing the growth characterization and chondrogenic capacity of SDSCs and MSCs, evaluating the application of SDSCs in regenerative medicine and diseases, and discussing potential future directions.

Hybrid lentivirus-phiC31-int-NLS vector allows site-specific recombination in murine and human cells but induces DNA damage.

PubMed

Grandchamp, Nicolas; Altémir, Dorothée; Philippe, Stéphanie; Ursulet, Suzanna; Pilet, Héloïse; Serre, Marie-Claude; Lenain, Aude; Serguera, Che; Mallet, Jacques; Sarkis, Chamsy

2014-01-01

Gene transfer allows transient or permanent genetic modifications of cells for experimental or therapeutic purposes. Gene delivery by HIV-derived lentiviral vector (LV) is highly effective but the risk of insertional mutagenesis is important and the random/uncontrollable integration of the DNA vector can deregulate the cell transcriptional activity. Non Integrative Lentiviral Vectors (NILVs) solve this issue in non-dividing cells, but they do not allow long term expression in dividing cells. In this context, obtaining stable expression while avoiding the problems inherent to unpredictable DNA vector integration requires the ability to control the integration site. One possibility is to use the integrase of phage phiC31 (phiC31-int) which catalyzes efficient site-specific recombination between the attP site in the phage genome and the chromosomal attB site of its Streptomyces host. Previous studies showed that phiC31-int is active in many eukaryotic cells, such as murine or human cells, and directs the integration of a DNA substrate into pseudo attP sites (pattP) which are homologous to the native attP site. In this study, we combined the efficiency of NILV for gene delivery and the specificity of phiC31-int for DNA substrate integration to engineer a hybrid tool for gene transfer with the aim of allowing long term expression in dividing and non-dividing cells preventing genotoxicity. We demonstrated the feasibility to target NILV integration in human and murine pattP sites with a dual NILV vectors system: one which delivers phiC31-int, the other which constitute the substrate containing an attB site in its DNA sequence. These promising results are however alleviated by the occurrence of significant DNA damages. Further improvements are thus required to prevent chromosomal rearrangements for a therapeutic use of the system. However, its use as a tool for experimental applications such as transgenesis is already applicable.

Teaching Geography and History through GIS: Application on Greek cultural sites

NASA Astrophysics Data System (ADS)

Skentos, Athanasios; Pavlopoulos, Kosmas; Galani, Apostolia; Theodorakopoulou, Katerina; Kritikos, Giorgos

2013-04-01

This study deals with the presentation of cultural succession in Greek space-time through a GIS application, associated with core concepts of geographic and historical education. Through the specific application students will be able to develop five distinct skills: sense of time-scale, historical and geographic comprehension, spatial analysis and interpretation, ability to perform geo-historical research, and procedure of geo-historical decision-making. The methodology is based on the calibration of a set of criteria for each cultural site that covers the topics of economy, geomorphology, society, religion, art and science. Further analysis of these data forms a geodatabase. In addition, palaeogeographic and historical maps of the cultural sites derived by the geodatabase provide information about temporal and spatial changes. As result, students will be able to develop a multidimensional and interdisciplinary approach, in order to reconstruct the evolution of the site.

Airborne remote sensors applied to engineering geology and civil works design investigations

NASA Technical Reports Server (NTRS)

Gelnett, R. H.

1975-01-01

The usefulness of various airborne remote sensing systems in the detection and identification of regional and specific geologic structural features that may affect the design and location of engineering structures on major civil works projects is evaluated. The Butler Valley Dam and Blue Lake Project in northern California was selected as a demonstration site. Findings derived from the interpretation of various kinds of imagery used are given.

Electronic structure of the organic semiconductor copper phthalocyanine: experiment and theory.

PubMed

Aristov, V Yu; Molodtsova, O V; Maslyuk, V V; Vyalikh, D V; Zhilin, V M; Ossipyan, Yu A; Bredow, T; Mertig, I; Knupfer, M

2008-01-21

The electronic structure of the organic semiconductor copper-phthalocyanine (CuPc) has been determined by a combination of conventional and resonant photoemission, near-edge x-ray absorption, as well as by the first-principles calculations. The experimentally obtained electronic valence band structure of CuPc is in very good agreement with the calculated density of states results, allowing the derivation of detailed site specific information.

Statistical adjustment of culture-independent diagnostic tests for trend analysis in the Foodborne Diseases Active Surveillance Network (FoodNet), USA.

PubMed

Gu, Weidong; Dutta, Vikrant; Patrick, Mary; Bruce, Beau B; Geissler, Aimee; Huang, Jennifer; Fitzgerald, Collette; Henao, Olga

2018-03-19

Culture-independent diagnostic tests (CIDTs) are increasingly used to diagnose Campylobacter infection in the Foodborne Diseases Active Surveillance Network (FoodNet). Because CIDTs have different performance characteristics compared with culture, which has been used historically and is still used to diagnose campylobacteriosis, adjustment of cases diagnosed by CIDT is needed to compare with culture-confirmed cases for monitoring incidence trends. We identified the necessary parameters for CIDT adjustment using culture as the gold standard, and derived formulas to calculate positive predictive values (PPVs). We conducted a literature review and meta-analysis to examine the variability in CIDT performance and Campylobacter prevalence applicable to FoodNet sites. We then developed a Monte Carlo method to estimate test-type and site-specific PPVs with their associated uncertainties. The uncertainty in our estimated PPVs was largely derived from uncertainty about the specificity of CIDTs and low prevalence of Campylobacter in tested samples. Stable CIDT-adjusted incidences of Campylobacter cases from 2012 to 2015 were observed compared with a decline in culture-confirmed incidence. We highlight the lack of data on the total numbers of tested samples as one of main limitations for CIDT adjustment. Our results demonstrate the importance of adjusting CIDTs for understanding trends in Campylobacter incidence in FoodNet.

Surface texture analysis of southern Tuli Basin sediments: Implications for Limpopo Valley geoarchaeological contexts

NASA Astrophysics Data System (ADS)

Le Baron, Joel C.; Grab, Stefan W.; Kuman, Kathleen

2011-03-01

The Hackthorne 1 site (southern Tuli Basin, South Africa) is situated on a sand-covered plateau adjacent to the Limpopo River Valley. Although the site is well known for its Stone Age archaeology, the past environmental contexts (particularly sedimentological/geomorphological) are not well known. We examine the Hackthorne sand grain surface textures, so as to provide some insight on the site specific and regional depositional history. Quartz sands at Hackthorne were collected from surface sands and from underlying weathered calcrete. SEM analysis was performed on sand grains, through which several mechanical and chemical microtextures were identified. Microtextures typical of fluvial environments were found only on grains derived from the plateau calcrete host sediment, whilst the surface sands exhibited only textures associated with aeolian environments. The results indicate that the calcrete host sediment is composed of alluvium, and that the surface sands mantling the Hackthorne Plateau are not deflated from the alluvial deposits in the Limpopo Valley, but may rather be derived from distant aeolian sources. The deposition of aeolian sands is consistent with OSL dates which place sand deposition, or remobilization, at 23 and 15 kya, periods in southern Africa associated with increased aridity.

Assessment of Provisional MODIS-derived Surfaces Related to the Global Carbon Cycle

NASA Astrophysics Data System (ADS)

Cohen, W. B.; Maiersperger, T. K.; Turner, D. P.; Gower, S. T.; Kennedy, R. E.; Running, S. W.

2002-12-01

The global carbon cycle is one of the most important foci of an emerging global biosphere monitoring system. A key component of such a system is the MODIS sensor, onboard the Terra satellite platform. Biosphere monitoring requires an integrated program of satellite observations, Earth-system models, and in situ data. Related to the carbon cycle, MODIS science teams routinely develop a variety of global surfaces such as land cover, leaf area index, and net primary production using MODIS data and functional algorithms. The quality of these surfaces must be evaluated to determine their effectiveness for global biosphere monitoring. A project called BigFoot (http://www.fsl.orst.edu/larse/bigfoot/) is an organized effort across nine biomes to assess the quality of the abovementioned surfaces: (1) Arctic tundra; (2) boreal evergreen needle-leaved forest; temperate (3) cropland, (4) grassland, (5) evergreen needle-leaved forest, and (6) deciduous broad-leaved forest; desert (7) grassland and (8) shrubland; and (9) tropical evergreen broad-leaved forest. Each biome is represented by a site that has an eddy-covariance flux tower that measures water vapor and CO2 fluxes. Flux tower footprints are relatively small-approximately 1 km2. BigFoot characterizes 25 km2 around each tower, using field data, Landsat ETM+ image data, and ecosystem process models. Our innovative field sampling design incorporates a nested spatial series to facilitate geostatistical analyses, samples the ecological variability at a site, and is logistically efficient. Field data are used both to develop site-specific algorithms for mapping/modeling the variables of interest and to characterize the errors in derived BigFoot surfaces. Direct comparisons of BigFoot- and MODIS-derived surfaces are made to help understand the sources of error in MODIS-derived surfaces and to facilitate improvements to MODIS algorithms. Results from four BigFoot sites will be presented.

Assessment of the Unintentional Reuse of Municipal Wastewater

NASA Astrophysics Data System (ADS)

Okasaki, S.; Fono, L.; Sedlak, D. L.; Dracup, J. A.

2002-12-01

Many surface waters that receive wastewater effluent also serve as source waters for drinking water treatment plants. Recent research has shown that a number of previously undiscovered wastewater-derived contaminants are present in these surface waters, including pharmaceuticals and human hormones, several of which are suspected carcinogens or endocrine disrupters and are, as of yet, unregulated through drinking water standards. This research has been designed to determine the extent of contamination of specific wastewater-derived contaminants in surface water bodies that both receive wastewater effluent and serve as a source of drinking water to a sizeable population. We are testing the hypothesis that surface water supplies during low flow are potentially of worse quality than carefully monitored reclaimed water. The first phase of our research involves: (1) the selection of sites for study; (2) a hydrologic analysis of the selected sites to determine average flow of the source water during median- and low-flow conditions; and (3) the development and testing of chemical analyses, including both conservative and reactive tracers that have been studied in microcosms and wetlands for attenuation rates. The second phase involves the development and use of the hydrologic model QUAL2E to simulate each of the selected watersheds in order to estimate potential stream water quality impairments at the drinking water intake at each site. The results of the model are verified with field sampling at designated locations at each site. We expect to identify several critical river basins where surface water at the drinking water intake contains sufficient wastewater-derived contaminants to warrant concern. If wastewater-derived contaminants are detected, we will estimate the average annual exposure of consumers of this water. We will compare these expected and actual concentrations with typical constituent concentrations found in wastewater that has undergone advanced treatment for reclamation. We may demonstrate that the surface water supplies during low flow are actually of worse quality than carefully monitored reclaimed water.

Spatio-temporal variation of urban ultrafine particle number concentrations

NASA Astrophysics Data System (ADS)

Ragettli, Martina S.; Ducret-Stich, Regina E.; Foraster, Maria; Morelli, Xavier; Aguilera, Inmaculada; Basagaña, Xavier; Corradi, Elisabetta; Ineichen, Alex; Tsai, Ming-Yi; Probst-Hensch, Nicole; Rivera, Marcela; Slama, Rémy; Künzli, Nino; Phuleria, Harish C.

2014-10-01

Methods are needed to characterize short-term exposure to ultrafine particle number concentrations (UFP) for epidemiological studies on the health effects of traffic-related UFP. Our aims were to assess season-specific spatial variation of short-term (20-min) UFP within the city of Basel, Switzerland, and to develop hybrid models for predicting short-term median and mean UFP levels on sidewalks. We collected measurements of UFP for periods of 20 min (MiniDiSC particle counter) and determined traffic volume along sidewalks at 60 locations across the city, during non-rush hours in three seasons. For each monitoring location, detailed spatial characteristics were locally recorded and potential predictor variables were derived from geographic information systems (GIS). We built multivariate regression models to predict local UFP, using concurrent UFP levels measured at a suburban background station, and combinations of meteorological, temporal, GIS and observed site characteristic variables. For a subset of sites, we assessed the relationship between UFP measured on the sidewalk and at the nearby residence (i.e., home outdoor exposure on e.g. balconies). The average median 20-min UFP levels at street and urban background sites were 14,700 ± 9100 particles cm-3 and 9900 ± 8600 particles cm-3, respectively, with the highest levels occurring in winter and the lowest in summer. The most important predictor for all models was the suburban background UFP concentration, explaining 50% and 38% of the variability of the median and mean, respectively. While the models with GIS-derived variables (R2 = 0.61) or observed site characteristics (R2 = 0.63) predicted median UFP levels equally well, mean UFP predictions using only site characteristic variables (R2 = 0.62) showed a better fit than models using only GIS variables (R2 = 0.55). The best model performance was obtained by using a combination of GIS-derived variables and locally observed site characteristics (median: R2 = 0.66; mean: R2 = 0.65). The 20-min UFP concentrations measured at the sidewalk were strongly related (R2 = 0.8) to the concurrent 20-min residential UFP levels nearby. Our results indicate that median UFP can be moderately predicted by means of a suburban background site and GIS-derived traffic and land use variables. In areas and regions where large-scale GIS data are not available, the spatial distribution of traffic-related UFP may be assessed reasonably well by collecting on-site short-term traffic and land-use data.

Quantitative Analysis of Human Pluripotency and Neural Specification by In-Depth (Phospho)Proteomic Profiling.

PubMed

Singec, Ilyas; Crain, Andrew M; Hou, Junjie; Tobe, Brian T D; Talantova, Maria; Winquist, Alicia A; Doctor, Kutbuddin S; Choy, Jennifer; Huang, Xiayu; La Monaca, Esther; Horn, David M; Wolf, Dieter A; Lipton, Stuart A; Gutierrez, Gustavo J; Brill, Laurence M; Snyder, Evan Y

2016-09-13

Controlled differentiation of human embryonic stem cells (hESCs) can be utilized for precise analysis of cell type identities during early development. We established a highly efficient neural induction strategy and an improved analytical platform, and determined proteomic and phosphoproteomic profiles of hESCs and their specified multipotent neural stem cell derivatives (hNSCs). This quantitative dataset (nearly 13,000 proteins and 60,000 phosphorylation sites) provides unique molecular insights into pluripotency and neural lineage entry. Systems-level comparative analysis of proteins (e.g., transcription factors, epigenetic regulators, kinase families), phosphorylation sites, and numerous biological pathways allowed the identification of distinct signatures in pluripotent and multipotent cells. Furthermore, as predicted by the dataset, we functionally validated an autocrine/paracrine mechanism by demonstrating that the secreted protein midkine is a regulator of neural specification. This resource is freely available to the scientific community, including a searchable website, PluriProt. Published by Elsevier Inc.

Long-range Electrostatic Complementarity Governs Substrate Recognition by Human Chymotrypsin C, a Key Regulator of Digestive Enzyme Activation*

PubMed Central

Batra, Jyotica; Szabó, András; Caulfield, Thomas R.; Soares, Alexei S.; Sahin-Tóth, Miklós; Radisky, Evette S.

2013-01-01

Human chymotrypsin C (CTRC) is a pancreatic serine protease that regulates activation and degradation of trypsinogens and procarboxypeptidases by targeting specific cleavage sites within their zymogen precursors. In cleaving these regulatory sites, which are characterized by multiple flanking acidic residues, CTRC shows substrate specificity that is distinct from that of other isoforms of chymotrypsin and elastase. Here, we report the first crystal structure of active CTRC, determined at 1.9-Å resolution, revealing the structural basis for binding specificity. The structure shows human CTRC bound to the small protein protease inhibitor eglin c, which binds in a substrate-like manner filling the S6-S5′ subsites of the substrate binding cleft. Significant binding affinity derives from burial of preferred hydrophobic residues at the P1, P4, and P2′ positions of CTRC, although acidic P2′ residues can also be accommodated by formation of an interfacial salt bridge. Acidic residues may also be specifically accommodated in the P6 position. The most unique structural feature of CTRC is a ring of intense positive electrostatic surface potential surrounding the primarily hydrophobic substrate binding site. Our results indicate that long-range electrostatic attraction toward substrates of concentrated negative charge governs substrate discrimination, which explains CTRC selectivity in regulating active digestive enzyme levels. PMID:23430245

Monoclonal Antibodies, Derived from Humans Vaccinated with the RV144 HIV Vaccine Containing the HVEM Binding Domain of Herpes Simplex Virus (HSV) Glycoprotein D, Neutralize HSV Infection, Mediate Antibody-Dependent Cellular Cytotoxicity, and Protect Mice from Ocular Challenge with HSV-1.

PubMed

Wang, Kening; Tomaras, Georgia D; Jegaskanda, Sinthujan; Moody, M Anthony; Liao, Hua-Xin; Goodman, Kyle N; Berman, Phillip W; Rerks-Ngarm, Supachai; Pitisuttithum, Punnee; Nitayapan, Sorachai; Kaewkungwal, Jaranit; Haynes, Barton F; Cohen, Jeffrey I

2017-10-01

The RV144 HIV vaccine trial included a recombinant HIV glycoprotein 120 (gp120) construct fused to a small portion of herpes simplex virus 1 (HSV-1) glycoprotein D (gD) so that the first 40 amino acids of gp120 were replaced by the signal sequence and the first 27 amino acids of the mature form of gD. This region of gD contains most of the binding site for HVEM, an HSV receptor important for virus infection of epithelial cells and lymphocytes. RV144 induced antibodies to HIV that were partially protective against infection, as well as antibodies to HSV. We derived monoclonal antibodies (MAbs) from peripheral blood B cells of recipients of the RV144 HIV vaccine and showed that these antibodies neutralized HSV-1 infection in cells expressing HVEM, but not the other major virus receptor, nectin-1. The MAbs mediated antibody-dependent cellular cytotoxicity (ADCC), and mice that received the MAbs and were then challenged by corneal inoculation with HSV-1 had reduced eye disease, shedding, and latent infection. To our knowledge, this is the first description of MAbs derived from human recipients of a vaccine that specifically target the HVEM binding site of gD. In summary, we found that monoclonal antibodies derived from humans vaccinated with the HVEM binding domain of HSV-1 gD (i) neutralized HSV-1 infection in a cell receptor-specific manner, (ii) mediated ADCC, and (iii) reduced ocular disease in virus-infected mice. IMPORTANCE Herpes simplex virus 1 (HSV-1) causes cold sores and neonatal herpes and is a leading cause of blindness. Despite many trials, no HSV vaccine has been approved. Nectin-1 and HVEM are the two major cellular receptors for HSV. These receptors are expressed at different levels in various tissues, and the role of each receptor in HSV pathogenesis is not well understood. We derived human monoclonal antibodies from persons who received the HIV RV144 vaccine that contained the HVEM binding domain of HSV-1 gD fused to HIV gp120. These antibodies were able to specifically neutralize HSV-1 infection in vitro via HVEM. Furthermore, we showed for the first time that HVEM-specific HSV-1 neutralizing antibodies protect mice from HSV-1 eye disease, indicating the critical role of HVEM in HSV-1 ocular infection. Copyright © 2017 American Society for Microbiology.

Peptidyl prolyl isomerase Pin1-inhibitory activity of D-glutamic and D-aspartic acid derivatives bearing a cyclic aliphatic amine moiety.

PubMed

Nakagawa, Hidehiko; Seike, Suguru; Sugimoto, Masatoshi; Ieda, Naoya; Kawaguchi, Mitsuyasu; Suzuki, Takayoshi; Miyata, Naoki

2015-12-01

Pin1 is a peptidyl prolyl isomerase that specifically catalyzes cis-trans isomerization of phosphorylated Thr/Ser-Pro peptide bonds in substrate proteins and peptides. Pin1 is involved in many important cellular processes, including cancer progression, so it is a potential target of cancer therapy. We designed and synthesized a novel series of Pin1 inhibitors based on a glutamic acid or aspartic acid scaffold bearing an aromatic moiety to provide a hydrophobic surface and a cyclic aliphatic amine moiety with affinity for the proline-binding site of Pin1. Glutamic acid derivatives bearing cycloalkylamino and phenylthiazole groups showed potent Pin1-inhibitory activity comparable with that of known inhibitor VER-1. The results indicate that steric interaction of the cyclic alkyl amine moiety with binding site residues plays a key role in enhancing Pin1-inhibitory activity. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Estimation of Source and Attenuation Parameters from Ground Motion Observations for Induced Seismicity in Alberta

NASA Astrophysics Data System (ADS)

Novakovic, M.; Atkinson, G. M.

2015-12-01

We use a generalized inversion to solve for site response, regional source and attenuation parameters, in order to define a region-specific ground-motion prediction equation (GMPE) from ground motion observations in Alberta, following the method of Atkinson et al. (2015 BSSA). The database is compiled from over 200 small to moderate seismic events (M 1 to 4.2) recorded at ~50 regional stations (distances from 30 to 500 km), over the last few years; almost all of the events have been identified as being induced by oil and gas activity. We remove magnitude scaling and geometric spreading functions from observed ground motions and invert for stress parameter, regional attenuation and site amplification. Resolving these parameters allows for the derivation of a regionally-calibrated GMPE that can be used to accurately predict amplitudes across the region in real time, which is useful for ground-motion-based alerting systems and traffic light protocols. The derived GMPE has further applications for the evaluation of hazards from induced seismicity.

Redundancy of primary RNA-binding functions of the bacterial transcription terminator Rho.

PubMed

Shashni, Rajesh; Qayyum, M Zuhaib; Vishalini, V; Dey, Debashish; Sen, Ranjan

2014-09-01

The bacterial transcription terminator, Rho, terminates transcription at half of the operons. According to the classical model derived from in vitro assays on a few terminators, Rho is recruited to the transcription elongation complex (EC) by recognizing specific sites (rut) on the nascent RNA. Here, we explored the mode of in vivo recruitment process of Rho. We show that sequence specific recognition of the rut site, in majority of the Rho-dependent terminators, can be compromised to a great extent without seriously affecting the genome-wide termination function as well as the viability of Escherichia coli. These terminators function optimally only through a NusG-assisted recruitment and activation of Rho. Our data also indicate that at these terminators, Rho-EC-bound NusG interaction facilitates the isomerization of Rho into a translocase-competent form by stabilizing the interactions of mRNA with the secondary RNA binding site, thereby overcoming the defects of the primary RNA binding functions. © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research.

Synthesis, Activity and Structural Analysis of Novel α-Hydroxytropolone Inhibitors of Human Immunodeficiency Virus Reverse Transcriptase-Associated Ribonuclease H

PubMed Central

Chung, Suhman; Himmel, Daniel M.; Jiang, Jian-Kang; Wojtak, Krzysztof; Bauman, Joseph D.; Rausch, Jason W.; Wilson, Jennifer A.; Beutler, John A.; Thomas, Craig J.; Arnold, Eddy; Le Grice, Stuart F.J.

2011-01-01

The α-hydroxytroplone, manicol (5,7-dihydroxy-2-isopropenyl-9-methyl-1,2,3,4-tetrahydro-benzocyclohepten-6-one) potently and specifically inhibits ribonuclease H (RNase H) activity of human immunodeficiency virus reverse transcriptase (HIV RT) in vitro. However, manicol was ineffective in reducing virus replication in culture. Ongoing efforts to improve the potency and specificity over the lead compound led us to synthesize 14 manicol derivatives that retain the divalent metal-chelating α-hydroxytropolone pharmacophore. These efforts were augmented by a high resolution structure of p66/p51 HIV-1 RT containing the nonnucleoside reverse transcriptase inhibitor (NNRTI), TMC278 and manicol in the DNA polymerase and RNase H active sites, respectively. We demonstrate here that several modified α-hydroxytropolones exhibit antiviral activity at non-cytotoxic concentrations. Inclusion of RNase H active site mutants indicated that manicol analogs can occupy an additional site in or around the DNA polymerase catalytic center. Collectively, our studies will promote future structure-based design of improved α-hydroxytropolones to complement the NRTI and NNRTI currently in clinical use. PMID:21568335

When is Mass Spectrometry Combined with Affinity Approaches Essential? A Case Study of Tyrosine Nitration in Proteins

NASA Astrophysics Data System (ADS)

Petre, Brînduşa-Alina; Ulrich, Martina; Stumbaum, Mihaela; Bernevic, Bogdan; Moise, Adrian; Döring, Gerd; Przybylski, Michael

2012-11-01

Tyrosine nitration in proteins occurs under physiologic conditions and is increased at disease conditions associated with oxidative stress, such as inflammation and Alzheimer's disease. Identification and quantification of tyrosine-nitrations are crucial for understanding nitration mechanism(s) and their functional consequences. Mass spectrometry (MS) is best suited to identify nitration sites, but is hampered by low stabilities and modification levels and possible structural changes induced by nitration. In this insight, we discuss methods for identifying and quantifying nitration sites by proteolytic affinity extraction using nitrotyrosine (NT)-specific antibodies, in combination with electrospray-MS. The efficiency of this approach is illustrated by identification of specific nitration sites in two proteins in eosinophil granules from several biological samples, eosinophil-cationic protein (ECP) and eosinophil-derived neurotoxin (EDN). Affinity extraction combined with Edman sequencing enabled the quantification of nitration levels, which were found to be 8 % and 15 % for ECP and EDN, respectively. Structure modeling utilizing available crystal structures and affinity studies using synthetic NT-peptides suggest a tyrosine nitration sequence motif comprising positively charged residues in the vicinity of the NT- residue, located at specific surface- accessible sites of the protein structure. Affinities of Tyr-nitrated peptides from ECP and EDN to NT-antibodies, determined by online bioaffinity- MS, provided nanomolar KD values. In contrast, false-positive identifications of nitrations were obtained in proteins from cystic fibrosis patients upon using NT-specific antibodies, and were shown to be hydroxy-tyrosine modifications. These results demonstrate affinity- mass spectrometry approaches to be essential for unequivocal identification of biological tyrosine nitrations.

Methylated site display (MSD)-AFLP, a sensitive and affordable method for analysis of CpG methylation profiles.

PubMed

Aiba, Toshiki; Saito, Toshiyuki; Hayashi, Akiko; Sato, Shinji; Yunokawa, Harunobu; Maruyama, Toru; Fujibuchi, Wataru; Kurita, Hisaka; Tohyama, Chiharu; Ohsako, Seiichiroh

2017-03-09

It has been pointed out that environmental factors or chemicals can cause diseases that are developmental in origin. To detect abnormal epigenetic alterations in DNA methylation, convenient and cost-effective methods are required for such research, in which multiple samples are processed simultaneously. We here present methylated site display (MSD), a unique technique for the preparation of DNA libraries. By combining it with amplified fragment length polymorphism (AFLP) analysis, we developed a new method, MSD-AFLP. Methylated site display libraries consist of only DNAs derived from DNA fragments that are CpG methylated at the 5' end in the original genomic DNA sample. To test the effectiveness of this method, CpG methylation levels in liver, kidney, and hippocampal tissues of mice were compared to examine if MSD-AFLP can detect subtle differences in the levels of tissue-specific differentially methylated CpGs. As a result, many CpG sites suspected to be tissue-specific differentially methylated were detected. Nucleotide sequences adjacent to these methyl-CpG sites were identified and we determined the methylation level by methylation-sensitive restriction endonuclease (MSRE)-PCR analysis to confirm the accuracy of AFLP analysis. The differences of the methylation level among tissues were almost identical among these methods. By MSD-AFLP analysis, we detected many CpGs showing less than 5% statistically significant tissue-specific difference and less than 10% degree of variability. Additionally, MSD-AFLP analysis could be used to identify CpG methylation sites in other organisms including humans. MSD-AFLP analysis can potentially be used to measure slight changes in CpG methylation level. Regarding the remarkable precision, sensitivity, and throughput of MSD-AFLP analysis studies, this method will be advantageous in a variety of epigenetics-based research.

Focused library with a core structure extracted from natural products and modified: application to phosphatase inhibitors and several biochemical findings.

PubMed

Hirai, Go; Sodeoka, Mikiko

2015-05-19

Synthesis of a focused library is an important strategy to create novel modulators of specific classes of proteins. Compounds in a focused library are composed of a common core structure and different diversity structures. In this Account, we describe our design and synthesis of libraries focused on selective inhibitors of protein phosphatases (PPases). We considered that core structures having structural and electronic features similar to those of PPase substrates, phosphate esters, would be a reasonable choice. Therefore, we extracted core structures from natural products already identified as PPase inhibitors. Since many PPases share similar active-site structures, such phosphate-mimicking core structures should interact with many enzymes in the same family, and therefore the choice of diversity structures is pivotal both to increase the binding affinity and to achieve specificity for individual enzymes. Here we present case studies of application of focused libraries to obtain PPase inhibitors, covering the overall process from selection of core structures to identification and evaluation of candidates in the focused libraries. To synthesize a library focused on protein serine-threonine phosphatases (PPs), we chose norcantharidin as a core structure, because norcantharidin dicarboxylate shows a broad inhibition profile toward several PPs. From the resulting focused library, we identified a highly selective PP2B inhibitor, NCA-01. On the other hand, to find inhibitors of dual-specificity protein phosphatases (DSPs), we chose 3-acyltetronic acid extracted from natural product RK-682 as a core structure, because its structure resembles the transition state in the dephosphorylation reaction of DSPs. However, a highly selective inhibitor was not found in the resulting focused library. Furthermore, an inherent drawback of compounds having the highly acidic 3-acyltetronic acid as a core structure is very weak potency in cellulo, probably due to poor cell membrane permeability. Therefore, we next modified the core structure from acidic to neutral by transformation to the enamine derivative and constructed a second-generation focused library (RE derivatives). The resulting compounds showed dramatically improved cell membrane permeability and inhibitory selectivity and included VHR (vaccinia VH1-related)-selective RE12 and CDC25A/B (cell division cycle 25A/B)-selective RE44. These inhibitors act on target enzymes in cellulo and do not generate reactive oxygen species, which is a potential problem with quinoid-type inhibitors of CDC25s. The cellular activity of RE12 was further improved by replacement of the side chain to afford RE176, which showed more potent antiproliferative activity than RE12 against HeLa cells. The dramatic change of inhibitory selectivity obtained by core structure modification from 3-acyltetronic acid to its enamine derivative was associated with a change in the mode of action. Namely, RE derivatives were found to be noncompetitive inhibitors with respect to a small-molecular substrate of CDC25A/B, whereas RK-682 was a competitive inhibitor of VHR. We identified the binding site of RE derivatives on the CDC25A as a pocket adjacent to the active site; this appears to be a promising target site for development of further novel inhibitors of CDC25s.

Evaluating the potential for site-specific modification of LiDAR DEM derivatives to improve environmental planning-scale wetland identification using Random Forest classification

NASA Astrophysics Data System (ADS)

O'Neil, Gina L.; Goodall, Jonathan L.; Watson, Layne T.

2018-04-01

Wetlands are important ecosystems that provide many ecological benefits, and their quality and presence are protected by federal regulations. These regulations require wetland delineations, which can be costly and time-consuming to perform. Computer models can assist in this process, but lack the accuracy necessary for environmental planning-scale wetland identification. In this study, the potential for improvement of wetland identification models through modification of digital elevation model (DEM) derivatives, derived from high-resolution and increasingly available light detection and ranging (LiDAR) data, at a scale necessary for small-scale wetland delineations is evaluated. A novel approach of flow convergence modelling is presented where Topographic Wetness Index (TWI), curvature, and Cartographic Depth-to-Water index (DTW), are modified to better distinguish wetland from upland areas, combined with ancillary soil data, and used in a Random Forest classification. This approach is applied to four study sites in Virginia, implemented as an ArcGIS model. The model resulted in significant improvement in average wetland accuracy compared to the commonly used National Wetland Inventory (84.9% vs. 32.1%), at the expense of a moderately lower average non-wetland accuracy (85.6% vs. 98.0%) and average overall accuracy (85.6% vs. 92.0%). From this, we concluded that modifying TWI, curvature, and DTW provides more robust wetland and non-wetland signatures to the models by improving accuracy rates compared to classifications using the original indices. The resulting ArcGIS model is a general tool able to modify these local LiDAR DEM derivatives based on site characteristics to identify wetlands at a high resolution.

A tiered approach for the human health risk assessment for consumption of vegetables from with cadmium-contaminated land in urban areas.

PubMed

Swartjes, Frank A; Versluijs, Kees W; Otte, Piet F

2013-10-01

Consumption of vegetables that are grown in urban areas takes place worldwide. In developing countries, vegetables are traditionally grown in urban areas for cheap food supply. In developing and developed countries, urban gardening is gaining momentum. A problem that arises with urban gardening is the presence of contaminants in soil, which can be taken up by vegetables. In this study, a scientifically-based and practical procedure has been developed for assessing the human health risks from the consumption of vegetables from cadmium-contaminated land. Starting from a contaminated site, the procedure follows a tiered approach which is laid out as follows. In Tier 0, the plausibility of growing vegetables is investigated. In Tier 1 soil concentrations are compared with the human health-based Critical soil concentration. Tier 2 offers the possibility for a detailed site-specific human health risk assessment in which calculated exposure is compared to the toxicological reference dose. In Tier 3, vegetable concentrations are measured and tested following a standardized measurement protocol. To underpin the derivation of the Critical soil concentrations and to develop a tool for site-specific assessment the determination of the representative concentration in vegetables has been evaluated for a range of vegetables. The core of the procedure is based on Freundlich-type plant-soil relations, with the total soil concentration and the soil properties as variables. When a significant plant-soil relation is lacking for a specific vegetable a geometric mean of BioConcentrationFactors (BCF) is used, which is normalized according to soil properties. Subsequently, a 'conservative' vegetable-group-consumption-rate-weighted BioConcentrationFactor is calculated as basis for the Critical soil concentration (Tier 1). The tool to perform site-specific human health risk assessment (Tier 2) includes the calculation of a 'realistic worst case' site-specific vegetable-group-consumption-rate-weighted BioConcentrationFactor. © 2013 Elsevier Inc. All rights reserved.

Comparison of unmanned aircraft systems (UAS) to LiDAR for streambank erosion measurement at the site-specific and river network scales

NASA Astrophysics Data System (ADS)

Hamshaw, S. D.; Dewoolkar, M. M.; Rizzo, D.; ONeil-Dunne, J.; Frolik, J.

2016-12-01

Measurement of rates and extent of streambank erosion along river corridors is an important component of many catchment studies and necessary for engineering projects such as river restoration, hazard assessment, and total maximum daily load (TMDL) development. A variety of methods have been developed to quantify streambank erosion, including bank pins, ground surveys, photogrammetry, LiDAR, and analytical models. However, these methods are not only resource intensive, but many are feasible and appropriate only for site-specific studies and not practical for erosion estimates at larger scales. Recent advancements in unmanned aircraft systems (UAS) and photogrammetry software provide capabilities for more rapid and economical quantification of streambank erosion and deposition at multiple scales (from site-specific to river network). At the site-specific scale, the capability of UAS to quantify streambank erosion was compared to terrestrial laser scanning (TLS) and RTK-GPS ground survey and assessed at seven streambank monitoring sites in central Vermont. Across all sites, the UAS-derived bank topography had mean errors of 0.21 m compared to TLS and GPS data. Highest accuracies were achieved in early spring conditions where mean errors approached 10 cm. The cross sectional area of bank erosion at a typical, vegetated streambank site was found to be reliably calculated within 10% of actual for erosion areas greater than 3.5 m2. At the river network-level scale, 20 km of river corridor along the New Haven, Winooski, and Mad Rivers was flown on multiple dates with UAS and used to generate digital elevation models (DEMs) that were then compared for change detection analysis. Airborne LiDAR data collected prior to UAS surveys was also compared to UAS data to determine multi-year rates of bank erosion. UAS-based photogrammetry for generation of fine scale topographic data shows promise for the monitoring of streambank erosion both at the individual site scale and river-network scale in areas that are not densely covered with vegetation year-round.

Neuroanatomical morphometric characterization of sex differences in youth using statistical learning.

PubMed

Sepehrband, Farshid; Lynch, Kirsten M; Cabeen, Ryan P; Gonzalez-Zacarias, Clio; Zhao, Lu; D'Arcy, Mike; Kesselman, Carl; Herting, Megan M; Dinov, Ivo D; Toga, Arthur W; Clark, Kristi A

2018-05-15

Exploring neuroanatomical sex differences using a multivariate statistical learning approach can yield insights that cannot be derived with univariate analysis. While gross differences in total brain volume are well-established, uncovering the more subtle, regional sex-related differences in neuroanatomy requires a multivariate approach that can accurately model spatial complexity as well as the interactions between neuroanatomical features. Here, we developed a multivariate statistical learning model using a support vector machine (SVM) classifier to predict sex from MRI-derived regional neuroanatomical features from a single-site study of 967 healthy youth from the Philadelphia Neurodevelopmental Cohort (PNC). Then, we validated the multivariate model on an independent dataset of 682 healthy youth from the multi-site Pediatric Imaging, Neurocognition and Genetics (PING) cohort study. The trained model exhibited an 83% cross-validated prediction accuracy, and correctly predicted the sex of 77% of the subjects from the independent multi-site dataset. Results showed that cortical thickness of the middle occipital lobes and the angular gyri are major predictors of sex. Results also demonstrated the inferential benefits of going beyond classical regression approaches to capture the interactions among brain features in order to better characterize sex differences in male and female youths. We also identified specific cortical morphological measures and parcellation techniques, such as cortical thickness as derived from the Destrieux atlas, that are better able to discriminate between males and females in comparison to other brain atlases (Desikan-Killiany, Brodmann and subcortical atlases). Copyright © 2018 Elsevier Inc. All rights reserved.

Common Data Model for Neuroscience Data and Data Model Exchange

PubMed Central

Gardner, Daniel; Knuth, Kevin H.; Abato, Michael; Erde, Steven M.; White, Thomas; DeBellis, Robert; Gardner, Esther P.

2001-01-01

Objective: Generalizing the data models underlying two prototype neurophysiology databases, the authors describe and propose the Common Data Model (CDM) as a framework for federating a broad spectrum of disparate neuroscience information resources. Design: Each component of the CDM derives from one of five superclasses—data, site, method, model, and reference—or from relations defined between them. A hierarchic attribute-value scheme for metadata enables interoperability with variable tree depth to serve specific intra- or broad inter-domain queries. To mediate data exchange between disparate systems, the authors propose a set of XML-derived schema for describing not only data sets but data models. These include biophysical description markup language (BDML), which mediates interoperability between data resources by providing a meta-description for the CDM. Results: The set of superclasses potentially spans data needs of contemporary neuroscience. Data elements abstracted from neurophysiology time series and histogram data represent data sets that differ in dimension and concordance. Site elements transcend neurons to describe subcellular compartments, circuits, regions, or slices; non-neuroanatomic sites include sequences to patients. Methods and models are highly domain-dependent. Conclusions: True federation of data resources requires explicit public description, in a metalanguage, of the contents, query methods, data formats, and data models of each data resource. Any data model that can be derived from the defined superclasses is potentially conformant and interoperability can be enabled by recognition of BDML-described compatibilities. Such metadescriptions can buffer technologic changes. PMID:11141510

The alpha-fetoprotein (AFP) third domain: a search for AFP interaction sites of cell cycle proteins.

PubMed

Mizejewski, G J

2016-09-01

The carboxy-terminal third domain of alpha-fetoprotein (AFP-3D) is known to harbor binding and/or interaction sites for hydrophobic ligands, receptors, and binding proteins. Such reports have established that AFP-3D consists of amino acid (AA) sequence stretches on the AFP polypeptide that engages in protein-to-protein interactions with various ligands and receptors. Using a computer software program specifically designed for such interactions, the present report identified AA sequence fragments on AFP-3D that could potentially interact with a variety of cell cycle proteins. The cell cycle proteins identified were (1) cyclins, (2) cyclin-dependent kinases, (3) cell cycle-associated proteins (inhibitors, checkpoints, initiators), and (4) ubiquitin ligases. Following detection of the AFP-3D to cell cycle protein interaction sites, the computer-derived AFP localization AA sequences were compared and aligned with previously reported hydrophobic ligand and receptor interaction sites on AFP-3D. A literature survey of the association of cell cycle proteins with AFP showed both positive relationships and correlations. Previous reports of experimental AFP-derived peptides effects on various cell cycle proteins served to confirm and verify the present computer cell cycle protein identifications. Cell cycle protein interactions with AFP-CD peptides have been reported in cultured MCF-7 breast cancer cells subjected to mRNA microarray analysis. After 7 days in culture with MCF-7 cells, the AFP-derived peptides were shown to downregulate cyclin E, SKP2, checkpoint suppressors, cyclin-dependent kinases, and ubiquitin ligases that modulate cyclin E/CdK2 transition from the G1 to the S-phase of the cell cycle. Thus, the experimental data on AFP-CD interaction with cell cycle proteins were consistent with the "in silico" findings.

Nuclear magnetic resonance studies of phenylalanine analog interactions with normal and sicklen hemoglobin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Y.H.

Several phenylalanine derivatives have been found to inhibit the gelation of deoxygenated sickle hemoglobin (deoxy HbS). Proton and /sup 19/F-NMR techniques were used to monitor the interaction of selected phenylalanine derivatives with the Hb molecule by using fluorine containing phenylalanine derivatives, Hb labeled at the ..beta..93 position with N-(2,2,2-trifluoroethyl) iodoacetamide (IA-F/sub 3/), and by monitoring the relaxation rates of the C2 and C4 histidine protons. The results show that the /sup 19/F spin-spin relaxation times of L-phenylalanin-4-fluorobenzylamide (PheNBz1-F), which has a deoxy HbS antigelling activity comparable to that of the amino acid, tryptophan, are affected much more strongly by interactionmore » with Hb than are those of glycin-4-fluorobenzylamide (GlyNBz1-F). In contrast, it is shown that N-(2,2,5,5-tetramethylpyrrolidin-1-oxy-3-carboxyl)-L-phenylalanine t-butyl ester (SL-Phe) exhibits specific binding to Hb, and an antigelling activity more than two orders of magnitude greater than that of phenylalanine. These results indicate that the fluorine nuclei strongly influenced by the presence of spin label nitroxide are located in a conformation within a few angstroms of the SL-Phe binding site. Proton NMR relaxation measurements of the C2 and C4 proton resonances from the ..beta..2, 4b143 and ..beta..146 histidine residues show significant and selective effects from the binding of SL-Phe to Hb, indicating that the SL-Phe binding site must be close to the side chains of these three residues. The strong antigelation activity of SL-Phe suggests that this binding site may be one of the intermolecular contact sites of importance to the deoxy HbS aggregation process.« less

VEGETATION COVER ANALYSIS OF HAZARDOUS WASTE SITES IN UTAH AND ARIZONA USING HYPERSPECTRAL REMOTE SENSING

DOE Office of Scientific and Technical Information (OSTI.GOV)

Serrato, M.; Jungho, I.; Jensen, J.

2012-01-17

Remote sensing technology can provide a cost-effective tool for monitoring hazardous waste sites. This study investigated the usability of HyMap airborne hyperspectral remote sensing data (126 bands at 2.3 x 2.3 m spatial resolution) to characterize the vegetation at U.S. Department of Energy uranium processing sites near Monticello, Utah and Monument Valley, Arizona. Grass and shrub species were mixed on an engineered disposal cell cover at the Monticello site while shrub species were dominant in the phytoremediation plantings at the Monument Valley site. The specific objectives of this study were to: (1) estimate leaf-area-index (LAI) of the vegetation using threemore » different methods (i.e., vegetation indices, red-edge positioning (REP), and machine learning regression trees), and (2) map the vegetation cover using machine learning decision trees based on either the scaled reflectance data or mixture tuned matched filtering (MTMF)-derived metrics and vegetation indices. Regression trees resulted in the best calibration performance of LAI estimation (R{sup 2} > 0.80). The use of REPs failed to accurately predict LAI (R{sup 2} < 0.2). The use of the MTMF-derived metrics (matched filter scores and infeasibility) and a range of vegetation indices in decision trees improved the vegetation mapping when compared to the decision tree classification using just the scaled reflectance. Results suggest that hyperspectral imagery are useful for characterizing biophysical characteristics (LAI) and vegetation cover on capped hazardous waste sites. However, it is believed that the vegetation mapping would benefit from the use of 1 higher spatial resolution hyperspectral data due to the small size of many of the vegetation patches (< 1m) found on the sites.« less

Spatial and temporal variability in the amount and source of dissolved organic carbon: Implications for ultraviolet exposure in amphibian habitats

USGS Publications Warehouse

Brooks, P.D.; O'Reilly, C. M.; Diamond, S.A.; Campbell, D.H.; Knapp, R.; Bradford, D.; Corn, P.S.; Hossack, B.; Tonnessen, K.

2005-01-01

The amount, chemical composition, and source of dissolved organic carbon (DOC), together with in situ ultraviolet (UV-B) attenuation, were measured at 1–2 week intervals throughout the summers of 1999, 2000, and 2001 at four sites in Rocky Mountain National Park (Colorado). Eight additional sites, four in Sequoia and Kings Canyon National Park/John Muir Wilderness (California) and four in Glacier National Park (Montana), were sampled during the summer of 2000. Attenuation of UV-B was significantly related to DOC concentrations over the three years in Rocky Mountain (R2 = 0.39, F = 25.71, P < 0.0001) and across all parks in 2000 (R2 = 0.44, F = 38.25, P < 0.0001). The relatively low R2 values, however, reflect significant temporal and spatial variability in the specific attenuation per unit DOC. Fluorescence analysis of the fulvic acid DOC fraction (roughly 600–2,000 Daltons) indicated that the source of DOC significantly affected the attenuation of UV-B. Sites in Sequoia–Kings Canyon were characterized by DOC derived primarily from algal sources and showed much deeper UV-B penetration, whereas sites in Glacier and Rocky Mountain contained a mix of algal and terrestrial DOC-dominated sites, with more terrestrially dominated sites characterized by greater UV-B attenuation per unit DOC. In general, site characteristics that promoted the accumulation of terrestrially derived DOC showed greater attenuation of UV-B per unit DOC; however, catchment vegetation and soil characteristics, precipitation, and local hydrology interacted to make it difficult to predict potential exposure from DOC concentrations.

Temperature Regimes Impact Coral Assemblages along Environmental Gradients on Lagoonal Reefs in Belize

PubMed Central

Townsend, Joseph E.; Courtney, Travis A.; Aichelman, Hannah E.; Davies, Sarah W.; Lima, Fernando P.; Castillo, Karl D.

2016-01-01

Coral reefs are increasingly threatened by global and local anthropogenic stressors such as rising seawater temperature, nutrient enrichment, sedimentation, and overfishing. Although many studies have investigated the impacts of local and global stressors on coral reefs, we still do not fully understand how these stressors influence coral community structure, particularly across environmental gradients on a reef system. Here, we investigate coral community composition across three different temperature and productivity regimes along a nearshore-offshore gradient on lagoonal reefs of the Belize Mesoamerican Barrier Reef System (MBRS). A novel metric was developed using ultra-high-resolution satellite-derived estimates of sea surface temperatures (SST) to classify reefs as exposed to low (lowTP), moderate (modTP), or high (highTP) temperature parameters over 10 years (2003 to 2012). Coral species richness, abundance, diversity, density, and percent cover were lower at highTP sites relative to lowTP and modTP sites, but these coral community traits did not differ significantly between lowTP and modTP sites. Analysis of coral life history strategies revealed that highTP sites were dominated by hardy stress-tolerant and fast-growing weedy coral species, while lowTP and modTP sites consisted of competitive, generalist, weedy, and stress-tolerant coral species. Satellite-derived estimates of Chlorophyll-a (chl-a) were obtained for 13-years (2003–2015) as a proxy for primary production. Chl-a concentrations were highest at highTP sites, medial at modTP sites, and lowest at lowTP sites. Notably, thermal parameters correlated better with coral community traits between site types than productivity, suggesting that temperature (specifically number of days above the thermal bleaching threshold) played a greater role in defining coral community structure than productivity on the MBRS. Dominance of weedy and stress-tolerant genera at highTP sites suggests that corals utilizing these two life history strategies may be better suited to cope with warmer oceans and thus may warrant protective status under climate change. PMID:27606598

Temperature Regimes Impact Coral Assemblages along Environmental Gradients on Lagoonal Reefs in Belize.

PubMed

Baumann, Justin H; Townsend, Joseph E; Courtney, Travis A; Aichelman, Hannah E; Davies, Sarah W; Lima, Fernando P; Castillo, Karl D

2016-01-01

Coral reefs are increasingly threatened by global and local anthropogenic stressors such as rising seawater temperature, nutrient enrichment, sedimentation, and overfishing. Although many studies have investigated the impacts of local and global stressors on coral reefs, we still do not fully understand how these stressors influence coral community structure, particularly across environmental gradients on a reef system. Here, we investigate coral community composition across three different temperature and productivity regimes along a nearshore-offshore gradient on lagoonal reefs of the Belize Mesoamerican Barrier Reef System (MBRS). A novel metric was developed using ultra-high-resolution satellite-derived estimates of sea surface temperatures (SST) to classify reefs as exposed to low (lowTP), moderate (modTP), or high (highTP) temperature parameters over 10 years (2003 to 2012). Coral species richness, abundance, diversity, density, and percent cover were lower at highTP sites relative to lowTP and modTP sites, but these coral community traits did not differ significantly between lowTP and modTP sites. Analysis of coral life history strategies revealed that highTP sites were dominated by hardy stress-tolerant and fast-growing weedy coral species, while lowTP and modTP sites consisted of competitive, generalist, weedy, and stress-tolerant coral species. Satellite-derived estimates of Chlorophyll-a (chl-a) were obtained for 13-years (2003-2015) as a proxy for primary production. Chl-a concentrations were highest at highTP sites, medial at modTP sites, and lowest at lowTP sites. Notably, thermal parameters correlated better with coral community traits between site types than productivity, suggesting that temperature (specifically number of days above the thermal bleaching threshold) played a greater role in defining coral community structure than productivity on the MBRS. Dominance of weedy and stress-tolerant genera at highTP sites suggests that corals utilizing these two life history strategies may be better suited to cope with warmer oceans and thus may warrant protective status under climate change.

Contacts between the factor TUF and RPG sequences.

PubMed

Vignais, M L; Huet, J; Buhler, J M; Sentenac, A

1990-08-25

The yeast TUF factor binds specifically to RPG-like sequences involved in multiple functions at enhancers, silencers, and telomeres. We have characterized the interaction of TUF with its optimal binding sequence, rpg-1 (1-ACACCCATACATTT-14), using a gel DNA-binding assay in combination with methylation protection and mutagenesis experiments. As many as 10 base pairs appear to be engaged in factor binding. Analysis of a collection of 30 different RPG mutants demonstrated the importance of 8 base pairs at position 2, 3, 4, 5, 6, 7, 10, and 12 and the critical role of the central GC pair at position 5. Methylation protection data on four different natural sites confirmed a close contact at positions 4, 5, 6, and 10 and suggested additional contacts at base pairs 8, 12, and 13. The derived consensus sequence was RCAAYCCRYNCAYY. A quantitative band shift analysis was used to determine the equilibrium dissociation constant for the complex of TUF and its optimal binding site rpg-1. The specific dissociation constant (K8) was found to be 1.3 x 10(-11) M. The comparison of the K8 value with the dissociation constant obtained for nonspecific DNA sites (Kn8 = 8.7 x 10(-6) M) shows the high binding selectivity of TUF for its specific RPG target.

Site-specific genome editing for correction of induced pluripotent stem cells derived from dominant dystrophic epidermolysis bullosa.

PubMed

Shinkuma, Satoru; Guo, Zongyou; Christiano, Angela M

2016-05-17

Genome editing with engineered site-specific endonucleases involves nonhomologous end-joining, leading to reading frame disruption. The approach is applicable to dominant negative disorders, which can be treated simply by knocking out the mutant allele, while leaving the normal allele intact. We applied this strategy to dominant dystrophic epidermolysis bullosa (DDEB), which is caused by a dominant negative mutation in the COL7A1 gene encoding type VII collagen (COL7). We performed genome editing with TALENs and CRISPR/Cas9 targeting the mutation, c.8068_8084delinsGA. We then cotransfected Cas9 and guide RNA expression vectors expressed with GFP and DsRed, respectively, into induced pluripotent stem cells (iPSCs) generated from DDEB fibroblasts. After sorting, 90% of the iPSCs were edited, and we selected four gene-edited iPSC lines for further study. These iPSCs were differentiated into keratinocytes and fibroblasts secreting COL7. RT-PCR and Western blot analyses revealed gene-edited COL7 with frameshift mutations degraded at the protein level. In addition, we confirmed that the gene-edited truncated COL7 could neither associate with normal COL7 nor undergo triple helix formation. Our data establish the feasibility of mutation site-specific genome editing in dominant negative disorders.

Strategic deployment of CHO expression platforms to deliver Pfizer's Monoclonal Antibody Portfolio.

PubMed

Scarcelli, John J; Shang, Tanya Q; Iskra, Tim; Allen, Martin J; Zhang, Lin

2017-11-01

Development of stable cell lines for expression of large-molecule therapeutics represents a significant portion of the time and effort required to advance a molecule to enabling regulatory toxicology studies and clinical evaluation. Our development strategy employs two different approaches for cell line development based on the needs of a particular project: a random integration approach for projects where high-level expression is critical, and a site-specific integration approach for projects in which speed and reduced employee time spend is a necessity. Here we describe both our random integration and site-specific integration platforms and their applications in support of monoclonal antibody development and production. We also compare product quality attributes of monoclonal antibodies produced with a nonclonal cell pool or clonal cell lines derived from the two platforms. Our data suggests that material source (pools vs. clones) does not significantly alter the examined product quality attributes. Our current practice is to leverage this observation with our site-specific integration platform, where material generated from cell pools is used for an early molecular assessment of a given candidate to make informed decisions around development strategy. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:1463-1467, 2017. © 2017 American Institute of Chemical Engineers.

Deriving Albedo from Coupled MERIS and MODIS Surface Products

NASA Technical Reports Server (NTRS)

Gao, Feng; Schaaf, Crystal; Jin, Yu-Fang; Lucht, Wolfgang; Strahler, Alan

2004-01-01

MERIS Level 2 surface reflectance products are now available to the scientific community. This paper demonstrates the production of MERIS-derived surface albedo and Nadir Bidirectional Reflectance Distribution Function (BRDF) adjusted reflectances by coupling the MERIS data with MODIS BRDF products. Initial efforts rely on the specification of surface anisotropy as provided by the global MODIS BRDF product for a first guess of the shape of the BRDF and then make use all of the coincidently available, partially atmospherically corrected, cloud cleared, MERIS observations to generate MERIS-derived BRDF and surface albedo quantities for each location. Comparisons between MODIS (aerosol-corrected) and MERIS (not-yet aerosol-corrected) surface values from April and May 2003 are also presented for case studies in Spain and California as well as preliminary comparisons with field data from the Devil's Rock Surfrad/BSRN site.

The 3-amino-derivative of gamma-cyclodextrin as chiral selector of Dns-amino acids in electrokinetic chromatography.

PubMed

Giuffrida, A; Contino, A; Maccarrone, G; Messina, M; Cucinotta, V

2009-04-24

The enantioseparation of the enantiomeric pairs of 10 Dns derivatives of alpha-amino acids was successfully carried out by using for the first time the 3-amino derivative of the gamma-cyclodextrin. The effects of pH and selector concentration on the migration times and the resolutions of analytes were studied in detail. 3-Deoxy-3-amino-2(S),3(R)-gamma-cyclodextrin (GCD3AM) shows very good chiral recognition ability even at very low concentrations at all the three investigated values of pH, as shown by the very large values of selectivity and resolution towards several pairs of amino acids. The role played by the cavity, the substitution site and the protonation equilibria on the observed properties of chiral selectivity, on varying the specific amino acid involved, is discussed.

A computational model of the nicotinic acetylcholine binding site

NASA Astrophysics Data System (ADS)

Gálvez-ruano, Enrique; Iriepa-Canalda, Isabel; Morreale, Antonio; Lipkowitz, Kenny B.

1999-01-01

We have derived a model of the nicotinic acetylcholine binding site. This was accomplished by using three known agonists (acetylcholine, nicotine and epibatidine) as templates around which polypeptide side chains, found to be part of the receptor cavity from published molecular biology studies, are allowed to flow freely in molecular dynamics simulations and mold themselves around these templates. The resulting supramolecular complex should thus be a complement, both in terms of steric effects as well as electronic effects, to the agonists and it should be a good estimation of the true receptor cavity structure. The shapes of those minireceptor cavities equilibrated rapidly on the simulation time scale and their structural congruence is very high, implying that a satisfactory model of the nicotinic acetylcholine binding site has been achieved. The computational methodology was internally tested against two rigid and specific antagonists (dihydro-β-erytroidine and erysoidine), that are expected to give rise to a somewhat differently shaped binding site compared to that derived from the agonists. Using these antagonists as templates there were structural reorganizations of the initial receptor cavities leading to distinctly different cavities compared to agonists. This indicates that adequate times and temperatures were used in our computational protocols to achieve equilibrium structures for the agonists. Overall, both minireceptor geometries for agonists and antagonists are similar with the exception of one amino acid (ARG209).

Critical review of decision support tools for sustainability assessment of site remediation options.

PubMed

Huysegoms, Lies; Cappuyns, Valérie

2017-07-01

In Europe alone, there are more than 2,5 million potentially contaminated sites of which 14% are expected to require remediation. Contaminated soil and groundwater can cause damage to human health as well as to valuable ecosystems. Globally more attention has been paid to this problem of soil contamination in the past decades. For example, more than 58 000 sites have been remediated in Europe between 2006 and 2011. Together with this increase in remediation projects there has been a surge in the development of new remediation technologies and decision support tools to be able to match every site and its specific characteristics to the best possible remediation alternative. In the past years the development of decision support tools (DST) has evolved in a more sustainable direction. Several DSTs added the claim not only to denote effective or technologically and economically feasible remediation alternatives but also to point out the more or most sustainable remediation alternatives. These trends in the evaluation of site remediation options left users with a confusing clew of possibly applicable tools to assist them in decision making for contaminated site remediation. This review provides a structured overview on the extent decision support tools for contaminated site remediation, that claim to assist in choosing the most sustainable remediation alternative, actually include the different elements of sustainability proposed in our assessment framework. The review contains an in-depth analysis of thirteen tools specifically developed to assess the sustainability of site remediation alternatives. This analysis is based on six criteria derived from the definition of sustainable development of the Brundtland report. The six criteria were concretized by using the three pillars of sustainability, applied to site remediation according to the SuRF-UK framework, two criteria derived from Life Cycle Assessment and Cost-Benefit Analysis, and an 'User friendly' criterion. These elements come together in a framework, drafted for this study, containing six criteria covering the environmental, economic, social, time, uncertainty aspects and user friendliness of a sustainable site remediation. The main remarks uncovered by this review are the imbalance of used indicators still expressing a strong preference for the environmental aspect at the expense of the economic and social aspects of sustainability, the lack of consistency in the terminology used within the field and the failure in adapting released tools to recent legislation or scientific advancements. Copyright © 2017 Elsevier Ltd. All rights reserved.

Glycyl-L-Glutamine: A Dipeptide Neurotransmitter Derived from Beta- Endorphin

DTIC Science & Technology

1994-03-31

pentobarbital anesthetized rats 15 min after 8-endorphin injection. S-Endorphin (0.5 nmol) followed by saline injection produced a rapid and sustained reduction ...glutamine did not influence the reduction in plasma pH caused by B-endorphin, however. When administered icy to rats that had not been pretreated...into specific thermoregulatory sites the medial preoptic area (mPOA) of the hypothalamus (Resch and Simpson, 1991). This finding provided us with an

Use of Landsat-derived temporal profiles for corn-soybean feature extraction and classification

NASA Technical Reports Server (NTRS)

Badhwar, G. D.; Carnes, J. G.; Austin, W. W.

1982-01-01

A physical model is presented, which has been derived from multitemporal-multispectral data acquired by Landsat satellites to describe the behavior and new features that are crop specific. A feasibility study over 40 sites was performed to classify the segment pixels into those of corn, soybeans, and others using the new features and a linear classifier. Results agree well with other existing methods, and it is shown the multitemporal-multispectral scanner data can be transformed into two parameters that are closely related to the target of interest and thus can be used in classification. The approach is less time intensive than other techniques and requires labeling of only pure pixels.

Emerging biological roles for erythropoietin in the nervous system.

PubMed

Brines, Michael; Cerami, Anthony

2005-06-01

Erythropoietin mediates an evolutionarily conserved, ancient immune response that limits damage to the heart, the nervous system and other tissues following injury. New evidence indicates that erythropoietin specifically prevents the destruction of viable tissue surrounding the site of an injury by signalling through a non-haematopoietic receptor. Engineered derivatives of erythropoietin that have a high affinity for this receptor have been developed, and these show robust tissue-protective effects in diverse preclinical models without stimulating erythropoiesis. A recent successful proof-of-concept clinical trial that used erythropoietin to treat human patients who had suffered a stroke encourages the evaluation of both this cytokine and non-erythropoietic derivatives as therapeutic agents to limit tissue injury.

Evaluating multicenter DTI data in Huntington's disease on site specific effects: An ex post facto approach☆

PubMed Central

Müller, Hans-Peter; Grön, Georg; Sprengelmeyer, Reiner; Kassubek, Jan; Ludolph, Albert C.; Hobbs, Nicola; Cole, James; Roos, Raymund A.C.; Duerr, Alexandra; Tabrizi, Sarah J.; Landwehrmeyer, G. Bernhard; Süssmuth, Sigurd D.

2013-01-01

Purpose Assessment of the feasibility to average diffusion tensor imaging (DTI) metrics of MRI data acquired in the course of a multicenter study. Materials and methods Sixty-one early stage Huntington's disease patients and forty healthy controls were studied using four different MR scanners at four European sites with acquisition protocols as close as possible to a given standard protocol. The potential and feasibility of averaging data acquired at different sites was evaluated quantitatively by region-of-interest (ROI) based statistical comparisons of coefficients of variation (CV) across centers, as well as by testing for significant group-by-center differences on averaged fractional anisotropy (FA) values between patients and controls. In addition, a whole-brain based statistical between-group comparison was performed using FA maps. Results The ex post facto statistical evaluation of CV and FA-values in a priori defined ROIs showed no differences between sites above chance indicating that data were not systematically biased by center specific factors. Conclusion Averaging FA-maps from DTI data acquired at different study sites and different MR scanner types does not appear to be systematically biased. A suitable recipe for testing on the possibility to pool multicenter DTI data is provided to permit averaging of DTI-derived metrics to differentiate patients from healthy controls at a larger scale. PMID:24179771

Computational active site analysis of molecular pathways to improve functional classification of enzymes.

PubMed

Ozyurt, A Sinem; Selby, Thomas L

2008-07-01

This study describes a method to computationally assess the function of homologous enzymes through small molecule binding interaction energy. Three experimentally determined X-ray structures and four enzyme models from ornithine cyclo-deaminase, alanine dehydrogenase, and mu-crystallin were used in combination with nine small molecules to derive a function score (FS) for each enzyme-model combination. While energy values varied for a single molecule-enzyme combination due to differences in the active sites, we observe that the binding energies for the entire pathway were proportional for each set of small molecules investigated. This proportionality of energies for a reaction pathway appears to be dependent on the amino acids in the active site and their direct interactions with the small molecules, which allows a function score (FS) to be calculated to assess the specificity of each enzyme. Potential of mean force (PMF) calculations were used to obtain the energies, and the resulting FS values demonstrate that a measurement of function may be obtained using differences between these PMF values. Additionally, limitations of this method are discussed based on: (a) larger substrates with significant conformational flexibility; (b) low homology enzymes; and (c) open active sites. This method should be useful in accurately predicting specificity for single enzymes that have multiple steps in their reactions and in high throughput computational methods to accurately annotate uncharacterized proteins based on active site interaction analysis. 2008 Wiley-Liss, Inc.

Knowledge-transfer learning for prediction of matrix metalloprotease substrate-cleavage sites.

PubMed

Wang, Yanan; Song, Jiangning; Marquez-Lago, Tatiana T; Leier, André; Li, Chen; Lithgow, Trevor; Webb, Geoffrey I; Shen, Hong-Bin

2017-07-18

Matrix Metalloproteases (MMPs) are an important family of proteases that play crucial roles in key cellular and disease processes. Therefore, MMPs constitute important targets for drug design, development and delivery. Advanced proteomic technologies have identified type-specific target substrates; however, the complete repertoire of MMP substrates remains uncharacterized. Indeed, computational prediction of substrate-cleavage sites associated with MMPs is a challenging problem. This holds especially true when considering MMPs with few experimentally verified cleavage sites, such as for MMP-2, -3, -7, and -8. To fill this gap, we propose a new knowledge-transfer computational framework which effectively utilizes the hidden shared knowledge from some MMP types to enhance predictions of other, distinct target substrate-cleavage sites. Our computational framework uses support vector machines combined with transfer machine learning and feature selection. To demonstrate the value of the model, we extracted a variety of substrate sequence-derived features and compared the performance of our method using both 5-fold cross-validation and independent tests. The results show that our transfer-learning-based method provides a robust performance, which is at least comparable to traditional feature-selection methods for prediction of MMP-2, -3, -7, -8, -9 and -12 substrate-cleavage sites on independent tests. The results also demonstrate that our proposed computational framework provides a useful alternative for the characterization of sequence-level determinants of MMP-substrate specificity.

Thermodynamic analysis of Bacillus subtilis endospore protonation using isothermal titration calorimetry

NASA Astrophysics Data System (ADS)

Harrold, Zoë R.; Gorman-Lewis, Drew

2013-05-01

Bacterial proton and metal adsorption reactions have the capacity to affect metal speciation and transport in aqueous environments. We coupled potentiometric titration and isothermal titration calorimetry (ITC) analyses to study Bacillus subtilis spore-proton adsorption. We modeled the potentiometric data using a four and five-site non-electrostatic surface complexation model (NE-SCM). Heats of spore surface protonation from coupled ITC analyses were used to determine site specific enthalpies of protonation based on NE-SCMs. The five-site model resulted in a substantially better model fit for the heats of protonation but did not significantly improve the potentiometric titration model fit. The improvement observed in the five-site protonation heat model suggests the presence of a highly exothermic protonation reaction circa pH 7 that cannot be resolved in the less sensitive potentiometric data. From the log Ks and enthalpies we calculated corresponding site specific entropies. Log Ks and site concentrations describing spore surface protonation are statistically equivalent to B. subtilis cell surface protonation constants. Spore surface protonation enthalpies, however, are more exothermic relative to cell based adsorption suggesting a different bonding environment. The thermodynamic parameters defined in this study provide insight on molecular scale spore-surface protonation reactions. Coupled ITC and potentiometric titrations can reveal highly exothermic, and possibly endothermic, adsorption reactions that are overshadowed in potentiometric models alone. Spore-proton adsorption NE-SCMs derived in this study provide a framework for future metal adsorption studies.

MicroRNA-363 negatively regulates the left ventricular determining transcription factor HAND1 in human embryonic stem cell-derived cardiomyocytes.

PubMed

Wagh, Vilas; Pomorski, Alexander; Wilschut, Karlijn J; Piombo, Sebastian; Bernstein, Harold S

2014-06-06

Posttranscriptional control of mRNA by microRNA (miRNA) has been implicated in the regulation of diverse biologic processes from directed differentiation of stem cells through organism development. We describe a unique pathway by which miRNA regulates the specialized differentiation of cardiomyocyte (CM) subtypes. We differentiated human embryonic stem cells (hESCs) to cardiac progenitor cells and functional CMs, and characterized the regulated expression of specific miRNAs that target transcriptional regulators of left/right ventricular-subtype specification. From >900 known human miRNAs in hESC-derived cardiac progenitor cells and functional CMs, a subset of differentially expressed cardiac miRNAs was identified, and in silico analysis predicted highly conserved binding sites in the 3'-untranslated regions (3'UTRs) of Hand-and-neural-crest-derivative-expressed (HAND) genes 1 and 2 that are involved in left and right ventricular development. We studied the temporal and spatial expression patterns of four miRNAs in differentiating hESCs, and found that expression of miRNA (miR)-363, miR-367, miR-181a, and miR-181c was specific for stage and site. Further analysis showed that miR-363 overexpression resulted in downregulation of HAND1 mRNA and protein levels. A dual luciferase reporter assay demonstrated functional interaction of miR-363 with the full-length 3'UTR of HAND1. Expression of anti-miR-363 in-vitro resulted in enrichment for HAND1-expressing CM subtype populations. We also showed that BMP4 treatment induced the expression of HAND2 with less effect on HAND1, whereas miR-363 overexpression selectively inhibited HAND1. These data show that miR-363 negatively regulates the expression of HAND1 and suggest that suppression of miR-363 could provide a novel strategy for generating functional left-ventricular CMs.

Site-specific characterization of envelope protein N-glycosylation on Sanofi Pasteur's tetravalent CYD dengue vaccine.

PubMed

Dubayle, Jean; Vialle, Sandrine; Schneider, Diane; Pontvianne, Jérémy; Mantel, Nathalie; Adam, Olivier; Guy, Bruno; Talaga, Philippe

2015-03-10

Recently, several virus studies have shown that protein glycosylation play a fundamental role in the virus-host cell interaction. Glycosylation characterization of the envelope proteins in both insect and mammalian cell-derived dengue virus (DENV) has established that two potential glycosylation residues, the asparagine 67 and 153 can potentially be glycosylated. Moreover, it appears that the glycosylation of these two residues can influence dramatically the virus production and the infection spreading in either mosquito or mammalian cells. The Sanofi Pasteur tetravalent dengue vaccine (CYD) consists of four chimeric viruses produced in mammalian vero cells. As DENV, the CYDs are able to infect human monocyte-derived dendritic cells in vitro via C-type lectins cell-surface molecules. Despite the importance of this interaction, the specific glycosylation pattern of the DENV has not been clearly documented so far. In this paper, we investigated the structure of the N-linked glycans in the four CYD serotypes. Using MALDI-TOF analysis, the N-linked glycans of CYDs were found to be a mix of high-mannose, hybrid and complex glycans. Site-specific N-glycosylation analysis of CYDs using nanoLC-ESI-MS/MS demonstrates that both asparagine residues 67 and 153 are glycosylated. Predominant glycoforms at asparagine 67 are high mannose-type structures while mainly complex- and hybrid-type structures are detected at asparagine 153. In vitro studies have shown that the immunological consequences of infection by the CYD dengue viruses 1-4 versus the wild type parents are comparable in human monocyte-derived dendritic cells. Our E-protein glycan characterizations of CYD are consistent with those observations from the wild type parents and thus support in vitro studies. In addition, these data provide new insights for the role of glycans in the dengue virus-host cell interactions. Copyright © 2015 Elsevier Ltd. All rights reserved.

Using dual classifications in the development of avian wetland indices of biological integrity for wetlands in West Virginia, USA.

PubMed

Veselka, Walter; Anderson, James T; Kordek, Walter S

2010-05-01

Considerable resources are being used to develop and implement bioassessment methods for wetlands to ensure that "biological integrity" is maintained under the United States Clean Water Act. Previous research has demonstrated that avian composition is susceptible to human impairments at multiple spatial scales. Using a site-specific disturbance gradient, we built avian wetland indices of biological integrity (AW-IBI) specific to two wetland classification schemes, one based on vegetative structure and the other based on the wetland's position in the landscape and sources of water. The resulting class-specific AW-IBI was comprised of one to four metrics that varied in their sensitivity to the disturbance gradient. Some of these metrics were specific to only one of the classification schemes, whereas others could discriminate varying levels of disturbance regardless of classification scheme. Overall, all of the derived biological indices specific to the vegetative structure-based classes of wetlands had a significant relation with the disturbance gradient; however, the biological index derived for floodplain wetlands exhibited a more consistent response to a local disturbance gradient. We suspect that the consistency of this response is due to the inherent nature of the connectivity of available habitat in floodplain wetlands.

Generation of TALE-Based Designer Epigenome Modifiers.

PubMed

Nitsch, Sandra; Mussolino, Claudio

2018-01-01

Manipulation of gene expression can be facilitated by editing the genome or the epigenome. Precise genome editing is traditionally achieved by using designer nucleases which are generally exploited to eliminate a specific gene product. Upon the introduction of a site-specific DNA double-strand break (DSB) by the nuclease, endogenous DSB repair mechanisms are in turn harnessed to induce DNA sequence changes that can result in target gene inactivation. Minimal off-target effects can be obtained by endowing designer nucleases with the highly specific DNA-binding domain (DBD) derived from transcription activator-like effectors (TALEs). In contrast, epigenome editing allows gene expression control without inducing changes in the DNA sequence by specifically altering epigenetic marks, as histone tails modifications or DNA methylation patterns within promoter or enhancer regions. Importantly, this approach allows both up- and downregulation of the target gene expression, and the effect is generally reversible. TALE-based designer epigenome modifiers combine the high specificity of TALE-derived DBDs with the power of epigenetic modifier domains to induce fast and long-lasting changes in the epigenetic landscape of a target gene and control its expression. Here we provide a detailed description for the generation of TALE-based designer epigenome modifiers and of a suitable reporter cell line to easily monitor their activity.

Continuous directed evolution of aminoacyl-tRNA synthetases

PubMed Central

Bryson, David I.; Fan, Chenguang; Guo, Li-Tao; Miller, Corwin; Söll, Dieter; Liu, David R.

2017-01-01

Directed evolution of orthogonal aminoacyl-tRNA synthetases (AARSs) enables site-specific installation of non-canonical amino acids (ncAAs) into proteins. Traditional evolution techniques typically produce AARSs with greatly reduced activity and selectivity compared to their wild-type counterparts. We designed phage-assisted continuous evolution (PACE) selections to rapidly produce highly active and selective orthogonal AARSs through hundreds of generations of evolution. PACE of a chimeric Methanosarcina spp. pyrrolysyl-tRNA synthetase (PylRS) improved its enzymatic efficiency (kcat/KMtRNA) 45-fold compared to the parent enzyme. Transplantation of the evolved mutations into other PylRS-derived synthetases improved yields of proteins containing non-canonical residues up to 9.7-fold. Simultaneous positive and negative selection PACE over 48 h greatly improved the selectivity of a promiscuous Methanocaldococcus jannaschii tyrosyl-tRNA synthetase variant for site-specific incorporation of p-iodo-L-phenylalanine. These findings offer new AARSs that increase the utility of orthogonal translation systems and establish the capability of PACE to efficiently evolve orthogonal AARSs with high activity and amino acid specificity. PMID:29035361

Human and livestock waste as a reduced carbon source contributing to the release of arsenic to shallow Bangladesh groundwater.

PubMed

Whaley-Martin, K J; Mailloux, B J; van Geen, A; Bostick, B C; Ahmed, K M; Choudhury, I; Slater, G F

2017-10-01

Recent studies have demonstrated that the supply of relatively young organic carbon stimulates the release of arsenic to groundwater in Bangladesh. This study explores the potential role of human and livestock waste as a significant source of this carbon in a densely populated rural area with limited sanitation. Profiles of aquifer sediment samples were analyzed for phytosterols and coprostanol to assess the relative contributions of plant-derived and human/livestock waste-derived organic carbon at two well-characterized sites in Araihazar. Coprostanol concentrations increased with depth from non-detection (<10m at Site B and <23m at Site F) to maxima of 1.3 and 0.5ng/g in aquifer sands recovered from 17m (Site B) and 26m (Site F), respectively. The commonly used sewage contamination index ([5β-coprostanol]/([5α-cholestanol]+[5β-coprostanol])) exceeds 0.7 between 12 and 19m at Site B and between 24 and 26m at Site F, indicating input of human/livestock waste to these depths. Urine/fecal input within the same depth range is supported by groundwater Cl/Br mass ratios >1000 compared to Cl/Br <500 at depths >50m. Installed tube wells in the area's study sites may act as a conduit for DOC and specifically human/livestock waste into the aquifer during flood events. The depth range of maximum input of human/livestock waste indicated by these independent markers coincides with the highest dissolved Fe (10-20mg/L) and As (200-400μg/L) concentrations in groundwater at both sites. The new findings suggest that the oxidation of human/livestock waste coupled to the reductive dissolution of iron-(oxy)-hydroxides and/or arsenate may enhance groundwater contamination with As. Copyright © 2017. Published by Elsevier B.V.

Defining indicators and standards for tourism impacts in protected areas: Cape Range National Park, Australia.

PubMed

Moore, Susan A; Polley, Amanda

2007-03-01

Visitors' perceptions of impacts and acceptable standards for environmental conditions can provide essential information for the sustainable management of tourist destinations, especially protected areas. To this end, visitor surveys were administered during the peak visitor season in Cape Range National Park, on the northwest coast of Western Australia and adjacent to the iconic Ningaloo Reef. The central focus was visitors' perceptions regarding environmental conditions and standards for potential indicators. Conditions considered of greatest importance in determining visitors' quality of experience included litter, inadequate disposal of human waste, presence of wildlife, levels of noise, and access to beach and ocean. Standards were determined, based on visitors' perceptions, for a range of site-specific and non-site-specific indicators, with standards for facilities (e.g., acceptable number of parking bays, signs) and for negative environmental impacts (e.g., levels of littering, erosion) sought. The proposed standards varied significantly between sites for the facilities indicators; however, there was no significant difference between sites for environmental impacts. For the facilities, the standards proposed by visitors were closely related to the existing situation, suggesting that they were satisfied with the status quo. These results are considered in the context of current research interest in the efficacy of visitor-derived standards as a basis for protected area management.

AMES Stereo Pipeline Derived DEM Accuracy Experiment Using LROC-NAC Stereopairs and Weighted Spatial Dependence Simulation for Lunar Site Selection

NASA Astrophysics Data System (ADS)

Laura, J. R.; Miller, D.; Paul, M. V.

2012-03-01

An accuracy assessment of AMES Stereo Pipeline derived DEMs for lunar site selection using weighted spatial dependence simulation and a call for outside AMES derived DEMs to facilitate a statistical precision analysis.

a Comparitive Study Using Geometric and Vertical Profile Features Derived from Airborne LIDAR for Classifying Tree Genera

NASA Astrophysics Data System (ADS)

Ko, C.; Sohn, G.; Remmel, T. K.

2012-07-01

We present a comparative study between two different approaches for tree genera classification using descriptors derived from tree geometry and those derived from the vertical profile analysis of LiDAR point data. The different methods provide two perspectives for processing LiDAR point clouds for tree genera identification. The geometric perspective analyzes individual tree crowns in relation to valuable information related to characteristics of clusters and line segments derived within crowns and overall tree shapes to highlight the spatial distribution of LiDAR points within the crown. Conversely, analyzing vertical profiles retrieves information about the point distributions with respect to height percentiles; this perspective emphasizes of the importance that point distributions at specific heights express, accommodating for the decreased point density with respect to depth of canopy penetration by LiDAR pulses. The targeted species include white birch, maple, oak, poplar, white pine and jack pine at a study site northeast of Sault Ste. Marie, Ontario, Canada.

Site-specific O-Glycosylation Analysis of Human Blood Plasma Proteins*

PubMed Central

Hoffmann, Marcus; Marx, Kristina; Reichl, Udo; Wuhrer, Manfred; Rapp, Erdmann

2016-01-01

Site-specific glycosylation analysis is key to investigate structure-function relationships of glycoproteins, e.g. in the context of antigenicity and disease progression. The analysis, though, is quite challenging and time consuming, in particular for O-glycosylated proteins. In consequence, despite their clinical and biopharmaceutical importance, many human blood plasma glycoproteins have not been characterized comprehensively with respect to their O-glycosylation. Here, we report on the site-specific O-glycosylation analysis of human blood plasma glycoproteins. To this end pooled human blood plasma of healthy donors was proteolytically digested using a broad-specific enzyme (Proteinase K), followed by a precipitation step, as well as a glycopeptide enrichment and fractionation step via hydrophilic interaction liquid chromatography, the latter being optimized for intact O-glycopeptides carrying short mucin-type core-1 and -2 O-glycans, which represent the vast majority of O-glycans on human blood plasma proteins. Enriched O-glycopeptide fractions were subjected to mass spectrometric analysis using reversed-phase liquid chromatography coupled online to an ion trap mass spectrometer operated in positive-ion mode. Peptide identity and glycan composition were derived from low-energy collision-induced dissociation fragment spectra acquired in multistage mode. To pinpoint the O-glycosylation sites glycopeptides were fragmented using electron transfer dissociation. Spectra were annotated by database searches as well as manually. Overall, 31 O-glycosylation sites and regions belonging to 22 proteins were identified, the majority being acute-phase proteins. Strikingly, also 11 novel O-glycosylation sites and regions were identified. In total 23 O-glycosylation sites could be pinpointed. Interestingly, the use of Proteinase K proved to be particularly beneficial in this context. The identified O-glycan compositions most probably correspond to mono- and disialylated core-1 mucin-type O-glycans (T-antigen). The developed workflow allows the identification and characterization of the major population of the human blood plasma O-glycoproteome and our results provide new insights, which can help to unravel structure-function relationships. The data were deposited to ProteomeXchange PXD003270. PMID:26598643

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection

PubMed Central

Gordon, Claire L.; Thome, Joseph J.C.; Igarashi, Suzu

2017-01-01

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. PMID:28130404

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection.

PubMed

Gordon, Claire L; Miron, Michelle; Thome, Joseph J C; Matsuoka, Nobuhide; Weiner, Joshua; Rak, Michael A; Igarashi, Suzu; Granot, Tomer; Lerner, Harvey; Goodrum, Felicia; Farber, Donna L

2017-03-06

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan. @Gordon et al.

TFBSshape: a motif database for DNA shape features of transcription factor binding sites.

PubMed

Yang, Lin; Zhou, Tianyin; Dror, Iris; Mathelier, Anthony; Wasserman, Wyeth W; Gordân, Raluca; Rohs, Remo

2014-01-01

Transcription factor binding sites (TFBSs) are most commonly characterized by the nucleotide preferences at each position of the DNA target. Whereas these sequence motifs are quite accurate descriptions of DNA binding specificities of transcription factors (TFs), proteins recognize DNA as a three-dimensional object. DNA structural features refine the description of TF binding specificities and provide mechanistic insights into protein-DNA recognition. Existing motif databases contain extensive nucleotide sequences identified in binding experiments based on their selection by a TF. To utilize DNA shape information when analysing the DNA binding specificities of TFs, we developed a new tool, the TFBSshape database (available at http://rohslab.cmb.usc.edu/TFBSshape/), for calculating DNA structural features from nucleotide sequences provided by motif databases. The TFBSshape database can be used to generate heat maps and quantitative data for DNA structural features (i.e., minor groove width, roll, propeller twist and helix twist) for 739 TF datasets from 23 different species derived from the motif databases JASPAR and UniPROBE. As demonstrated for the basic helix-loop-helix and homeodomain TF families, our TFBSshape database can be used to compare, qualitatively and quantitatively, the DNA binding specificities of closely related TFs and, thus, uncover differential DNA binding specificities that are not apparent from nucleotide sequence alone.

TFBSshape: a motif database for DNA shape features of transcription factor binding sites

PubMed Central

Yang, Lin; Zhou, Tianyin; Dror, Iris; Mathelier, Anthony; Wasserman, Wyeth W.; Gordân, Raluca; Rohs, Remo

2014-01-01

Transcription factor binding sites (TFBSs) are most commonly characterized by the nucleotide preferences at each position of the DNA target. Whereas these sequence motifs are quite accurate descriptions of DNA binding specificities of transcription factors (TFs), proteins recognize DNA as a three-dimensional object. DNA structural features refine the description of TF binding specificities and provide mechanistic insights into protein–DNA recognition. Existing motif databases contain extensive nucleotide sequences identified in binding experiments based on their selection by a TF. To utilize DNA shape information when analysing the DNA binding specificities of TFs, we developed a new tool, the TFBSshape database (available at http://rohslab.cmb.usc.edu/TFBSshape/), for calculating DNA structural features from nucleotide sequences provided by motif databases. The TFBSshape database can be used to generate heat maps and quantitative data for DNA structural features (i.e., minor groove width, roll, propeller twist and helix twist) for 739 TF datasets from 23 different species derived from the motif databases JASPAR and UniPROBE. As demonstrated for the basic helix-loop-helix and homeodomain TF families, our TFBSshape database can be used to compare, qualitatively and quantitatively, the DNA binding specificities of closely related TFs and, thus, uncover differential DNA binding specificities that are not apparent from nucleotide sequence alone. PMID:24214955

DOE Office of Scientific and Technical Information (OSTI.GOV)

Pancost, R.D.; Damste, J.S.S.; Lint, S. De

Although abundant geochemical data indicate that anaerobic methane oxidation occurs in marine sediments, the linkage to specific microorganisms remains unclear. In order to examine processes of methane consumption and oxidation, sediment samples from mud volcanoes at two distinct sites on the Mediterranean consumption and oxidation, sediment samples from mud volcanoes at two distinct sites on the Mediterranean Ridge were collected via the submersible Nautile. Geochemical data strongly indicate that methane is oxidized under aerobic conditions, and compound-specific carbon isotope analyses indicate that methane is oxidized under anaerobic conditions, and compound-specific carbon isotope analyses indicate that this reaction is facilitated bymore » a consortium of archaea and bacteria. Specifically, these methane-rich sediments contain high abundances of methanogen-specific biomarkers that are significantly depleted in {sup 13}C ({delta}{sup 13}C values are as low as {minus}95%). Biomarkers inferred to derive from sulfate-reducing bacteria and other heterotrophic bacteria are similarly depleted. Consistent with previous work, such depletion can be explained by consumption of {sup 13}C-depleted methane by methanogens operating in reverse and as part a consortium of organisms in which sulfate serves as the terminal electron acceptor. Moreover, their results indicate that this process is widespread in Mediterranean mud volcanoes and in some localized settings in the predominant microbiological process.« less

Identification of human-to-human transmissibility factors in PB2 proteins of influenza A by large-scale mutual information analysis

PubMed Central

Miotto, Olivo; Heiny, AT; Tan, Tin Wee; August, J Thomas; Brusic, Vladimir

2008-01-01

Background The identification of mutations that confer unique properties to a pathogen, such as host range, is of fundamental importance in the fight against disease. This paper describes a novel method for identifying amino acid sites that distinguish specific sets of protein sequences, by comparative analysis of matched alignments. The use of mutual information to identify distinctive residues responsible for functional variants makes this approach highly suitable for analyzing large sets of sequences. To support mutual information analysis, we developed the AVANA software, which utilizes sequence annotations to select sets for comparison, according to user-specified criteria. The method presented was applied to an analysis of influenza A PB2 protein sequences, with the objective of identifying the components of adaptation to human-to-human transmission, and reconstructing the mutation history of these components. Results We compared over 3,000 PB2 protein sequences of human-transmissible and avian isolates, to produce a catalogue of sites involved in adaptation to human-to-human transmission. This analysis identified 17 characteristic sites, five of which have been present in human-transmissible strains since the 1918 Spanish flu pandemic. Sixteen of these sites are located in functional domains, suggesting they may play functional roles in host-range specificity. The catalogue of characteristic sites was used to derive sequence signatures from historical isolates. These signatures, arranged in chronological order, reveal an evolutionary timeline for the adaptation of the PB2 protein to human hosts. Conclusion By providing the most complete elucidation to date of the functional components participating in PB2 protein adaptation to humans, this study demonstrates that mutual information is a powerful tool for comparative characterization of sequence sets. In addition to confirming previously reported findings, several novel characteristic sites within PB2 are reported. Sequence signatures generated using the characteristic sites catalogue characterize concisely the adaptation characteristics of individual isolates. Evolutionary timelines derived from signatures of early human influenza isolates suggest that characteristic variants emerged rapidly, and remained remarkably stable through subsequent pandemics. In addition, the signatures of human-infecting H5N1 isolates suggest that this avian subtype has low pandemic potential at present, although it presents more human adaptation components than most avian subtypes. PMID:18315849

Analysis of Protein-RNA and Protein-Peptide Interactions in Equine Infectious Anemia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Jae-Hyung

2007-01-01

Macromolecular interactions are essential for virtually all cellular functions including signal transduction processes, metabolic processes, regulation of gene expression and immune responses. This dissertation focuses on the characterization of two important macromolecular interactions involved in the relationship between Equine Infectious Anemia Virus (EIAV) and its host cell in horse: (1) the interaction between the EIAV Rev protein and its binding site, the Rev-responsive element (RRE) and (2) interactions between equine MHC class I molecules and epitope peptides derived from EIAV proteins. EIAV, one of the most divergent members of the lentivirus family, has a single-stranded RNA genome and carries severalmore » regulatory and structural proteins within its viral particle. Rev is an essential EIAV regulatory encoded protein that interacts with the viral RRE, a specific binding site in the viral mRNA. Using a combination of experimental and computational methods, the interactions between EIAV Rev and RRE were characterized in detail. EIAV Rev was shown to have a bipartite RNA binding domain contain two arginine rich motifs (ARMs). The RRE secondary structure was determined and specific structural motifs that act as cis-regulatory elements for EIAV Rev-RRE interaction were identified. Interestingly, a structural motif located in the high affinity Rev binding site is well conserved in several diverse lentiviral genoes, including HIV-1. Macromolecular interactions involved in the immune response of the horse to EIAV infection were investigated by analyzing complexes between MHC class I proteins and epitope peptides derived from EIAV Rev, Env and Gag proteins. Computational modeling results provided a mechanistic explanation for the experimental finding that a single amino acid change in the peptide binding domain of the quine MHC class I molecule differentially affectes the recognitino of specific epitopes by EIAV-specific CTL. Together, the findings in this dissertation provide novel insights into the strategy used by EIAV to replicate itself, and provide new details about how the host cell responds to and defends against EIAV upon the infection. Moreover, they have contributed to the understanding of the macromolecular recognition events that regulate these processes.« less

Value of a dual-polarized gap-filling radar in support of southern California post-fire debris-flow warnings

USGS Publications Warehouse

Jorgensen, David P.; Hanshaw, Maiana N.; Schmidt, Kevin M.; Laber, Jayme L; Staley, Dennis M.; Kean, Jason W.; Restrepo, Pedro J.

2011-01-01

A portable truck-mounted C-band Doppler weather radar was deployed to observe rainfall over the Station Fire burn area near Los Angeles, California, during the winter of 2009/10 to assist with debris-flow warning decisions. The deployments were a component of a joint NOAA–U.S. Geological Survey (USGS) research effort to improve definition of the rainfall conditions that trigger debris flows from steep topography within recent wildfire burn areas. A procedure was implemented to blend various dual-polarized estimators of precipitation (for radar observations taken below the freezing level) using threshold values for differential reflectivity and specific differential phase shift that improves the accuracy of the rainfall estimates over a specific burn area sited with terrestrial tipping-bucket rain gauges. The portable radar outperformed local Weather Surveillance Radar-1988 Doppler (WSR-88D) National Weather Service network radars in detecting rainfall capable of initiating post-fire runoff-generated debris flows. The network radars underestimated hourly precipitation totals by about 50%. Consistent with intensity–duration threshold curves determined from past debris-flow events in burned areas in Southern California, the portable radar-derived rainfall rates exceeded the empirical thresholds over a wider range of storm durations with a higher spatial resolution than local National Weather Service operational radars. Moreover, the truck-mounted C-band radar dual-polarimetric-derived estimates of rainfall intensity provided a better guide to the expected severity of debris-flow events, based on criteria derived from previous events using rain gauge data, than traditional radar-derived rainfall approaches using reflectivity–rainfall relationships for either the portable or operational network WSR-88D radars. Part of the reason for the improvement was due to siting the radar closer to the burn zone than the WSR-88Ds, but use of the dual-polarimetric variables improved the rainfall estimation by ~12% over the use of traditional Z–R relationships.

Population-Genomic Insights into Variation in Prevotella intermedia and Prevotella nigrescens Isolates and Its Association with Periodontal Disease

PubMed Central

Zhang, Yifei; Zhen, Min; Zhan, Yalin; Song, Yeqing; Zhang, Qian; Wang, Jinfeng

2017-01-01

High-throughput sequencing has helped to reveal the close relationship between Prevotella and periodontal disease, but the roles of subspecies diversity and genomic variation within this genus in periodontal diseases still need to be investigated. We performed a comparative genome analysis of 48 Prevotella intermedia and Prevotella nigrescens isolates that from the same cohort of subjects to identify the main drivers of their pathogenicity and adaptation to different environments. The comparisons were done between two species and between disease and health based on pooled sequences. The results showed that both P. intermedia and P. nigrescens have highly dynamic genomes and can take up various exogenous factors through horizontal gene transfer. The major differences between disease-derived and health-derived samples of P. intermedia and P. nigrescens were factors related to genome modification and recombination, indicating that the Prevotella isolates from disease sites may be more capable of genomic reconstruction. We also identified genetic elements specific to each sample, and found that disease groups had more unique virulence factors related to capsule and lipopolysaccharide synthesis, secretion systems, proteinases, and toxins, suggesting that strains from disease sites may have more specific virulence, particularly for P. intermedia. The differentially represented pathways between samples from disease and health were related to energy metabolism, carbohydrate and lipid metabolism, and amino acid metabolism, consistent with data from the whole subgingival microbiome in periodontal disease and health. Disease-derived samples had gained or lost several metabolic genes compared to healthy-derived samples, which could be linked with the difference in virulence performance between diseased and healthy sample groups. Our findings suggest that P. intermedia and P. nigrescens may serve as “crucial substances” in subgingival plaque, which may reflect changes in microbial and environmental dynamics in subgingival microbial ecosystems. This provides insight into the potential of P. intermedia and P. nigrescens as new predictive biomarkers and targets for effective interventions in periodontal disease. PMID:28983469

Population-Genomic Insights into Variation in Prevotella intermedia and Prevotella nigrescens Isolates and Its Association with Periodontal Disease.

PubMed

Zhang, Yifei; Zhen, Min; Zhan, Yalin; Song, Yeqing; Zhang, Qian; Wang, Jinfeng

2017-01-01

High-throughput sequencing has helped to reveal the close relationship between Prevotella and periodontal disease, but the roles of subspecies diversity and genomic variation within this genus in periodontal diseases still need to be investigated. We performed a comparative genome analysis of 48 Prevotella intermedia and Prevotella nigrescens isolates that from the same cohort of subjects to identify the main drivers of their pathogenicity and adaptation to different environments. The comparisons were done between two species and between disease and health based on pooled sequences. The results showed that both P. intermedia and P. nigrescens have highly dynamic genomes and can take up various exogenous factors through horizontal gene transfer. The major differences between disease-derived and health-derived samples of P. intermedia and P. nigrescens were factors related to genome modification and recombination, indicating that the Prevotella isolates from disease sites may be more capable of genomic reconstruction. We also identified genetic elements specific to each sample, and found that disease groups had more unique virulence factors related to capsule and lipopolysaccharide synthesis, secretion systems, proteinases, and toxins, suggesting that strains from disease sites may have more specific virulence, particularly for P. intermedia . The differentially represented pathways between samples from disease and health were related to energy metabolism, carbohydrate and lipid metabolism, and amino acid metabolism, consistent with data from the whole subgingival microbiome in periodontal disease and health. Disease-derived samples had gained or lost several metabolic genes compared to healthy-derived samples, which could be linked with the difference in virulence performance between diseased and healthy sample groups. Our findings suggest that P. intermedia and P. nigrescens may serve as "crucial substances" in subgingival plaque, which may reflect changes in microbial and environmental dynamics in subgingival microbial ecosystems. This provides insight into the potential of P. intermedia and P. nigrescens as new predictive biomarkers and targets for effective interventions in periodontal disease.

Evaluation of a maleimido derivative of CHX-A” DTPA for site-specific labeling of Affibody molecules

PubMed Central

Tolmachev, Vladimir; Xu, Heng; Wållberg, Helena; Ahlgren, Sara; Hjertman, Magnus; Sjöberg, Anna; Sandström, Mattias; Abrahmsén, Lars; Brechbiel, Martin W.; Orlova, Anna

2008-01-01

Affibody molecules are a new class of small targeting proteins based on a common threehelix bundle structure. Affibody molecules binding a desired target may be selected using phage-display technology. An Affibody molecule ZHER2:342 binding with subnanomolar affinity to the tumor antigen HER2 has recently been developed for radionuclide imaging in vivo. Introduction of a single cysteine into the cysteine-free Affibody scaffold provides a unique thiol group for site-specific labeling of recombinant Affibody molecules. The recently developed maleimido-CHX-A” DTPA was site-specifically conjugated at the C-terminal cysteine of ZHER2:2395-C, a variant of ZHER2:342, providing a homogenous conjugate with a dissociation constant of 56 pM. The yield of labeling with 111In was > 99% after 10 min at room temperature. In vitro cell tests demonstrated specific binding of 111In-CHX-A” DTPAZ2395-C to HER2-expressing cell-line SKOV-3 and good cellular retention of radioactivity. In normal mice, the conjugate demonstrated rapid clearance from all non-specific organs except kidney. In mice bearing SKOV-3 xenografts, the tumor uptake of 111In-CHX-A” DTPAZ2395-C was 17.3 ± 4.8 % IA/g and the tumor-to-blood ratio 86 ± 46 (4 h post-injection). HER2-exprssing xenografts were clearly visualized 1 h post-injection. In conclusion, coupling of maleimido-CHX-A” DTPA to cysteine-containing Affibody molecules provides welldefined uniform conjugate, which can be rapidly labeled at room temperature and provides high-contrast imaging of molecular targets in vivo. PMID:18620447

Photoaffinity labeling the substance P receptor using a derivative of substance P containing para-benzoylphenylalanine

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boyd, N.D.; White, C.F.; Leeman, S.E.

A novel photoreactive substance P (SP) analogue has been synthesized by solid-phase peptide synthesis methodology to incorporate the amino acid p-benzoyl-L-phenylalanine (L-Phe(pBz)) in place of the Phe{sup 8} residue of SP. (Phe{sup 8}(OpBz))SP was equipotent with SP in competing for SP binding sites on rat submaxillary gland membranes and had potent sialagogic activity in vivo. In the absence of light, the {sup 125}I-labeled Bolton-Hunter conjugate of (Phe{sup 8}(pBz))SP bound in a saturable and reversible manner to an apparently homogeneous class of binding sites with an affinity K{sub D} = 0.4 nM. The binding of {sup 125}I-(Phe{sup 8}(pBz))SP was inhibited competitivelymore » by various tachykinin peptides and analogues with the appropriate specificity for SP/NK-1 receptors. Upon photolysis, up to 70{percent} of the specifically bound {sup 125}I-(Phe{sup 8}(pBz))SP underwent covalent linkage to two polypeptides of M{sub r} = 53 000 and 46 000, identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Quantitative analysis of the inhibitory effects of SP and related peptides on {sup 125}I-(Phe{sup 8}(pBz))SP photoincorporation indicated that the binding sites of the two photolabeled polypeptides have the same peptide specificity, namely, that typical of NK-1-type SP receptors. Further information on the relationship between the two labeled SP binding sites was provided by enzymatic digestion studies. The highly specific and remarkably efficient photolabeling achieved with {sup 125}I-(Phe{sup 8}(pBz))SP suggests that this photoaffinity probe will be of considerable value for the characterization of the molecular structure of the SP receptor.« less

Late Middle Pleistocene hominin teeth from Panxian Dadong, South China.

PubMed

Liu, Wu; Schepartz, Lynne A; Xing, Song; Miller-Antonio, Sari; Wu, Xiujie; Trinkaus, Erik; Martinón-Torres, María

2013-05-01

The hominin teeth and evidence of hominin activities recovered from 1991 to 2005 at the Panxian Dadong site in South China are dated to the late Middle Pleistocene (MIS 8-6 or ca. 130-300 ka), a period for which very little is known about the morphology of Asian populations. The present study provides the first detailed morphometric description and comparisons of four hominin teeth (I(1), C1, P(3) and P3) from this site. Our study shows that the Panxian Dadong teeth combine archaic and derived features that align them with Middle and Upper Pleistocene fossils from East and West Asia and Europe. These teeth do not display any typical Neanderthal features and they are generally more derived than other contemporaneous populations from Asia and Africa. However, the derived traits are not diagnostic enough to specifically link the Panxian Dadong teeth to Homo sapiens, a common problem when analyzing the Middle Pleistocene dental record from Africa and Asia. These findings are contextualized in the discussion of the evolutionary course of Asian Middle Pleistocene hominins, and they highlight the necessity of incorporating the Asian fossil record in the still open debate about the origin of H. sapiens. Copyright © 2013 Elsevier Ltd. All rights reserved.

Photoaffinity labelling and solubilization on the central 5-HT1A receptor binding site.

PubMed

Gozlan, H; Emerit, M B; el Mestikawy, S; Cossery, J M; Marquet, A; Besselievre, R; Hamon, M

1987-01-01

Two complementary approaches, covalent labelling and solubilization, have been used to study the biochemical properties of the central 5-HT1A receptor binding site. We have first designed a photoaffinity ligand containing the structure of 8-OH-DPAT, a potent and specific agonist of 5-HT1A sites. Thus, 8-methoxy-2[N-n-propyl,N-3-(2-nitro-4-azido-phenyl)- aminopropyl]aminotetralin or 8-methoxy-3'-NAP-amino-PAT, was found to displace, in the dark, [3H]8-OH-DPAT from 5-HT1A sites in rat hippocampal membranes with an IC50 of 6.6 nM. Under two cumulative UV irradiations (366 nm, for 20 min at 4 degrees C), 8-methoxy-3-'-NAP-amino-PAT (30 nM) blocked irreversibly 55-60% of 5-HT1A binding sites. This blockade was specific of 5-HT1A sites since the other serotoninergic sites, 5-HT1B, 5-HT2 and also the presynaptic 5-HT3 sites were not affected by the treatment. In addition, the binding of [3H]Spiperone and [3H]7-OH-DPAT to striatal dopamine sites remained unchanged under similar photolysis conditions. The tritiated derivative of the photoaffinity ligand (92 Ci/mmol) was then synthesized for the identification of the covalently bound protein(s). SDS-PAGE of solubilized membranes irradiated in the presence of 20 nM 3H-8-methoxy-3'-NAP-amino-PAT allowed the detection of a 63 kD protein whose labelling appeared specific. Thus, 3H-incorporation into the 63 kD band could be prevented by microM concentrations of 5-HT, 8-OH-DPAT and other selective 5-HT1A ligands such as isapirone. In contrast, the 5-HT2 antagonist ketanserin, norepinephrine and dopamine-related ligands (including 7-OH-DPAT) were ineffective. Direct solubilization of 5-HT1A receptor binding sites was also attempted from rat hippocampal membranes. The best results were obtained using CHAPS (10 mM) plus NaCl (0.2 M), which led to 50% recovery of 5-HT1A sites in the 100,000 g supernatant. The pharmacological properties and sensitivity to N-ethyl-maleimide and GppNHp of soluble sites appeared near identical to those of membrane-bound 5-HT1A sites.

Cas6 is an endoribonuclease that generates guide RNAs for invader defense in prokaryotes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Carte, Jason; Wang, Ruiying; Li, Hong

An RNA-based gene silencing pathway that protects bacteria and archaea from viruses and other genome invaders is hypothesized to arise from guide RNAs encoded by CRISPR loci and proteins encoded by the cas genes. CRISPR loci contain multiple short invader-derived sequences separated by short repeats. The presence of virus-specific sequences within CRISPR loci of prokaryotic genomes confers resistance against corresponding viruses. The CRISPR loci are transcribed as long RNAs that must be processed to smaller guide RNAs. Here we identified Pyrococcus furiosus Cas6 as a novel endoribonuclease that cleaves CRISPR RNAs within the repeat sequences to release individual invader targetingmore » RNAs. Cas6 interacts with a specific sequence motif in the 5{prime} region of the CRISPR repeat element and cleaves at a defined site within the 3{prime} region of the repeat. The 1.8 angstrom crystal structure of the enzyme reveals two ferredoxin-like folds that are also found in other RNA-binding proteins. The predicted active site of the enzyme is similar to that of tRNA splicing endonucleases, and concordantly, Cas6 activity is metal-independent. cas6 is one of the most widely distributed CRISPR-associated genes. Our findings indicate that Cas6 functions in the generation of CRISPR-derived guide RNAs in numerous bacteria and archaea.« less

Estimating economic thresholds for site-specific weed control using manual weed counts and sensor technology: an example based on three winter wheat trials.

PubMed

Keller, Martina; Gutjahr, Christoph; Möhring, Jens; Weis, Martin; Sökefeld, Markus; Gerhards, Roland

2014-02-01

Precision experimental design uses the natural heterogeneity of agricultural fields and combines sensor technology with linear mixed models to estimate the effect of weeds, soil properties and herbicide on yield. These estimates can be used to derive economic thresholds. Three field trials are presented using the precision experimental design in winter wheat. Weed densities were determined by manual sampling and bi-spectral cameras, yield and soil properties were mapped. Galium aparine, other broad-leaved weeds and Alopecurus myosuroides reduced yield by 17.5, 1.2 and 12.4 kg ha(-1) plant(-1)  m(2) in one trial. The determined thresholds for site-specific weed control with independently applied herbicides were 4, 48 and 12 plants m(-2), respectively. Spring drought reduced yield effects of weeds considerably in one trial, since water became yield limiting. A negative herbicide effect on the crop was negligible, except in one trial, in which the herbicide mixture tended to reduce yield by 0.6 t ha(-1). Bi-spectral cameras for weed counting were of limited use and still need improvement. Nevertheless, large weed patches were correctly identified. The current paper presents a new approach to conducting field trials and deriving decision rules for weed control in farmers' fields. © 2013 Society of Chemical Industry.

Molecular Properties of neurotoxin receptors sites associated with sodium channels from mammalian brain.

PubMed

Catterall, W A; Hartshorne, R P; Beneski, D A

1982-01-01

Neurotoxins that act at specific receptor sites on voltage-sensitive sodium channels have been used as molecular probes to identify and purify protein components of sodium channels from mammalian brain. Photoreactive derivatives of scorpion toxin have been prepared and used to covalently label sodium channels in intact synaptosomes. Two polypeptides, alpha with Mr approximately 270,000 and beta with Mr approximately 38,000, are specifically labeled indicating that they are components of the scorpion toxin receptor site on the sodium channel. The sodium channel can be solubilized with retention of specific binding of [3H] saxitoxin using nonionic detergents such as Triton X-100. The solubilized saxitoxin receptor has molecular weight of 316,000 +/- 63,000 and binds 0.9 g of Triton X-100 and phospholipid per g of protein. The solubilized receptor can be purified 750-fold by ion exchange chromatography, wheat germ lectin/Sepharose chromatography and sucrose gradient sedimentation to a final specific activity of 1488 pmol/mg. Analysis of the polypeptide chain composition of the most highly purified fractions indicates that alpha and beta comprise 65% of the protein of these fractions and are only the polypeptides whose presence correlates with saxitoxin binding activity. These studies lead to a working hypothesis of sodium channel structure in which the intact channel is comprised of a complex with Mr of approximately 316,000 containing one mole of alpha (Mr approximately 270,000) and one to three moles of beta (Mr approximately 38,000).

Genetic dissection of TrkB activated signalling pathways required for specific aspects of the taste system

PubMed Central

2014-01-01

Background Neurotrophin-4 (NT-4) and brain derived neurotrophic factor (BDNF) bind to the same receptor, Ntrk2/TrkB, but play distinct roles in the development of the rodent gustatory system. However, the mechanisms underlying these processes are lacking. Results Here, we demonstrate, in vivo, that single or combined point mutations in major adaptor protein docking sites on TrkB receptor affect specific aspects of the mouse gustatory development, known to be dependent on BDNF or NT-4. In particular, mice with a mutation in the TrkB-SHC docking site had reduced gustatory neuron survival at both early and later stages of development, when survival is dependent on NT-4 and BDNF, respectively. In addition, lingual innervation and taste bud morphology, both BDNF-dependent functions, were altered in these mutants. In contrast, mutation of the TrkB-PLCγ docking site alone did not affect gustatory neuron survival. Moreover, innervation to the tongue was delayed in these mutants and taste receptor expression was altered. Conclusions We have genetically dissected pathways activated downstream of the TrkB receptor that are required for specific aspects of the taste system controlled by the two neurotrophins NT-4 and BDNF. In addition, our results indicate that TrkB also regulate the expression of specific taste receptors by distinct signalling pathways. These results advance our knowledge of the biology of the taste system, one of the fundamental sensory systems crucial for an organism to relate to the environment. PMID:25256039

Human 15-LOX-1 active site mutations alter inhibitor binding and decrease potency.

PubMed

Armstrong, Michelle; van Hoorebeke, Christopher; Horn, Thomas; Deschamps, Joshua; Freedman, J Cody; Kalyanaraman, Chakrapani; Jacobson, Matthew P; Holman, Theodore

2016-11-01

Human 15-lipoxygenase-1 (h15-LOX-1 or h12/15-LOX) reacts with polyunsaturated fatty acids and produces bioactive lipid derivatives that are implicated in many important human diseases. One such disease is stroke, which is the fifth leading cause of death and the first leading cause of disability in America. The discovery of h15-LOX-1 inhibitors could potentially lead to novel therapeutics in the treatment of stroke, however, little is known about the inhibitor/active site interaction. This study utilizes site-directed mutagenesis, guided in part by molecular modeling, to gain a better structural understanding of inhibitor interactions within the active site. We have generated eight mutants (R402L, R404L, F414I, F414W, E356Q, Q547L, L407A, I417A) of h15-LOX-1 to determine whether these active site residues interact with two h15-LOX-1 inhibitors, ML351 and an ML094 derivative, compound 18. IC 50 values and steady-state inhibition kinetics were determined for the eight mutants, with four of the mutants affecting inhibitor potency relative to wild type h15-LOX-1 (F414I, F414W, E356Q and L407A). The data indicate that ML351 and compound 18, bind in a similar manner in the active site to an aromatic pocket close to F414 but have subtle differences in their specific binding modes. This information establishes the binding mode for ML094 and ML351 and will be leveraged to develop next-generation inhibitors. Copyright © 2016 Elsevier Ltd. All rights reserved.

Digested sludge-derived three-dimensional hierarchical porous carbon for high-performance supercapacitor electrode.

PubMed

Zhang, Jia-Jia; Fan, Hao-Xiang; Dai, Xiao-Hu; Yuan, Shi-Jie

2018-04-01

Digested sludge, as the main by-product of the sewage sludge anaerobic digestion process, still contains considerable organic compounds. In this protocol, we report a facile method for preparing digested sludge-derived self-doped porous carbon material for high-performance supercapacitor electrodes via a sustainable pyrolysis/activation process. The obtained digested sludge-derived carbon material (HPDSC) exhibits versatile O-, N-doped hierarchical porous framework, high specific surface area (2103.6 m 2  g -1 ) and partial graphitization phase, which can facilitate ion transport, provide more storage sites for electrolyte ions and enhance the conductivity of active electrode materials. The HPDSC-based supercapacitor electrodes show favourable energy storage performance, with a specific capacitance of 245 F g -1 at 1.0 A g -1 in 0.5 M Na 2 SO 4 ; outstanding cycling stability, with 98.4% capacitance retention after 2000 cycles; and good rate performance (211 F g -1 at 11 A g -1 ). This work provides a unique self-doped three-dimensional hierarchical porous carbon material with a favourable charge storage capacity and at the same time finds a high value-added and environment-friendly strategy for disposal and recycling of digested sludge.

Digested sludge-derived three-dimensional hierarchical porous carbon for high-performance supercapacitor electrode

NASA Astrophysics Data System (ADS)

Zhang, Jia-Jia; Fan, Hao-Xiang; Dai, Xiao-Hu; Yuan, Shi-Jie

2018-04-01

Digested sludge, as the main by-product of the sewage sludge anaerobic digestion process, still contains considerable organic compounds. In this protocol, we report a facile method for preparing digested sludge-derived self-doped porous carbon material for high-performance supercapacitor electrodes via a sustainable pyrolysis/activation process. The obtained digested sludge-derived carbon material (HPDSC) exhibits versatile O-, N-doped hierarchical porous framework, high specific surface area (2103.6 m2 g-1) and partial graphitization phase, which can facilitate ion transport, provide more storage sites for electrolyte ions and enhance the conductivity of active electrode materials. The HPDSC-based supercapacitor electrodes show favourable energy storage performance, with a specific capacitance of 245 F g-1 at 1.0 A g-1 in 0.5 M Na2SO4; outstanding cycling stability, with 98.4% capacitance retention after 2000 cycles; and good rate performance (211 F g-1 at 11 A g-1). This work provides a unique self-doped three-dimensional hierarchical porous carbon material with a favourable charge storage capacity and at the same time finds a high value-added and environment-friendly strategy for disposal and recycling of digested sludge.

Tree-, stand- and site-specific controls on landscape-scale patterns of transpiration

NASA Astrophysics Data System (ADS)

Kathrin Hassler, Sibylle; Weiler, Markus; Blume, Theresa

2018-01-01

Transpiration is a key process in the hydrological cycle, and a sound understanding and quantification of transpiration and its spatial variability is essential for management decisions as well as for improving the parameterisation and evaluation of hydrological and soil-vegetation-atmosphere transfer models. For individual trees, transpiration is commonly estimated by measuring sap flow. Besides evaporative demand and water availability, tree-specific characteristics such as species, size or social status control sap flow amounts of individual trees. Within forest stands, properties such as species composition, basal area or stand density additionally affect sap flow, for example via competition mechanisms. Finally, sap flow patterns might also be influenced by landscape-scale characteristics such as geology and soils, slope position or aspect because they affect water and energy availability; however, little is known about the dynamic interplay of these controls.We studied the relative importance of various tree-, stand- and site-specific characteristics with multiple linear regression models to explain the variability of sap velocity measurements in 61 beech and oak trees, located at 24 sites across a 290 km2 catchment in Luxembourg. For each of 132 consecutive days of the growing season of 2014 we modelled the daily sap velocity and derived sap flow patterns of these 61 trees, and we determined the importance of the different controls.Results indicate that a combination of mainly tree- and site-specific factors controls sap velocity patterns in the landscape, namely tree species, tree diameter, geology and aspect. For sap flow we included only the stand- and site-specific predictors in the models to ensure variable independence. Of those, geology and aspect were most important. Compared to these predictors, spatial variability of atmospheric demand and soil moisture explains only a small fraction of the variability in the daily datasets. However, the temporal dynamics of the explanatory power of the tree-specific characteristics, especially species, are correlated to the temporal dynamics of potential evaporation. We conclude that transpiration estimates on the landscape scale would benefit from not only consideration of hydro-meteorological drivers, but also tree, stand and site characteristics in order to improve the spatial and temporal representation of transpiration for hydrological and soil-vegetation-atmosphere transfer models.

SCS-CN and GIS-based approach for identifying potential water harvesting sites in the Kali Watershed, Mahi River Basin, India

NASA Astrophysics Data System (ADS)

Ramakrishnan, D.; Bandyopadhyay, A.; Kusuma, K. N.

2009-08-01

The Kali sub-watershed is situated in the semi-arid region of Gujarat, India and forms a part of the Mahi River Watershed. This watershed receives an average annual rainfall of 900mm mainly between July and September. Due to high runoff potential, evapo-transpiration and poor infiltration, drought like situation prevails in this area from December to June almost every year. In this paper, augmentation of water resource is proposed by construction of runoff harvesting structures like check dam, percolation pond, farm pond, well and subsurface dyke. The site suitability for different water harvesting structures is determined by considering spatially varying parameters like runoff potential, slope, fracture pattern and micro-watershed area. GIS is utilised as a tool to store, analyse and integrate spatial and attribute information pertaining to runoff, slope, drainage and fracture. The runoff derived by SCS-CN method is a function of runoff potential which can be expressed in terms of runoff coefficient (ratio between the runoff and rainfall) which can be classified into three classes, viz., high (>40%), moderate (20-40%) and low (<20%). In addition to IMSD, FAO specifications for water harvesting/recharging structures, parameters such as effective storage, rock mass permeability are herein considered to augment effective storage. Using the overlay and decision tree concepts in GIS, potential water harvesting sites are identified. The derived sites are field investigated for suitability and implementation. In all, the accuracy of the site selection at implementation level varies from 80-100%.

Cross-calibration of Medium Resolution Earth Observing Satellites by Using EO-1 Hyperion-derived Spectral Surface Reflectance from "Lunar Cal Sites"

NASA Astrophysics Data System (ADS)

Ungar, S.

2017-12-01

Over the past 3 years, the Earth Observing-one (EO-1) Hyperion imaging spectrometer was used to slowly scan the lunar surface at a rate which results in up to 32X oversampling to effectively increase the SNR. Several strategies, including comparison against the USGS RObotic Lunar Observatory (ROLO) mode,l are being employed to estimate the absolute and relative accuracy of the measurement set. There is an existing need to resolve discrepancies as high as 10% between ROLO and solar based calibration of current NASA EOS assets. Although the EO-1 mission was decommissioned at the end of March 2017, the development of a well-characterized exoatmospheric spectral radiometric database, for a range of lunar phase angles surrounding the fully illuminated moon, continues. Initial studies include a comprehensive analysis of the existing 17-year collection of more than 200 monthly lunar acquisitions. Specific lunar surface areas, such as a lunar mare, are being characterized as potential "lunar calibration sites" in terms of their radiometric stability in the presence of lunar nutation and libration. Site specific Hyperion-derived lunar spectral reflectance are being compared against spectrographic measurements made during the Apollo program. Techniques developed through this activity can be employed by future high-quality orbiting imaging spectrometers (such as HyspIRI and EnMap) to further refine calibration accuracies. These techniques will enable the consistent cross calibration of existing and future earth observing systems (spectral and multi-spectral) including those that do not have lunar viewing capability. When direct lunar viewing is not an option for an earth observing asset, orbiting imaging spectrometers can serve as transfer radiometers relating that asset's sensor response to lunar values through near contemporaneous observations of well characterized stable CEOS test sites. Analysis of this dataset will lead to the development of strategies to ensure more accurate cross calibrations when employing the more capable, future imaging spectrometers.

Context-dependent environmental quality standards of soil nitrate for terrestrial plant communities.

PubMed

van Goethem, Thomas M W J; Schipper, Aafke M; Wamelink, G W Wieger; Huijbregts, Mark A J

2016-10-01

Environmental quality standards (EQS) specify the maximum permissible concentration or level of a specific environmental stressor. Here, a procedure is proposed to derive EQS that are specific to a representative species pool and conditional on confounding environmental factors. To illustrate the procedure, a dataset was used with plant species richness observations of grasslands and forests and accompanying soil nitrate-N and pH measurements collected from 981 sampling sites in the Netherlands. Species richness was related to soil nitrate-N and pH with quantile regression allowing for interaction effects. The resulting regression models were used to derive EQS for nitrate conditional on pH, quantified as the nitrate-N concentrations at a specific pH level corresponding with a species richness equal to 95% of the species pool, for both grasslands and forest communities. The EQS varied between 1.8 mg/kg nitrate-N at pH 9-65 mg/kg nitrate-N at pH 4. EQS for forests and grasslands were similar, but EQS based on Red List species richness were considerably lower (more stringent) than those based on overall species richness, particularly at high pH levels. The results indicate that both natural background pH conditions and Red List species are important factors to consider in the derivation of EQS for soil nitrate-N for terrestrial ecosystems. Copyright © 2016 Elsevier Ltd. All rights reserved.

Determination of the affinity of drugs toward serum albumin by measurement of the quenching of the intrinsic tryptophan fluorescence of the protein.

PubMed

Epps, D E; Raub, T J; Caiolfa, V; Chiari, A; Zamai, M

1999-01-01

Binding of new chemical entities to serum proteins is an issue confronting pharmaceutical companies during development of potential therapeutic agents. Most drugs bind to the most abundant plasma protein, human serum albumin (HSA), at two major binding sites. Excepting fluorescence spectroscopy, existing methods for assaying drug binding to serum albumin are insensitive to higher-affinity compounds and can be labour-intensive, time-consuming, and usually require compound-specific assays. This led us to examine alternative ways to measure drug-albumin interaction. One method described here uses fluorescence quenching of the single tryptophan (Trp) residue in HSA excited at 295 nm to measure drug-binding affinity. Unfortunately, many compounds absorb, fluoresce, or both, in this UV wavelength region of the spectrum. Several types of binding phenomenon and spectral interference were identified by use of six structurally unrelated compounds and the equations necessary to make corrections mathematically were derived and applied to calculate binding constants accurately. The general cases were: direct quenching of Trp fluorescence by optically transparent ligands with low or high affinities; binding of optically transparent, non-fluorescent ligands to two specific sites where both sites or only one site result in Trp fluorescence quenching; and chromophores whose absorption either overlaps the Trp emission and quenches by energy transfer or absorbs light at the Trp fluorescence excitation wavelength producing absorptive screening as well as fluorescence quenching. Unless identification of the site specificity of drug binding to serum albumin is desired, quenching of the Trp fluorescence of albumin by titration with ligand is a rapid and facile method for determining the binding affinities of drugs for serum albumin.

Cation disorder in Ga1212.

PubMed

Greenwood, K B; Ko, D; Vander Griend, D A; Sarjeant, G M; Milgram, J W; Garrity, E S; DeLoach, D I; Poeppelmeier, K R; Salvador, P A; Mason, T O

2000-07-24

Substitution of calcium for strontium in LnSr2-xCaxCu2GaO7 (Ln = La, Pr, Nd, Gd, Ho, Er, Tm, and Yb) materials at ambient pressure and 975 degrees C results in complete substitution of calcium for strontium in the lanthanum and praseodymium systems and partial substitution in the other lanthanide systems. The calcium saturation level depends on the size of the Ln cation, and in all cases, a decrease in the lattice parameters with calcium concentration was observed until a common, lower bound, average A-cation size is reached. Site occupancies from X-ray and neutron diffraction experiments for LnSr2-xCaxCu2GaO7 (x = 0 and x = 2) confirm that the A-cations distribute between the two blocking-layer sites and the active-layer site based on size. A quantitative link between cation distribution and relative site-specific cation enthalpy for calcium, strontium, and lanthanum within the gallate structure is derived. The cation distribution in other similar materials can potentially be modeled.

Site-specific mesenchymal control of inflammatory pain to yeast challenge in vulvodynia afflicted and pain-free women

PubMed Central

Foster, David C.; Falsetta, Megan L.; Woeller, Collynn F.; Pollock, Stephen J.; Song, Kunchang; Bonham, Adrienne; Haidaris, Constantine G.; Stodgell, Chris J.; Messing, Susan P.; Iadarola, Michael; Phipps, Richard P.

2015-01-01

Fibroblast strains were derived from two regions of the lower genital tract of localized provoked vulvodynia (LPV) cases and pain-free controls. Sixteen strains were derived from four cases and four controls, age and race matched, following pre-sampling mechanical pain threshold assessments. Strains were challenged with six separate stimuli: live yeast species (C. albicans, C. glabrata, C. tropicalis, and S. cerevisiae), yeast extract (zymosan), or inactive vehicle. Production of prostaglandin E2 (PGE2) and interleukin-6 (IL-6) were pro-inflammatory response measures. Highest IL-6 and PGE2 occurred with vestibular strains following C. albicans, C. glabrata, and zymosan challenges, resulting in the ability to significantly predict IL-6 and PGE2 production by genital tract location. Following C. albicans and C. glabrata challenge of all sixteen fibroblast strains, adjusting for dual sampling of subjects, PGE2 and IL-6 production significantly predicted the pre-sampling pain threshold from the genital tract site of sampling. At the same location of pain assessment and fibroblast sampling, in situ immunohistochemical (IHC)(+) fibroblasts for IL-6 and Cox-2 were quantified microscopically. The correlation between IL-6 production and IL-6 IHC(+) was statistically significant yet biological significance is unknown because of the small number of IHC(+) IL-6 fibroblasts identified. A low fibroblast IL-6 IHC(+) count may result from most IL-6 produced by fibroblasts existing in a secreted, extracellular state. Enhanced, site-specific, innate immune responsiveness to yeast pathogens by fibroblasts may be an early step in LPV pathogenesis. Fibroblast strain testing may offer an attractive/objective marker of LPV pathology in women with vulvodynia of inflammatory origin. PMID:25679469

Mechanism of Error-Free DNA Replication Past Lucidin-Derived DNA Damage by Human DNA Polymerase κ.

PubMed

Yockey, Oliver P; Jha, Vikash; Ghodke, Pratibha P; Xu, Tianzuo; Xu, Wenyan; Ling, Hong; Pradeepkumar, P I; Zhao, Linlin

2017-11-20

DNA damage impinges on genetic information flow and has significant implications in human disease and aging. Lucidin-3-O-primeveroside (LuP) is an anthraquinone derivative present in madder root, which has been used as a coloring agent and food additive. LuP can be metabolically converted to genotoxic compound lucidin, which subsequently forms lucidin-specific N 2 -2'-deoxyguanosine (N 2 -dG) and N 6 -2'-deoxyadenosine (N 6 -dA) DNA adducts. Lucidin is mutagenic and carcinogenic in rodents but has low carcinogenic risks in humans. To understand the molecular mechanism of low carcinogenicity of lucidin in humans, we performed DNA replication assays using site-specifically modified oligodeoxynucleotides containing a structural analogue (LdG) of lucidin-N 2 -dG DNA adduct and determined the crystal structures of DNA polymerase (pol) κ in complex with LdG-bearing DNA and an incoming nucleotide. We examined four human pols (pol η, pol ι, pol κ, and Rev1) in their efficiency and accuracy during DNA replication with LdG; these pols are key players in translesion DNA synthesis. Our results demonstrate that pol κ efficiently and accurately replicates past the LdG adduct, whereas DNA replication by pol η, pol ι is compromised to different extents. Rev1 retains its ability to incorporate dCTP opposite the lesion albeit with decreased efficiency. Two ternary crystal structures of pol κ illustrate that the LdG adduct is accommodated by pol κ at the enzyme active site during insertion and postlesion-extension steps. The unique open active site of pol κ allows the adducted DNA to adopt a standard B-form for accurate DNA replication. Collectively, these biochemical and structural data provide mechanistic insights into the low carcinogenic risk of lucidin in humans.

Site- and strand-specific nicking of DNA by fusion proteins derived from MutH and I-SceI or TALE repeats.

PubMed

Gabsalilow, Lilia; Schierling, Benno; Friedhoff, Peter; Pingoud, Alfred; Wende, Wolfgang

2013-04-01

Targeted genome engineering requires nucleases that introduce a highly specific double-strand break in the genome that is either processed by homology-directed repair in the presence of a homologous repair template or by non-homologous end-joining (NHEJ) that usually results in insertions or deletions. The error-prone NHEJ can be efficiently suppressed by 'nickases' that produce a single-strand break rather than a double-strand break. Highly specific nickases have been produced by engineering of homing endonucleases and more recently by modifying zinc finger nucleases (ZFNs) composed of a zinc finger array and the catalytic domain of the restriction endonuclease FokI. These ZF-nickases work as heterodimers in which one subunit has a catalytically inactive FokI domain. We present two different approaches to engineer highly specific nickases; both rely on the sequence-specific nicking activity of the DNA mismatch repair endonuclease MutH which we fused to a DNA-binding module, either a catalytically inactive variant of the homing endonuclease I-SceI or the DNA-binding domain of the TALE protein AvrBs4. The fusion proteins nick strand specifically a bipartite recognition sequence consisting of the MutH and the I-SceI or TALE recognition sequences, respectively, with a more than 1000-fold preference over a stand-alone MutH site. TALE-MutH is a programmable nickase.

Ebselen: Mechanisms of Glutamate Dehydrogenase and Glutaminase Enzyme Inhibition.

PubMed

Yu, Yan; Jin, Yanhong; Zhou, Jie; Ruan, Haoqiang; Zhao, Han; Lu, Shiying; Zhang, Yue; Li, Di; Ji, Xiaoyun; Ruan, Benfang Helen

2017-12-15

Ebselen modulates target proteins through redox reactions with selenocysteine/cysteine residues, or through binding to the zinc finger domains. However, a recent contradiction in ebselen inhibition of kidney type glutaminase (KGA) stimulated our interest in investigating its inhibition mechanism with glutamate dehydrogenase (GDH), KGA, thioredoxin reductase (TrxR), and glutathione S-transferase. Fluorescein- or biotin-labeled ebselen derivatives were synthesized for mechanistic analyses. Biomolecular interaction analyses showed that only GDH, KGA, and TrxR proteins can bind to the ebselen derivative, and the binding to GDH and KGA could be competed off by glutamine or glutamate. From the gel shift assays, the fluorescein-labeled ebselen derivative could co-migrate with hexameric GDH and monomeric/dimeric TrxR in a dose-dependent manner; it also co-migrated with KGA but disrupted the tetrameric form of the KGA enzyme at a high compound concentration. Further proteomic analysis demonstrated that the ebselen derivative could cross-link with proteins through a specific cysteine at the active site of GDH and TrxR proteins, but for KGA protein, the binding site is at the N-terminal appendix domain outside of the catalytic domain, which might explain why ebselen is not a potent KGA enzyme inhibitor in functional assays. In conclusion, ebselen could inhibit enzyme activity by binding to the catalytic domain or disruption of the protein complex. In addition, ebselen is a relatively potent selective GDH inhibitor that might provide potential therapeutic opportunities for hyperinsulinism-hyperammonemia syndrome patients who have the mutational loss of GTP inhibition.

Development of growth and yield models for southern hardwoods: site index determinations

Treesearch

John Paul McTague; Daniel J. Robison; David O' Loughlin; Joseph Roise; Robert Kellison

2006-01-01

Growth and yield data from across 13 southern States, collected from 1967 to 2004 from fully-stocked even-aged southern hardwood forests on a variety of site types, was used to calculate site index curves. These derived curves provide an efficient means to evaluate the productivity-age relation which varies across many sites. These curves were derived for mixed-species...

Regiospecific Ester Hydrolysis by Orange Peel Esterase - An Undergraduate Experiment.

NASA Astrophysics Data System (ADS)

Bugg, Timothy D. H.; Lewin, Andrew M.; Catlin, Eric R.

1997-01-01

A simple but effective experiment has been developed to demonstrate the regiospecificity of enzyme catalysis using an esterase activity easily isolated from orange peel. The experiment involves the preparation of diester derivatives of para-, meta- and ortho-hydroxybenzoic acid (e.g. methyl 4-acetoxy-benzoic acid). The derivatives are incubated with orange peel esterase, as a crude extract, and with commercially available pig liver esterase and porcine pancreatic lipase. The enzymatic hydrolysis reactions are monitored by thin layer chromatography, revealing which of the two ester groups is hydrolysed, and the rate of the enzyme-catalysed reaction. The results of a group experiment revealed that in all cases hydrolysis was observed with at least one enzyme, and in most cases the enzymatic hydrolysis was specific for production of either the hydroxy-ester or acyl-acid product. Specificity towards the ortho-substituted series was markedly different to that of the para-substituted series, which could be rationalised in the case of pig liver esterase by a published active site model.

Remedial investigation report on Waste Area Grouping 5 at Oak Ridge National Laboratory, Oak Ridge, Tennessee. Volume 3 -- Appendix B: Technical findings and conclusions

DOE Office of Scientific and Technical Information (OSTI.GOV)

NONE

1995-09-01

This document provides the Environmental Restoration Program with information about the results of investigations performed at Waste Area Grouping (WAG) 5. It includes information on risk assessments that have evaluated long-term impacts to human health and the environment. Information provided in this document forms the basis for decisions regarding the need for subsequent remediation work at WAG 5. Sections B1.1 through B1.4 present an overview of the environmental setting of WAG 5, including location, population, land uses, ecology, and climate, and Sects. B1.5 through B1.7 give site-specific details (e.g., topography, soils, geology, and hydrology). The remediation investigation (RI) of WAGmore » 5 did not entail en exhaustive characterization of all physical attributes of the site; the information presented here focuses on those most relevant to the development and verification of the WAG 5 conceptual model. Most of the information presented in this appendix was derived from the RI field investigation, which was designed to complement the existing data base from earlier, site-specific studies of Solid Waste Storage Area (SWSA) 5 and related areas.« less

Recombinant Human Factor IX Produced from Transgenic Porcine Milk

PubMed Central

Lee, Meng-Hwan; Lin, Yin-Shen; Tu, Ching-Fu; Yen, Chon-Ho

2014-01-01

Production of biopharmaceuticals from transgenic animal milk is a cost-effective method for highly complex proteins that cannot be efficiently produced using conventional systems such as microorganisms or animal cells. Yields of recombinant human factor IX (rhFIX) produced from transgenic porcine milk under the control of the bovine α-lactalbumin promoter reached 0.25 mg/mL. The rhFIX protein was purified from transgenic porcine milk using a three-column purification scheme after a precipitation step to remove casein. The purified protein had high specific activity and a low ratio of the active form (FIXa). The purified rhFIX had 11.9 γ-carboxyglutamic acid (Gla) residues/mol protein, which approached full occupancy of the 12 potential sites in the Gla domain. The rhFIX was shown to have a higher isoelectric point and lower sialic acid content than plasma-derived FIX (pdFIX). The rhFIX had the same N-glycosylation sites and phosphorylation sites as pdFIX, but had a higher specific activity. These results suggest that rhFIX produced from porcine milk is physiologically active and they support the use of transgenic animals as bioreactors for industrial scale production in milk. PMID:24955355

Drought effects on water quality in the South Platte River Basin, Colorado

USGS Publications Warehouse

Sprague, Lori A.

2005-01-01

Twenty-three stream sites representing a range of forested, agricultural, and urban land uses were sampled in the South Platte River Basin of Colorado from July through September 2002 to characterize water quality during drought conditions. With a few exceptions, dissolved ammonia, Kjeldahl nitrogen, total phosphorus, and dissolved orthophosphate concentrations were similar to seasonal historical levels in all land use areas during the drought. At some agricultural sites, decreased dilution of irrigation return flow may have contributed to higher concentrations of some nutrient species, increased primary productivity, and higher dissolved oxygen concentrations. At some urban sites, decreased dilution of base flow and wastewater treatment plant effluent may have contributed to higher dissolved nitrite-plus-nitrate concentrations, increased primary productivity, and higher dissolved oxygen concentrations. Total pesticide concentrations in urban and agricultural areas were not consistently higher or lower during the drought. At most forested sites, decreased dilution of ground water-derived calcium bicarbonate type base flow likely led to elevated pH and specific-conductance values. Water temperatures at many of the forested sites also were higher, contributing to lower dissolved oxygen concentrations during the drought.

Geologic implications of gas hydrates in the offshore of India: results of the National Gas Hydrate Program Expedition 01

USGS Publications Warehouse

Collett, Timothy S.; Boswell, Ray; Cochran, J.R.; Kumar, Pushpendra; Lall, Malcolm; Mazumdar, Aninda; Ramana, Mangipudi Venkata; Ramprasad, Tammisetti; Riedel, Michael; Sain, Kalachand; Sathe, Arun Vasant; Vishwanath, Krishna

2014-01-01

One of the specific objectives of this expedition was to test gas hydrate formation models and constrain model parameters, especially those that account for the formation of concentrated gas hydrate accumulations. The necessary data for characterizing the occurrence of in situ gas hydrate, such as interstitial water chlorinities, core-derived gas chemistry, physical and sedimentological properties, thermal images of the recovered cores, and downhole measured logging data (LWD and/or conventional wireline log data), were obtained from most of the drill sites established during NGHP-01. Almost all of the drill sites yielded evidence for the occurrence of gas hydrate; however, the inferred in situ concentration of gas hydrate varied substantially from site to site. For the most part, the interpretation of downhole logging data, core thermal images, interstitial water analyses, and pressure core images from the sites drilled during NGHP-01 indicate that the occurrence of concentrated gas hydrate is mostly associated with the presence of fractures in the sediments, and in some limited cases, by coarser grained (mostly sand-rich) sediments.

Fabrication and Characterization of Vertically Aligned ZnO Nanorod Arrays via Inverted Monolayer Colloidal Crystals Mask

NASA Astrophysics Data System (ADS)

Chen, Cheng; Ding, Taotao; Qi, Zhiqiang; Zhang, Wei; Zhang, Jun; Xu, Juan; Chen, Jingwen; Dai, Jiangnan; Chen, Changqing

2018-04-01

The periodically ordered ZnO nanorod (NR) arrays have been successfully synthesized via a hydrothermal approach on the silicon substrates by templating of the TiO2 ring deriving from the polystyrene (PS) nanosphere monolayer colloidal crystals (MCC). With the inverted MCC mask, sol-gel-derived ZnO seeds could serve as the periodic nucleation positions for the site-specific growth of ZnO NRs. The large-scale patterned arrays of single ZnO NR with good side-orientation can be readily produced. According to the experimental results, the as-integrated ZnO NR arrays showed an excellent crystal quality and optical property, very suitable for optoelectronic applications such as stimulated emitters and ZnO photonic crystal devices.

SINE sequences detect DNA fingerprints in salmonid fishes.

PubMed

Spruell, P; Thorgaard, G H

1996-04-01

DNA probes homologous to two previously described salmonid short interspersed nuclear elements (SINEs) detected DNA fingerprint patterns in 14 species of salmonid fishes. The probes showed more homology to some species than to others and little homology to three nonsalmonid fishes. The DNA fingerprint patterns derived from the SINE probes are individual-specific and inherited in a Mendelian manner. Probes derived from different regions of the same SINE detect only partially overlapping banding patterns, reflecting a more complex SINE structure than has been previously reported. Like the human Alu sequence, the SINEs found in salmonids could provide useful genetic markers and primer sites for PCR-based techniques. These elements may be more desirable for some applications than traditional DNA fingerprinting probes that detect tandemly repeated arrays.

Identity of a peptide domain of human C9 that is bound by the cell-surface complement inhibitor, CD59.

PubMed

Chang, C P; Hüsler, T; Zhao, J; Wiedmer, T; Sims, P J

1994-10-21

The CD59 antigen is a plasma membrane glycoprotein that serves as an inhibitor of the C5b-9 complex of complement. This inhibitory activity appears related to the capacity of CD59 to bind with high affinity to sites that are nascently exposed in the alpha-chain subunit of human C8, as well as within the C9b domain (amino acid residues 245-538) of human C9, during assembly of the C5b-9 complex on the target membrane (Ninomiya, H., and Sims, P. J. (1992) J. Biol. Chem. 267, 13675-13680). The CD59 binding site in C9 was first investigated by N-terminal sequencing of CD59-binding peptides generated by limited digest of the isolated C9b domain. These experiments revealed a 17-kDa fragment (starting at C9 residue Thr-320) that retained affinity for CD59, suggesting the possibility for localizing the CD59 binding site by mapping with small C9-derived peptides. Peptides spanning the entire C9b sequence were expressed in Escherichia coli and then probed with CD59. CD59 bound specifically to all peptides starting N-terminal to C9 residue 359 with C termini extending beyond residue 411. Little to no CD59 binding was observed for various C9-derived peptides that started C-terminal to residue 359 or that were truncated N-terminal to residue 411. Affinity-purified antibody against C9 residues 320-411 inhibited CD59 binding to C9 by > 50% and completely inhibited its binding to the isolated C9b domain. Little to no specific binding of CD59 was detected for peptides restricted to the putative hinge domain within C9b (residues 245-271). These results indicate that a CD59 binding site is located between residues 320 and 411 of the C9 polypeptide and suggest that the affinity of this site is principally determined by residues 359-411.

Engineering applications of strong ground motion simulation

NASA Astrophysics Data System (ADS)

Somerville, Paul

1993-02-01

The formulation, validation and application of a procedure for simulating strong ground motions for use in engineering practice are described. The procedure uses empirical source functions (derived from near-source strong motion recordings of small earthquakes) to provide a realistic representation of effects such as source radiation that are difficult to model at high frequencies due to their partly stochastic behavior. Wave propagation effects are modeled using simplified Green's functions that are designed to transfer empirical source functions from their recording sites to those required for use in simulations at a specific site. The procedure has been validated against strong motion recordings of both crustal and subduction earthquakes. For the validation process we choose earthquakes whose source models (including a spatially heterogeneous distribution of the slip of the fault) are independently known and which have abundant strong motion recordings. A quantitative measurement of the fit between the simulated and recorded motion in this validation process is used to estimate the modeling and random uncertainty associated with the simulation procedure. This modeling and random uncertainty is one part of the overall uncertainty in estimates of ground motions of future earthquakes at a specific site derived using the simulation procedure. The other contribution to uncertainty is that due to uncertainty in the source parameters of future earthquakes that affect the site, which is estimated from a suite of simulations generated by varying the source parameters over their ranges of uncertainty. In this paper, we describe the validation of the simulation procedure for crustal earthquakes against strong motion recordings of the 1989 Loma Prieta, California, earthquake, and for subduction earthquakes against the 1985 Michoacán, Mexico, and Valparaiso, Chile, earthquakes. We then show examples of the application of the simulation procedure to the estimatation of the design response spectra for crustal earthquakes at a power plant site in California and for subduction earthquakes in the Seattle-Portland region. We also demonstrate the use of simulation methods for modeling the attenuation of strong ground motion, and show evidence of the effect of critical reflections from the lower crust in causing the observed flattening of the attenuation of strong ground motion from the 1988 Saguenay, Quebec, and 1989 Loma Prieta earthquakes.

An Extreme Meteorological Events Analysis For Nuclear Power Plant (NPP) Siting Project at Bangka Island, Indonesia

NASA Astrophysics Data System (ADS)

Septiadi, Deni; S, Yarianto Sugeng B.; Sriyana; Anzhar, Kurnia; Suntoko, Hadi

2018-03-01

The potential sources of meteorological phenomena in Nuclear Power Plant (NPP) area of interest are identified and the extreme values of the possible resulting hazards associated which such phenomena are evaluated to derive the appropriate design bases for the NPP. The appropriate design bases shall be determined according to the Nuclear Energy Regulatory Agency (Bapeten) applicable regulations, which presently do not indicate quantitative criteria for purposes of determining the design bases for meteorological hazards. These meteorological investigations are also carried out to evaluate the regional and site specific meteorological parameters which affect the transport and dispersion of radioactive effluents on the environment of the region around the NPP site. The meteorological hazards are to be monitored and assessed periodically over the lifetime of the plant to ensure that consistency with the design assumptions is maintained throughout the full lifetime of the facility.

Initial Validation of NDVI time seriesfrom AVHRR, VEGETATION, and MODIS

NASA Technical Reports Server (NTRS)

Morisette, Jeffrey T.; Pinzon, Jorge E.; Brown, Molly E.; Tucker, Jim; Justice, Christopher O.

2004-01-01

The paper will address Theme 7: Multi-sensor opportunities for VEGETATION. We present analysis of a long-term vegetation record derived from three moderate resolution sensors: AVHRR, VEGETATION, and MODIS. While empirically based manipulation can ensure agreement between the three data sets, there is a need to validate the series. This paper uses atmospherically corrected ETM+ data available over the EOS Land Validation Core Sites as an independent data set with which to compare the time series. We use ETM+ data from 15 globally distributed sites, 7 of which contain repeat coverage in time. These high-resolution data are compared to the values of each sensor by spatially aggregating the ETM+ to each specific sensors' spatial coverage. The aggregated ETM+ value provides a point estimate for a specific site on a specific date. The standard deviation of that point estimate is used to construct a confidence interval for that point estimate. The values from each moderate resolution sensor are then evaluated with respect to that confident interval. Result show that AVHRR, VEGETATION, and MODIS data can be combined to assess temporal uncertainties and address data continuity issues and that the atmospherically corrected ETM+ data provide an independent source with which to compare that record. The final product is a consistent time series climate record that links historical observations to current and future measurements.

Methylene Blue Inhibits Caspases by Oxidation of the Catalytic Cysteine.

PubMed

Pakavathkumar, Prateep; Sharma, Gyanesh; Kaushal, Vikas; Foveau, Bénédicte; LeBlanc, Andrea C

2015-09-24

Methylene blue, currently in phase 3 clinical trials against Alzheimer Disease, disaggregates the Tau protein of neurofibrillary tangles by oxidizing specific cysteine residues. Here, we investigated if methylene blue can inhibit caspases via the oxidation of their active site cysteine. Methylene blue, and derivatives, azure A and azure B competitively inhibited recombinant Caspase-6 (Casp6), and inhibited Casp6 activity in transfected human colon carcinoma cells and in serum-deprived primary human neuron cultures. Methylene blue also inhibited recombinant Casp1 and Casp3. Furthermore, methylene blue inhibited Casp3 activity in an acute mouse model of liver toxicity. Mass spectrometry confirmed methylene blue and azure B oxidation of the catalytic Cys163 cysteine of Casp6. Together, these results show a novel inhibitory mechanism of caspases via sulfenation of the active site cysteine. These results indicate that methylene blue or its derivatives could (1) have an additional effect against Alzheimer Disease by inhibiting brain caspase activity, (2) be used as a drug to prevent caspase activation in other conditions, and (3) predispose chronically treated individuals to cancer via the inhibition of caspases.

Methylene Blue Inhibits Caspases by Oxidation of the Catalytic Cysteine

PubMed Central

Pakavathkumar, Prateep; Sharma, Gyanesh; Kaushal, Vikas; Foveau, Bénédicte; LeBlanc, Andrea C.

2015-01-01

Methylene blue, currently in phase 3 clinical trials against Alzheimer Disease, disaggregates the Tau protein of neurofibrillary tangles by oxidizing specific cysteine residues. Here, we investigated if methylene blue can inhibit caspases via the oxidation of their active site cysteine. Methylene blue, and derivatives, azure A and azure B competitively inhibited recombinant Caspase-6 (Casp6), and inhibited Casp6 activity in transfected human colon carcinoma cells and in serum-deprived primary human neuron cultures. Methylene blue also inhibited recombinant Casp1 and Casp3. Furthermore, methylene blue inhibited Casp3 activity in an acute mouse model of liver toxicity. Mass spectrometry confirmed methylene blue and azure B oxidation of the catalytic Cys163 cysteine of Casp6. Together, these results show a novel inhibitory mechanism of caspases via sulfenation of the active site cysteine. These results indicate that methylene blue or its derivatives could (1) have an additional effect against Alzheimer Disease by inhibiting brain caspase activity, (2) be used as a drug to prevent caspase activation in other conditions, and (3) predispose chronically treated individuals to cancer via the inhibition of caspases. PMID:26400108

Domain repertoires as a tool to derive protein recognition rules.

PubMed

Zucconi, A; Panni, S; Paoluzi, S; Castagnoli, L; Dente, L; Cesareni, G

2000-08-25

Several approaches, some of which are described in this issue, have been proposed to assemble a complete protein interaction map. These are often based on high throughput methods that explore the ability of each gene product to bind any other element of the proteome of the organism. Here we propose that a large number of interactions can be inferred by revealing the rules underlying recognition specificity of a small number (a few hundreds) of families of protein recognition modules. This can be achieved through the construction and characterization of domain repertoires. A domain repertoire is assembled in a combinatorial fashion by allowing each amino acid position in the binding site of a given protein recognition domain to vary to include all the residues allowed at that position in the domain family. The repertoire is then searched by phage display techniques with any target of interest and from the primary structure of the binding site of the selected domains one derives rules that are used to infer the formation of complexes between natural proteins in the cell.

Structural Model for the Flip-Flop Action in Thiamin Pyrophosphate-Dependent Human Pyruvate Dehydrogenase

NASA Technical Reports Server (NTRS)

Ciszak, Ewa; Dominiak, Paulina

2003-01-01

The derivative of vitamin B1 thiamin pyrophosphate (TPP) is a cofactor of enzymes performing catalysis in pathways of energy production, including (i) decarboxylation of alpha-keto acids followed by (ii) transketolation. These enzymes have shown a common mechanism of TPP activation by imposing an active V-conformation of this coenzyme that brings the N4 atom of the aminopyrimidine ring to the distance required for the intramolecular C-H N hydrogen-bonding with the C2- atom of the thiazolium ring. The reactive C2 atom of TPP is the nucleophile that attacks the carbonyl carbon of different substrates used by the TPP-dependent enzymes. The structure of the heterotetrameric human pyruvate dehydrogenase (Elp) recently determined in our laboratory (1) revealed the association pattern of the subunits and the specifics of two chemically equivalent cofactor binding sites. Dynamic nonequivalence of these two cofactor sites directs the flip-flop action of this enzyme, depending upon which two active sites effect each other (2). The crystal structure derived from the holo-form of Elp provided the basis for the model of the flip-flop action of Elp in which different steps of the catalytic reaction are performed in each of the two cofactor sites at any given moment, where these steps are governed by the concerted shuttle-like motion of the subunits. It is further proposed that balancing a hydrogen-bond network and related cofactor geometry determine the continuity of catalytic events.

Hydroxyhomocitrulline Is a Collagen-Specific Carbamylation Mark that Affects Cross-link Formation.

PubMed

Taga, Yuki; Tanaka, Keisuke; Hamada, Chieko; Kusubata, Masashi; Ogawa-Goto, Kiyoko; Hattori, Shunji

2017-10-19

Carbamylation is a non-enzymatic post-translational modification that physiologically occurs during aging and is a risk factor for various diseases. The most common product of carbamylation is homocitrulline (HCit), where a lysine (Lys) amino group has reacted with urea-derived cyanate. HCit has recently been detected in collagen; however, given that 15%-90% of total Lys in collagen is hydroxylated, it is unclear how hydroxylation affects collagen carbamylation. Here, we identified a collagen-specific carbamylation product, hydroxyhomocitrulline (HHCit), and showed that high levels of HHCit are correlated with age in rat tissue collagen and in vivo carbamylation in mice, as well as with the decline of kidney function in the serum of dialysis patients. Proteomic analysis of the carbamylated collagens identified α2(I) Lys 933 , a major cross-linking site, as a preferential HHCit site. Furthermore, our results suggest that hydroxylysine carbamylation affects the mechanical properties of connective tissue by competitively inhibiting collagen cross-link formation. Copyright © 2017 Elsevier Ltd. All rights reserved.

Energy efficiency trade-offs drive nucleotide usage in transcribed regions

PubMed Central

Chen, Wei-Hua; Lu, Guanting; Bork, Peer; Hu, Songnian; Lercher, Martin J.

2016-01-01

Efficient nutrient usage is a trait under universal selection. A substantial part of cellular resources is spent on making nucleotides. We thus expect preferential use of cheaper nucleotides especially in transcribed sequences, which are often amplified thousand-fold compared with genomic sequences. To test this hypothesis, we derive a mutation-selection-drift equilibrium model for nucleotide skews (strand-specific usage of ‘A' versus ‘T' and ‘G' versus ‘C'), which explains nucleotide skews across 1,550 prokaryotic genomes as a consequence of selection on efficient resource usage. Transcription-related selection generally favours the cheaper nucleotides ‘U' and ‘C' at synonymous sites. However, the information encoded in mRNA is further amplified through translation. Due to unexpected trade-offs in the codon table, cheaper nucleotides encode on average energetically more expensive amino acids. These trade-offs apply to both strand-specific nucleotide usage and GC content, causing a universal bias towards the more expensive nucleotides ‘A' and ‘G' at non-synonymous coding sites. PMID:27098217

Effects of Site-Specific Guanine C8-Modifications on an Intramolecular DNA G-Quadruplex

PubMed Central

Lech, Christopher Jacques; Cheow Lim, Joefina Kim; Wen Lim, Jocelyn Mei; Amrane, Samir; Heddi, Brahim; Phan, Anh Tuân

2011-01-01

Understanding the fundamentals of G-quadruplex formation is important both for targeting G-quadruplexes formed by natural sequences and for engineering new G-quadruplexes with desired properties. Using a combination of experimental and computational techniques, we have investigated the effects of site-specific substitution of a guanine with C8-modified guanine derivatives, including 8-bromo-guanine, 8-O-methyl-guanine, 8-amino-guanine, and 8-oxo-guanine, within a well-defined (3 + 1) human telomeric G-quadruplex platform. The effects of substitutions on the stability of the G-quadruplex were found to depend on the type and position of the modification among different guanines in the structure. An interesting modification-dependent NMR chemical-shift effect was observed across basepairing within a guanine tetrad. This effect was reproduced by ab initio quantum mechanical computations, which showed that the observed variation in imino proton chemical shift is largely influenced by changes in hydrogen-bond geometry within the guanine tetrad. PMID:22004753

Bispecific small molecule-antibody conjugate targeting prostate cancer.

PubMed

Kim, Chan Hyuk; Axup, Jun Y; Lawson, Brian R; Yun, Hwayoung; Tardif, Virginie; Choi, Sei Hyun; Zhou, Quan; Dubrovska, Anna; Biroc, Sandra L; Marsden, Robin; Pinstaff, Jason; Smider, Vaughn V; Schultz, Peter G

2013-10-29

Bispecific antibodies, which simultaneously target CD3 on T cells and tumor-associated antigens to recruit cytotoxic T cells to cancer cells, are a promising new approach to the treatment of hormone-refractory prostate cancer. Here we report a site-specific, semisynthetic method for the production of bispecific antibody-like therapeutics in which a derivative of the prostate-specific membrane antigen-binding small molecule DUPA was selectively conjugated to a mutant αCD3 Fab containing the unnatural amino acid, p-acetylphenylalanine, at a defined site. Homogeneous conjugates were generated in excellent yields and had good solubility. The efficacy of the conjugate was optimized by modifying the linker structure, relative binding orientation, and stoichiometry of the ligand. The optimized conjugate showed potent and selective in vitro activity (EC50 ~ 100 pM), good serum half-life, and potent in vivo activity in prophylactic and treatment xenograft mouse models. This semisynthetic approach is likely to be applicable to the generation of additional bispecific agents using drug-like ligands selective for other cell-surface receptors.

Water-sediment controversy in setting environmental standards for selenium

USGS Publications Warehouse

Hamilton, Steven J.; Lemly, A. Dennis

1999-01-01

A substantial amount of laboratory and field research on selenium effects to biota has been accomplished since the national water quality criterion was published for selenium in 1987. Many articles have documented adverse effects on biota at concentrations below the current chronic criterion of 5 μg/L. This commentary will present information to support a national water quality criterion for selenium of 2 μg/L, based on a wide array of support from federal, state, university, and international sources. Recently, two articles have argued for a sediment-based criterion and presented a model for deriving site-specific criteria. In one example, they calculate a criterion of 31 μg/L for a stream with a low sediment selenium toxicity threshold and low site-specific sediment total organic carbon content, which is substantially higher than the national criterion of 5 μg/L. Their basic premise for proposing a sediment-based method has been critically reviewed and problems in their approach are discussed.

Blood-Derived RNA- and microRNA-Hydrolyzing IgG Antibodies in Schizophrenia Patients.

PubMed

Ermakov, E A; Ivanova, S A; Buneva, V N; Nevinsky, G A

2018-05-01

Abzymes with various catalytic activities are the earliest statistically significant markers of existing and developing autoimmune diseases (AIDs). Currently, schizophrenia (SCZD) is not considered to be a typical AID. It was demonstrated recently that antibodies from SCZD patients efficiently hydrolyze DNA and myelin basic protein. Here, we showed for the first time that autoantibodies from 35 SCZD patients efficiently hydrolyze RNA (cCMP > poly(C) > poly(A) > yeast RNA) and analyzed site-specific hydrolysis of microRNAs involved in the regulation of several genes in SCZD (miR-137, miR-9-5p, miR-219-2-3p, and miR-219a-5p). All four microRNAs were cleaved by IgG preparations (n = 21) from SCZD patients in a site-specific manner. The RNase activity of the abzymes correlated with SCZD clinical parameters. The data obtained showed that SCZD patients might display signs of typical autoimmune processes associated with impaired functioning of microRNAs resulting from their hydrolysis by the abzymes.

ACTS Ka-band Propagation Research in a Spatially Diversified Network with Two USAT Ground Stations

NASA Technical Reports Server (NTRS)

Kalu, Alex; Acousta, R.; Durand, S.; Emrich, Carol; Ventre, G.; Wilson, W.

1999-01-01

Congestion in the radio spectrum below 18 GHz is stimulating greater interest in the Ka (20/30 GHz) frequency band. Transmission at these shorter wavelengths is greatly influenced by rain resulting in signal attenuation and decreased link availability. The size and projected cost of Ultra Small Aperture Terminals (USATS) make site diversity methodology attractive for rain fade compensation. Separation distances between terminals must be small to be of interest commercially. This study measures diversity gain at a separation distance <5 km and investigates utilization of S-band weather radar reflectivity in predicting diversity gain. Two USAT ground stations, separated by 2.43 km for spatial diversity, received a continuous Ka-band tone sent from NASA Glenn Research Center via the Advanced Communications Technology Satellite (ACTS) steerable antenna beam. Received signal power and rainfall were measured, and Weather Surveillance Radar-1998 Doppler (WSR-88D) data were obtained as a measure of precipitation along the USAT-to-ACTS slant path. Signal attenuation was compared for the two sites, and diversity gain was calculated for fades measured on eleven days. Correlation of WSR-88D S-band reflectivity with measured Ka-band attenuation consisted of locating radar volume elements along each slant path, converting reflectivity to Ka-band attenuation with rain rate calculation as an intermediate step. Specific attenuation for each associated path segment was summed, resulting in total attenuation along the slant path. Derived Ka-band attenuation did not correlate closely with empirical data (r = 0.239), but a measured signal fade could be matched with an increase in radar reflectivity in all fade events. Applying a low pass filter to radar reflectivity prior to deriving Ka-band attenuation improved the correlation between measured and derived signal attenuation (r = 0.733). Results indicate that site diversity at small separation distances is a viable means of rain fade compensation, and that existing models underestimate diversity gain for a subtropical climate such as Florida. Also, filtered WSR-88D reflectivity can be used for optimizing diversity terminal placement by comparing derived Ka- band attenuation between the diversity sites.

Molecular characterization of dissolved organic matter from subtropical wetlands: a comparative study through the analysis of optical properties, NMR and FTICR/MS

NASA Astrophysics Data System (ADS)

Hertkorn, Norbert; Harir, Mourad; Cawley, Kaelin M.; Schmitt-Kopplin, Philippe; Jaffé, Rudolf

2016-04-01

Wetlands provide quintessential ecosystem services such as maintenance of water quality, water supply and biodiversity, among others; however, wetlands are also among the most threatened ecosystems worldwide. Natural dissolved organic matter (DOM) is an abundant and critical component in wetland biogeochemistry. This study describes the first detailed, comparative, molecular characterization of DOM in subtropical, pulsed, wetlands, namely the Everglades (USA), the Pantanal (Brazil) and the Okavango Delta (Botswana), using optical properties, high-field nuclear magnetic resonance (NMR) and ultrahigh-resolution mass spectrometry (FT-ICRMS), and compares compositional features to variations in organic matter sources and flooding characteristics (i.e., differences in hydroperiod). While optical properties showed a high degree of variability within and between the three wetlands, analogies in DOM fluorescence properties were such that an established excitation emission matrix fluorescence parallel factor analysis (EEM-PARAFAC) model for the Everglades was perfectly applicable to the other two wetlands. Area-normalized 1H NMR spectra of selected samples revealed clear distinctions of samples while a pronounced congruence within the three pairs of wetland DOM readily suggested the presence of an individual wetland-specific molecular signature. Within sample pairs (long- vs. short-hydroperiod sites), internal differences mainly referred to intensity variations (denoting variable abundance) rather than to alterations of NMR resonances positioning (denoting diversity of molecules). The relative disparity was largest between the Everglades long- and short-hydroperiod samples, whereas Pantanal and Okavango samples were more alike among themselves. Otherwise, molecular divergence was most obvious in the case of unsaturated protons (δH > 5 ppm). 2-D NMR spectroscopy for a particular sample revealed a large richness of aliphatic and unsaturated substructures, likely derived from microbial sources such as periphyton in the Everglades. In contrast, the chemical diversity of aromatic wetland DOM likely originates from a combination of higher plant sources, progressive microbial and photochemical oxidation, and contributions from combustion-derived products (e.g., black carbon). FT-ICRMS spectra of both Okavango and Pantanal showed near 57 ± 2 % CHO, 8 ± 2 % CHOS, 33 ± 2 % CHNO and < 1 % CHNOS molecules, whereas those of Everglades samples were markedly enriched in CHOS and CHNOS at the expense of CHO and CHNO compounds. In particular, the Everglades short-hydroperiod site showed a large set of aromatic and oxygen-deficient "black sulfur" compounds whereas the long-hydroperiod site contained oxygenated sulfur attached to fused-ring polyphenols. The elevated abundance of CHOS compounds for the Everglades samples likely results from higher inputs of agriculture-derived and sea-spray-derived sulfate. Although wetland DOM samples were found to share many molecular features, each sample was unique in its composition, which reflected specific environmental drivers and/or specific biogeochemical processes.

Natural radionuclides and stable elements in weaver ants (Oecophylla smaragdina) from tropical northern Australia.

PubMed

Medley, Peter; Doering, Che; Evans, Fiona; Bollhöfer, Andreas

2017-11-01

Natural radionuclides and stable elements were measured in weaver ants, leaves and soils collected from three sites in tropical northern Australia. Radionuclide concentration ratios for ants relative to soil were derived from the measurements and used to refine the current environmental radiological assessment for remediation of Ranger uranium mine. Use of site-specific concentration ratios for weaver ants gave a more conservative estimate of environmental exposure to the arthropod wildlife group than use of default concentration ratios in the ERICA Tool. This was primarily because the 226 Ra concentration ratio for weaver ants was more than 7 times greater than for generic arthropods. Crown Copyright © 2017. Published by Elsevier Ltd. All rights reserved.

Settlement patterns, GIS, remote sensing, and the late prehistory of the Black Prairie in east central Mississippi

NASA Technical Reports Server (NTRS)

Johnson, Jay K.

1991-01-01

Data recovered as the result of a recent field project designed to test a model of the distribution of protohistoric settlement in an unusual physiographic zone in eastern Mississippi are examined using GIS based techniques to manipulate soil and stream distance information. Significant patterning is derived. The generally thin soils and uniform substratum of the Black Prairie in combination with a distinctive settlement pattern offer a promising opportunity for the search for site specific characteristics within airborne imagery. Landsat TM data provide information on modern ground cover which is used as a mask to select areas in which a multivariate search for archaeological site signatures within a TIMS image is most likely to prove fruitful.

Preferential Redistribution of Fine-Grained Particles in the Panama Basin and Potential Errors in 230Th-Derived Focusing Factors

NASA Astrophysics Data System (ADS)

Marcantonio, F.; Lyle, M. W.; Ibrahim, R.

2013-12-01

The 230Th constant-flux proxy technique, commonly used in paleoceanography to estimate sediment fluxes, is thought to differentiate lateral from vertical fluxes of sediment at sites that have undergone sediment redistribution. However, redistribution processes (focusing or winnowing) are expected to fractionate fine particles from those that are coarse. Since fine particles with greater surface area are known to contain greater concentrations of 230Th, one might expect that sediment redistribution would bias 230Th-derived sediment mass accumulation rates (MARs). We investigate this possibility in two regions of the Panama Basin where significant sediment focusing has been hypothesized to occur. We examine multicore sediments from paired sites at two locations, one close to the equator at the southern limit of the Panama Basin (Carnegie Ridge) where upwelling and primary productivity are high, and one at 6°N at the northern boundary of the Panama Basin (Cocos Ridge), where primary productivity is lower. The multicores, which are constrained by radiocarbon ages that span the latest Holocene at each paired site, represent regions that have undergone potential winnowing and focusing (thin vs thick sediment drapes identified using seismic reflection) at each Panama Basin location. Since the distance separating the paired sites at each location is no more than about 50 km, one would expect the 230Th-derived MARs to be similar, i.e., the rain rate should not be significantly different at each of the paired sites. The radiocarbon-derived sand fraction (>63-μm) MARs, which likely represent the vertical rain of particles not transported by bottom currents, are identical at each of the paired sites, with fluxes at the Carnegie Ridge about 3.5 times greater than those at the Cocos Ridge over the past several thousand years. Over the same time period, the 230Th-normalized MARs are relatively similar at both the Carnegie and Cocos sites, but are different by about 60% at each of the paired sites, with the higher MARs always at the potentially winnowed sites. The 'thick' Carnegie Ridge site has a 230Th-derived focusing factor of about 6. However, we believe that, given our observations of the radiocarbon-derived fluxes of sand, the high 230Th-derived focusing factor is likely an overestimate of the degree of focusing, and the average 230Th-derived MAR is likely an underestimate of the true MAR. Similarly, the biasing for the 'thin' Cocos site that has a 230Th-derived focusing factor of 0.2 (i.e., potential winnowing) is expected to be the opposite, namely, 230Th-derived fluxes are likely being overestimated there. The quantitative extent to which 230Th-derived focusing factors have been over- and under-estimated in the Panama Basin will be discussed. We hypothesize that size fractionation, as well as 230Th focusing errors, occur most frequently at lower current velocities where the fine and coarse fraction of the sediment components are more effectively sorted from each other.

Characterization of the rat RALDH1 promoter. A functional CCAAT and octamer motif are critical for basal promoter activity.

PubMed

Guimond, Julie; Devost, Dominic; Brodeur, Helene; Mader, Sylvie; Bhat, Pangala V

2002-12-12

Retinal dehydrogenase type 1 (RALDH1) catalyzes the oxidation of retinal to retinoic acid (RA), a metabolite of vitamin A important for embryogenesis and tissue differentiation. Rat RALDH1 is expressed to high levels in developing kidney, and in stomach, intestine epithelia. To understand the mechanisms of the transcriptional regulation of rat RALDH1, we cloned a 1360-base pair (bp) 5'-flanking region of RALDH1 gene. Using luciferase reporter constructs transfected into HEK 293 and LLCPK (kidney-derived) cells, basal promoter activity was associated with sequences between -80 and +43. In this minimal promoter region, TATA and CCAAT cis-acting elements as well as SP1, AP1 and octamer (Oct)-binding sites were present. The CCAAT box and Oct-binding site, located between positions -72 and -68 and -56 and -49, respectively, were shown by deletion analysis and site-directed mutation to be critical for promoter activity. Nuclear extracts from kidney cells contain proteins specifically binding the Oct and CCAAT sequences, resulting in the formation of six complexes, while different patterns of complexes were observed with non-kidney cell extracts. Gel shift assays using either single or double mutations of the Oct and CCAAT sequences as well as super shift assays demonstrated single and double occupancy of these two sites by Oct-1 and CBF-A. In addition, unidentified proteins also bound the Oct motif specifically in the absence of CBF-A binding. These results demonstrate specific involvement of Oct and CCAAT-binding proteins in the regulation of RALDH1 gene.

Prediction of earthquake ground motion at rock sites in Japan: evaluation of empirical and stochastic approaches for the PEGASOS Refinement Project

NASA Astrophysics Data System (ADS)

Edwards, Benjamin; Fäh, Donat

2017-11-01

Strong ground-motion databases used to develop ground-motion prediction equations (GMPEs) and calibrate stochastic simulation models generally include relatively few recordings on what can be considered as engineering rock or hard rock. Ground-motion predictions for such sites are therefore susceptible to uncertainty and bias, which can then propagate into site-specific hazard and risk estimates. In order to explore this issue we present a study investigating the prediction of ground motion at rock sites in Japan, where a wide range of recording-site types (from soil to very hard rock) are available for analysis. We employ two approaches: empirical GMPEs and stochastic simulations. The study is undertaken in the context of the PEGASOS Refinement Project (PRP), a Senior Seismic Hazard Analysis Committee (SSHAC) Level 4 probabilistic seismic hazard analysis of Swiss nuclear power plants, commissioned by swissnuclear and running from 2008 to 2013. In order to reduce the impact of site-to-site variability and expand the available data set for rock and hard-rock sites we adjusted Japanese ground-motion data (recorded at sites with 110 m s-1 < Vs30 < 2100 m s-1) to a common hard-rock reference. This was done through deconvolution of: (i) empirically derived amplification functions and (ii) the theoretical 1-D SH amplification between the bedrock and surface. Initial comparison of a Japanese GMPE's predictions with data recorded at rock and hard-rock sites showed systematic overestimation of ground motion. A further investigation of five global GMPEs' prediction residuals as a function of quarter-wavelength velocity showed that they all presented systematic misfit trends, leading to overestimation of median ground motions at rock and hard-rock sites in Japan. In an alternative approach, a stochastic simulation method was tested, allowing the direct incorporation of site-specific Fourier amplification information in forward simulations. We use an adjusted version of the model developed for Switzerland during the PRP. The median simulation prediction at true rock and hard-rock sites (Vs30 > 800 m s-1) was found to be comparable (within expected levels of epistemic uncertainty) to predictions using an empirical GMPE, with reduced residual misfit. As expected, due to including site-specific information in the simulations, the reduction in misfit could be isolated to a reduction in the site-related within-event uncertainty. The results of this study support the use of finite or pseudo-finite fault stochastic simulation methods in estimating strong ground motions in regions of weak and moderate seismicity, such as central and northern Europe. Furthermore, it indicates that weak-motion data has the potential to allow estimation of between- and within-site variability in ground motion, which is a critical issue in site-specific seismic hazard analysis, particularly for safety critical structures.

Chemoselective synthesis of functional homocysteine residues in polypeptides and peptides.

PubMed

Gharakhanian, Eric G; Deming, Timothy J

2016-04-18

A methodology was developed for efficient, chemoselective transformation of methionine residues into stable, functional homocysteine derivatives. Methionine residues can undergo highly chemoselective alkylation reactions at low pH to yield stable sulfonium ions, which could then be selectively demethylated to give stable alkyl homocysteine residues. This mild, two-step process is chemoselective, efficient, tolerates many functional groups, and provides a means for creation of new functional biopolymers, site-specific peptide tagging, and synthesis of biomimetic and structural analogs of peptides.

The Use of Aptamers in Breast Cancer Anti-Metastasis Therapy

DTIC Science & Technology

2001-09-01

with serum-free medium (IRL-41) supplemented with 0.4 mM ascorbic acid and antibiotics for 72 h (2,3). Recombinant P4(108_343)-HA was purified from...mCLCA1 and hCLCA1 (9, calcium -activated chloride channel; cystic fibrosis 11). Although the protein processing and function ap- pear to be conserved among...AAC) to glutamine ( CAA ) mutation that abolishes a derived primers followed by internal gene-specific primers consensus glycosylation site (NxS/T-QxS/T

KCNE1 induces fenestration in the Kv7.1/KCNE1 channel complex that allows for highly specific pharmacological targeting

NASA Astrophysics Data System (ADS)

Wrobel, Eva; Rothenberg, Ina; Krisp, Christoph; Hundt, Franziska; Fraenzel, Benjamin; Eckey, Karina; Linders, Joannes T. M.; Gallacher, David J.; Towart, Rob; Pott, Lutz; Pusch, Michael; Yang, Tao; Roden, Dan M.; Kurata, Harley T.; Schulze-Bahr, Eric; Strutz-Seebohm, Nathalie; Wolters, Dirk; Seebohm, Guiscard

2016-10-01

Most small-molecule inhibitors of voltage-gated ion channels display poor subtype specificity because they bind to highly conserved residues located in the channel's central cavity. Using a combined approach of scanning mutagenesis, electrophysiology, chemical ligand modification, chemical cross-linking, MS/MS-analyses and molecular modelling, we provide evidence for the binding site for adamantane derivatives and their putative access pathway in Kv7.1/KCNE1 channels. The adamantane compounds, exemplified by JNJ303, are highly potent gating modifiers that bind to fenestrations that become available when KCNE1 accessory subunits are bound to Kv7.1 channels. This mode of regulation by auxiliary subunits may facilitate the future development of potent and highly subtype-specific Kv channel inhibitors.

Physical constraints determine the logic of bacterial promoter architectures

PubMed Central

Ezer, Daphne; Zabet, Nicolae Radu; Adryan, Boris

2014-01-01

Site-specific transcription factors (TFs) bind to their target sites on the DNA, where they regulate the rate at which genes are transcribed. Bacterial TFs undergo facilitated diffusion (a combination of 3D diffusion around and 1D random walk on the DNA) when searching for their target sites. Using computer simulations of this search process, we show that the organization of the binding sites, in conjunction with TF copy number and binding site affinity, plays an important role in determining not only the steady state of promoter occupancy, but also the order at which TFs bind. These effects can be captured by facilitated diffusion-based models, but not by standard thermodynamics. We show that the spacing of binding sites encodes complex logic, which can be derived from combinations of three basic building blocks: switches, barriers and clusters, whose response alone and in higher orders of organization we characterize in detail. Effective promoter organizations are commonly found in the E. coli genome and are highly conserved between strains. This will allow studies of gene regulation at a previously unprecedented level of detail, where our framework can create testable hypothesis of promoter logic. PMID:24476912

Evolution of I-SceI Homing Endonucleases with Increased DNA Recognition Site Specificity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Joshi, Rakesh; Ho, Kwok Ki; Tenney, Kristen

2013-09-18

Elucidating how homing endonucleases undergo changes in recognition site specificity will facilitate efforts to engineer proteins for gene therapy applications. I-SceI is a monomeric homing endonuclease that recognizes and cleaves within an 18-bp target. It tolerates limited degeneracy in its target sequence, including substitution of a C:G{sub +4} base pair for the wild-type A:T{sub +4} base pair. Libraries encoding randomized amino acids at I-SceI residue positions that contact or are proximal to A:T{sub +4} were used in conjunction with a bacterial one-hybrid system to select I-SceI derivatives that bind to recognition sites containing either the A:T{sub +4} or the C:G{submore » +4} base pairs. As expected, isolates encoding wild-type residues at the randomized positions were selected using either target sequence. All I-SceI proteins isolated using the C:G{sub +4} recognition site included small side-chain substitutions at G100 and either contained (K86R/G100T, K86R/G100S and K86R/G100C) or lacked (G100A, G100T) a K86R substitution. Interestingly, the binding affinities of the selected variants for the wild-type A:T{sub +4} target are 4- to 11-fold lower than that of wild-type I-SceI, whereas those for the C:G{sub +4} target are similar. The increased specificity of the mutant proteins is also evident in binding experiments in vivo. These differences in binding affinities account for the observed -36-fold difference in target preference between the K86R/G100T and wild-type proteins in DNA cleavage assays. An X-ray crystal structure of the K86R/G100T mutant protein bound to a DNA duplex containing the C:G{sub +4} substitution suggests how sequence specificity of a homing enzyme can increase. This biochemical and structural analysis defines one pathway by which site specificity is augmented for a homing endonuclease.« less

Structure of a highly acidic β-lactamase from the moderate halophile Chromohalobacter sp. 560 and the discovery of a Cs{sup +}-selective binding site

DOE Office of Scientific and Technical Information (OSTI.GOV)

Arai, Shigeki; Yonezawa, Yasushi; Okazaki, Nobuo

2015-03-01

The tertiary structure of a β-lactamase derived from the halobacterium Chromohalobacter sp. 560 (HaBLA) was determined by X-ray crystallography. Three unique Sr{sup 2+}-binding sites and one Cs{sup +}-binding site were discovered in the HaBLA molecule. Environmentally friendly absorbents are needed for Sr{sup 2+} and Cs{sup +}, as the removal of the radioactive Sr{sup 2+} and Cs{sup +} that has leaked from the Fukushima Nuclear Power Plant is one of the most important problems in Japan. Halophilic proteins are known to have many acidic residues on their surface that can provide specific binding sites for metal ions such as Cs{sup +}more » or Sr{sup 2+}. The crystal structure of a halophilic β-lactamase from Chromohalobacter sp. 560 (HaBLA) was determined to resolutions of between 1.8 and 2.9 Å in space group P3{sub 1} using X-ray crystallography. Moreover, the locations of bound Sr{sup 2+} and Cs{sup +} ions were identified by anomalous X-ray diffraction. The location of one Cs{sup +}-specific binding site was identified in HaBLA even in the presence of a ninefold molar excess of Na{sup +} (90 mM Na{sup +}/10 mM Cs{sup +}). From an activity assay using isothermal titration calorimetry, the bound Sr{sup 2+} and Cs{sup +} ions do not significantly affect the enzymatic function of HaBLA. The observation of a selective and high-affinity Cs{sup +}-binding site provides important information that is useful for the design of artificial Cs{sup +}-binding sites that may be useful in the bioremediation of radioactive isotopes.« less

Altered Substrate Specificity of Drug-Resistant Human Immunodeficiency Virus Type 1 Protease

PubMed Central

Dauber, Deborah S.; Ziermann, Rainer; Parkin, Neil; Maly, Dustin J.; Mahrus, Sami; Harris, Jennifer L.; Ellman, Jon A.; Petropoulos, Christos; Craik, Charles S.

2002-01-01

Resistance to human immunodeficiency virus type 1 protease (HIV PR) inhibitors results primarily from the selection of multiple mutations in the protease region. Because many of these mutations are selected for the ability to decrease inhibitor binding in the active site, they also affect substrate binding and potentially substrate specificity. This work investigates the substrate specificity of a panel of clinically derived protease inhibitor-resistant HIV PR variants. To compare protease specificity, we have used positional-scanning, synthetic combinatorial peptide libraries as well as a select number of individual substrates. The subsite preferences of wild-type HIV PR determined by using the substrate libraries are consistent with prior reports, validating the use of these libraries to compare specificity among a panel of HIV PR variants. Five out of seven protease variants demonstrated subtle differences in specificity that may have significant impacts on their abilities to function in viral maturation. Of these, four variants demonstrated up to fourfold changes in the preference for valine relative to alanine at position P2 when tested on individual peptide substrates. This change correlated with a common mutation in the viral NC/p1 cleavage site. These mutations may represent a mechanism by which severely compromised, drug-resistant viral strains can increase fitness levels. Understanding the altered substrate specificity of drug-resistant HIV PR should be valuable in the design of future generations of protease inhibitors as well as in elucidating the molecular basis of regulation of proteolysis in HIV. PMID:11773410

Characterization, solubilization and partial purification of serotonin 5-HT1C receptors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yagaloff, K.A.

1986-01-01

/sup 125/I-Lysergic acid diethylamide (/sup 125/I-LSD) binds with high affinity to a unique serotonergic site on rat choroid plexus. These sites were localized to choroid plexus epithelial cells using a novel high resolution autoradiographic technique. In membrane preparations, the serotonergic site density was 3100 fmol/mg protein, which is 10 fold higher than the density of any other serotonergic site in brain homogenates. The pharmacology of this site, termed the 5-HT1c site, does not match that of 5-Ht1a, 5-HT1b or 5HT2 serotonergic sites. 5-Ht1c sites were solubilized from pig choroid plexus using the zwitterionic detergent, CHAPS. High affinity labelling of themore » solubilized site was obtained using the serotonergic radioligand, N1-methyl-2-(/sup 125/I)lysergic acid diethylamide (/sup 125/I-MIL). Choroid plexus tumors obtained from transgenic mice were examined for the presence of serotonin 5-HT1c receptors. /sup 125/I-LSD binding to choroid plexus tumors displays a pharmacological profile that matches the properties of 5-HT1c receptors in normal choroid plexus. The tumor exhibits the highest site density of serotonin receptors (6600 fmol/mg protein) found in any tissue. /sup 125/I-LSD autoradiography of brain sections from transgenic mice shows high levels of specific labelling over the tumor. The affinities of various indolealkyl, phenlakyl and beta-carboline derivatives for the serotonin 5-HT1c receptor were measured in pig choroid plexus using /sup 125/I-MIL. Serotonin precursors and metabolites were all very weak inhibitors of specific /sup 125/I-MIL binding. Structure-affinity relationships were determined for a number of indolealkylamine analogues. Only serotonin is present in cerebrospinal fluid at concentrations near its 5-HT1c inhibition constant, suggesting that serotonin is the natural 5-HT1c agonist.« less

Coronary CT Angiography-derived Fractional Flow Reserve: Machine Learning Algorithm versus Computational Fluid Dynamics Modeling.

PubMed

Tesche, Christian; De Cecco, Carlo N; Baumann, Stefan; Renker, Matthias; McLaurin, Tindal W; Duguay, Taylor M; Bayer, Richard R; Steinberg, Daniel H; Grant, Katharine L; Canstein, Christian; Schwemmer, Chris; Schoebinger, Max; Itu, Lucian M; Rapaka, Saikiran; Sharma, Puneet; Schoepf, U Joseph

2018-04-10

Purpose To compare two technical approaches for determination of coronary computed tomography (CT) angiography-derived fractional flow reserve (FFR)-FFR derived from coronary CT angiography based on computational fluid dynamics (hereafter, FFR CFD ) and FFR derived from coronary CT angiography based on machine learning algorithm (hereafter, FFR ML )-against coronary CT angiography and quantitative coronary angiography (QCA). Materials and Methods A total of 85 patients (mean age, 62 years ± 11 [standard deviation]; 62% men) who had undergone coronary CT angiography followed by invasive FFR were included in this single-center retrospective study. FFR values were derived on-site from coronary CT angiography data sets by using both FFR CFD and FFR ML . The performance of both techniques for detecting lesion-specific ischemia was compared against visual stenosis grading at coronary CT angiography, QCA, and invasive FFR as the reference standard. Results On a per-lesion and per-patient level, FFR ML showed a sensitivity of 79% and 90% and a specificity of 94% and 95%, respectively, for detecting lesion-specific ischemia. Meanwhile, FFR CFD resulted in a sensitivity of 79% and 89% and a specificity of 93% and 93%, respectively, on a per-lesion and per-patient basis (P = .86 and P = .92). On a per-lesion level, the area under the receiver operating characteristics curve (AUC) of 0.89 for FFR ML and 0.89 for FFR CFD showed significantly higher discriminatory power for detecting lesion-specific ischemia compared with that of coronary CT angiography (AUC, 0.61) and QCA (AUC, 0.69) (all P < .0001). Also, on a per-patient level, FFR ML (AUC, 0.91) and FFR CFD (AUC, 0.91) performed significantly better than did coronary CT angiography (AUC, 0.65) and QCA (AUC, 0.68) (all P < .0001). Processing time for FFR ML was significantly shorter compared with that of FFR CFD (40.5 minutes ± 6.3 vs 43.4 minutes ± 7.1; P = .042). Conclusion The FFR ML algorithm performs equally in detecting lesion-specific ischemia when compared with the FFR CFD approach. Both methods outperform accuracy of coronary CT angiography and QCA in the detection of flow-limiting stenosis. © RSNA, 2018.

How does precipitation become runoff? Comparison of hydrologic thresholds across hillslope and catchment scales

NASA Astrophysics Data System (ADS)

Ross, C.; Ali, G.; Oswald, C. J.; McMillan, H. K.; Walter, K.

2017-12-01

A hydrologic threshold is a critical point in time when runoff behavior rapidly changes, often in response to the activation of specific storage-driven or intensity-driven processes. Hydrologic thresholds can be viewed as characteristic signatures of hydrosystems, which makes them useful for site comparison as long as their presence (or lack thereof) can be evaluated in a standard manner across a range of environments. While several previous studies have successfully identified thresholds at a variety of individual sites, only a limited number have compared dynamics prevailing at the hillslope versus catchment scale, or distinguished the role of storage versus intensity thresholds. The objective of this study was therefore to examine precipitation input thresholds as well as "precipitation minus evapotranspiration" thresholds in environments with contrasted climatic and geographic characteristics. Historical climate and hydrometric datasets were consolidated for one hillslope site located at the Panola Mountain Research Watershed (Southeastern USA) and catchments located in the HJ Andrew's Experimental Forest (Northwestern USA), the Catfish Creek Watershed (Canadian prairies), the Experimental Lakes Area (Canadian boreal ecozone), the Tarrawarra catchment (Australia) and the Mahurangi catchment (New Zealand). Individual precipitation-runoff events were delineated using the newly introduced software HydRun to derive event-specific hydrograph parameters as well surrogate measures of antecedent moisture conditions and evapotranspiration in an automated and consistent manner. Various hydrograph parameters were then plotted against those surrogate measures to detect and evaluate site-specific threshold dynamics. Preliminary results show that a range of threshold shapes (e.g., "hockey stick", heaviside and dirac) were observed across sites. The influence of antecedent precipitation on threshold magnitude and shape also appeared stronger at sites with lower topographic relief and drier climate. Future analyses will focus on the interaction between storage and intensity thresholds in order to evaluate the importance of considering both for comparative hydrological studies.

Enzymatic Specific Production and Chemical Functionalization of Phenylpropanone Platform Monomers from Lignin

PubMed Central

Hasegawa, Ryoichi; Kurosawa, Kanako; Maeda, Allyn H.; Koizumi, Toshio; Nishimura, Hiroshi; Okada, Hitomi; Qu, Chen; Saito, Kaori; Watanabe, Takashi; Hatada, Yuji

2016-01-01

Abstract Enzymatic catalysis is an ecofriendly strategy for the production of high‐value low‐molecular‐weight aromatic compounds from lignin. Although well‐definable aromatic monomers have been obtained from synthetic lignin‐model dimers, enzymatic‐selective synthesis of platform monomers from natural lignin has not been accomplished. In this study, we successfully achieved highly specific synthesis of aromatic monomers with a phenylpropane structure directly from natural lignin using a cascade reaction of β‐O‐4‐cleaving bacterial enzymes in one pot. Guaiacylhydroxylpropanone (GHP) and the GHP/syringylhydroxylpropanone (SHP) mixture are exclusive monomers from lignin isolated from softwood (Cryptomeria japonica) and hardwood (Eucalyptus globulus). The intermediate products in the enzymatic reactions show the capacity to accommodate highly heterologous substrates at the substrate‐binding sites of the enzymes. To demonstrate the applicability of GHP as a platform chemical for bio‐based industries, we chemically generate value‐added GHP derivatives for bio‐based polymers. Together with these chemical conversions for the valorization of lignin‐derived phenylpropanone monomers, the specific and enzymatic production of the monomers directly from natural lignin is expected to provide a new stream in “white biotechnology” for sustainable biorefineries. PMID:27878983

Production of rosamicin derivatives in Micromonospora rosaria by introduction of D-mycinose biosynthetic gene with PhiC31-derived integration vector pSET152.

PubMed

Anzai, Yojiro; Iizaka, Yohei; Li, Wei; Idemoto, Naoki; Tsukada, Shu-ichi; Koike, Kazuo; Kinoshita, Kenji; Kato, Fumio

2009-08-01

Some of the polyketide-derived bioactive compounds contain sugars attached to the aglycone core, and these sugars often impart specific biological activity to the molecule or enhance this activity. Mycinamicin II, a 16-member macrolide antibiotic produced by Micromonospora griseorubida A11725, contains a branched lactone and two different deoxyhexose sugars, D-desosamine and D-mycinose, at the C-5 and C-21 positions, respectively. The D-mycinose biosynthesis genes, mycCI, mycCII, mycD, mycE, mycF, mydH, and mydI, present in the M. griseorubida A11725 chromosome were introduced into pSET152 under the regulation of the promoter of the apramycin-resistance gene aac(3)IV. The resulting plasmid pSETmycinose was introduced into Micromonospora rosaria IFO13697 cells, which produce the 16-membered macrolide antibiotic rosamicin containing a branched lactone and D-desosamine at the C-5 position. Although the M. rosaria TPMA0001 transconjugant exhibited low rosamicin productivity, two new compounds, IZI and IZII, were detected in the ethylacetate extract from the culture broth. IZI was identified as a mycinosyl rosamicin derivative, 23-O-mycinosyl-20-deoxo-20-dihydro-12,13-deepoxyrosamicin (MW 741), which has previously been synthesized by a bioconversion technique. This is the first report on production of mycinosyl rosamicin-derivatives by a engineered biosynthesis approach. The integration site PhiC31attB was identified on M. rosaria IFO13697 chromosome, and the site lay within an ORF coding a pirin homolog protein. The pSETmycinose could be useful for stimulating the production of "unnatural" natural mycinosyl compounds by various actinomycete strains using the bacteriophage PhiC31 att/int system.

Nitrogen-doped two-dimensional porous carbon sheets derived from clover biomass for high performance supercapacitors

NASA Astrophysics Data System (ADS)

Wang, Cunjing; Wu, Dapeng; Wang, Hongju; Gao, Zhiyong; Xu, Fang; Jiang, Kai

2017-09-01

Highly porous carbon sheets were prepared from fresh clover stems under air atmosphere via a facile potassium chloride salt-sealing technique, which not only avoids using the high cost inert gas protection but also spontaneously introduce multi-level porosity into the carbon structure taking advantage of the trace of oxygen in the molten salt system. The as-obtained porous carbon sheets possess high specific surface area of 2244 m2 g-1 and interconnected hierarchical pore structures from micro-to macro-scale, which provide abundant storage active sites and fast ion diffusion channels. In addition, the spontaneously formed N (2.55 at%) and O (6.94 at%) doping sites not only improve the electron conductivity of the electrode but also enhance the specific capacitance by introducing pseudocapacitance. When employed as supercapacitor electrodes, a high specific capacitance of 436 F g-1 at 1 A g-1 and an excellent rate capacity with capacitance remaining 290 F g-1 at 50 A g-1 are demonstrated. Furthermore, the assembled symmetric supercapacitor delivers a high specific capacitance of 420 F g-1 at 0.5 A g-1, excellent energy density of 58.4 Wh kg-1 and good cycling stability which retains 99.4% of the initial capacitance at 5 A g-1 after 30,000 cycles.

Cytochemical demonstration of oxidative damage in Alzheimer disease by immunochemical enhancement of the carbonyl reaction with 2,4-dinitrophenylhydrazine.

PubMed

Smith, M A; Sayre, L M; Anderson, V E; Harris, P L; Beal, M F; Kowall, N; Perry, G

1998-06-01

Formation of carbonyls derived from lipids, proteins, carbohydrates, and nucleic acids is common during oxidative stress. For example, metal-catalyzed, "site-specific" oxidation of several amino acid side-chains produces aldehydes or ketones, and peroxidation of lipids generates reactive aldehydes such as malondialdehyde and hydroxynonenal. Here, using in situ 2,4-dinitrophenylhydrazine labeling linked to an antibody system, we describe a highly sensitive and specific cytochemical technique to specifically localize biomacromolecule-bound carbonyl reactivity. When this technique was applied to tissues from cases of Alzheimer disease, in which oxidative events including lipoperoxidative, glycoxidative, and other oxidative protein modifications have been reported, we detected free carbonyls not only in the disease-related intraneuronal lesions but also in other neurons. In marked contrast, free carbonyls were not found in neurons or glia in age-matched control cases. Importantly, this assay was highly specific for detecting disease-related oxidative damage because the site of oxidative damage can be assessed in the midst of concurrent age-related increases in free carbonyls in vascular basement membrane that would contaminate biochemical samples subjected to bulk analysis. These findings demonstrate that oxidative imbalance and stress are key elements in the pathogenesis of Alzheimer disease.

Development of [3H]2-Carboxy-4,6-dichloro-1H-indole-3-propionic Acid ([3H]PSB-12150): A Useful Tool for Studying GPR17

PubMed Central

2014-01-01

The recently described synthetic GPR17 agonist 2-carboxy-4,6-dichloro-1H-indole-3-propionic acid (1) was prepared in tritium-labeled form by catalytic hydrogenation of the corresponding propenoic acid derivative 8 with tritium gas. The radioligand [3H]PSB-12150 (9) was obtained with a specific activity of 17 Ci/mmol (629 GBq/mmol). It showed specific and saturable binding to a single binding site in membrane preparations from Chinese hamster ovary cells recombinantly expressing the human GPR17. A competition assay procedure was established, which allows the determination of ligand binding affinities. PMID:24900835

Development of [(3)H]2-Carboxy-4,6-dichloro-1H-indole-3-propionic Acid ([(3)H]PSB-12150): A Useful Tool for Studying GPR17.

PubMed

Köse, Meryem; Ritter, Kirsten; Thiemke, Katharina; Gillard, Michel; Kostenis, Evi; Müller, Christa E

2014-04-10

The recently described synthetic GPR17 agonist 2-carboxy-4,6-dichloro-1H-indole-3-propionic acid (1) was prepared in tritium-labeled form by catalytic hydrogenation of the corresponding propenoic acid derivative 8 with tritium gas. The radioligand [(3)H]PSB-12150 (9) was obtained with a specific activity of 17 Ci/mmol (629 GBq/mmol). It showed specific and saturable binding to a single binding site in membrane preparations from Chinese hamster ovary cells recombinantly expressing the human GPR17. A competition assay procedure was established, which allows the determination of ligand binding affinities.

Identification and characterization of (/sup 3/H)-rauwolscine binding to alpha2-adrenoceptors in the canine saphenous vein

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gout, B.

1988-01-01

The biochemical exploration of the alpha2-adrenergic receptors was investigated in the canine saphenous vein using the highly selective alpha2-adrenergic antagonist rauwolscine as a tritiated ligand. Following an enzymatic digestive pretreatment, the authors isolated a purified smooth muscle cell membranes fraction from saphenous veins in quantity sufficient to permit them to study the venous alpha2-adrenoreceptor content. The binding of tritiated rauwolscine was rapid, specific, saturable and reversible. The presence of high affinity sites with a density of binding Bmax of 125.2 /+ -/ 43.1 fmol/mg protein was demonstrated on a unique class of non interacting sites. The kinetically derived Kd wasmore » 1.28 nM, in good agreement with the value obtained from saturation isotherms. The pharmacological profile of these sites was assessed by the comparison of the potency of alpha-adrenergic agonists and antagonists to inhibit 1 nM (/sup 3/H)-rauwolscine. Their efficacy was respectively: rauwolscine > phentolamine > RX 781094 > clonidine >> prazosin > (-)-phenylephrine > (-)-noradrenaline. The results showed that (/sup 3/H)-rauwolscine bound specifically to sites in their membranal preparation, which had the pharmacological characteristics of the alpha2-adrenoceptors. The correlation between biochemical and pharmacological data revealed the usefulness of binding methods in the further study of adrenergic mechanisms in the canine saphenous vein.« less

LncRNA/DNA binding analysis reveals losses and gains and lineage specificity of genomic imprinting in mammals.

PubMed

Liu, Haihua; Shang, Xiaoxiao; Zhu, Hao

2017-05-15

Genomic imprinting is regulated by lncRNAs and is important for embryogenesis, physiology and behaviour in mammals. Aberrant imprinting causes diseases and disorders. Experimental studies have examined genomic imprinting primarily in humans and mice, thus leaving some fundamental issues poorly addressed. The cost of experimentally examining imprinted genes in many tissues in diverse species makes computational analysis of lncRNAs' DNA binding sites valuable. We performed lncRNA/DNA binding analysis in imprinting clusters from multiple mammalian clades and discovered the following: (i) lncRNAs and imprinting sites show significant losses and gains and distinct lineage-specificity; (ii) binding of lncRNAs to promoters of imprinted genes may occur widely throughout the genome; (iii) a considerable number of imprinting sites occur in only evolutionarily more derived species; and (iv) multiple lncRNAs may bind to the same imprinting sites, and some lncRNAs have multiple DNA binding motifs. These results suggest that the occurrence of abundant lncRNAs in mammalian genomes makes genomic imprinting a mechanism of adaptive evolution at the epigenome level. The data and program are available at the database LongMan at lncRNA.smu.edu.cn. zhuhao@smu.edu.cn. Supplementary data are available at Bioinformatics online. © The Author 2017. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com

Cyclophilin B binding to platelets supports calcium-dependent adhesion to collagen.

PubMed

Allain, F; Durieux, S; Denys, A; Carpentier, M; Spik, G

1999-08-01

We have recently reported that cyclophilin B (CyPB), a secreted cyclosporine-binding protein, could bind to T lymphocytes through interactions with two types of binding sites. The first ones, referred to as type I, involve interactions with the conserved domain of CyPB and promote the endocytosis of surface-bound ligand, while the second type of binding sites, termed type II, are represented by glycosaminoglycans (GAG). Here, we further investigated the interactions of CyPB with blood cell populations. In addition to lymphocytes, CyPB was found to interact mainly with platelets. The binding is specific, with a dissociation constant (kd) of 9 +/- 3 nmol/L and the number of sites estimated at 960 +/- 60 per cell. Platelet glycosaminoglycans are not required for the interactions, but the binding is dramatically reduced by active cyclosporine derivatives. We then analyzed the biologic effects of CyPB and found a significant increase in platelet adhesion to collagen. Concurrently, CyPB initiates a transmembranous influx of Ca(2+) and induces the phosphorylation of the P-20 light chains of myosin. Taken together, the present results demonstrate for the first time that extracellular CyPB specifically interacts with platelets through a functional receptor related to the lymphocyte type I binding sites and might act by regulating the activity of a receptor-operated membrane Ca(2+) channel.

Genome-wide analysis of AR binding and comparison with transcript expression in primary human fetal prostate fibroblasts and cancer associated fibroblasts.

PubMed

Nash, Claire; Boufaied, Nadia; Mills, Ian G; Franco, Omar E; Hayward, Simon W; Thomson, Axel A

2017-05-05

The androgen receptor (AR) is a transcription factor, and key regulator of prostate development and cancer, which has discrete functions in stromal versus epithelial cells. AR expressed in mesenchyme is necessary and sufficient for prostate development while loss of stromal AR is predictive of prostate cancer progression. Many studies have characterized genome-wide binding of AR in prostate tumour cells but none have used primary mesenchyme or stroma. We applied ChIPseq to identify genomic AR binding sites in primary human fetal prostate fibroblasts and patient derived cancer associated fibroblasts, as well as the WPMY1 cell line overexpressing AR. We identified AR binding sites that were specific to fetal prostate fibroblasts (7534), cancer fibroblasts (629), WPMY1-AR (2561) as well as those common among all (783). Primary fibroblasts had a distinct AR binding profile versus prostate cancer cell lines and tissue, and showed a localisation to gene promoter binding sites 1 kb upstream of the transcriptional start site, as well as non-classical AR binding sequence motifs. We used RNAseq to define transcribed genes associated with AR binding sites and derived cistromes for embryonic and cancer fibroblasts as well as a cistrome common to both. These were compared to several in vivo ChIPseq and transcript expression datasets; which identified subsets of AR targets that were expressed in vivo and regulated by androgens. This analysis enabled us to deconvolute stromal AR targets active in stroma within tumour samples. Taken together, our data suggest that the AR shows significantly different genomic binding site locations in primary prostate fibroblasts compared to that observed in tumour cells. Validation of our AR binding site data with transcript expression in vitro and in vivo suggests that the AR target genes we have identified in primary fibroblasts may contribute to clinically significant and biologically important AR-regulated changes in prostate tissue. Copyright © 2017. Published by Elsevier B.V.

Electrocorticographic high gamma activity versus electrical cortical stimulation mapping of naming.

PubMed

Sinai, Alon; Bowers, Christopher W; Crainiceanu, Ciprian M; Boatman, Dana; Gordon, Barry; Lesser, Ronald P; Lenz, Frederick A; Crone, Nathan E

2005-07-01

Subdural electrocorticographic (ECoG) recordings in patients undergoing epilepsy surgery have shown that functional activation is associated with event-related broadband gamma activity in a higher frequency range (>70 Hz) than previously studied in human scalp EEG. To investigate the utility of this high gamma activity (HGA) for mapping language cortex, we compared its neuroanatomical distribution with functional maps derived from electrical cortical stimulation (ECS), which remains the gold standard for predicting functional impairment after surgery for epilepsy, tumours or vascular malformations. Thirteen patients had undergone subdural electrode implantation for the surgical management of intractable epilepsy. Subdural ECoG signals were recorded while each patient verbally named sequentially presented line drawings of objects, and estimates of event-related HGA (80-100 Hz) were made at each recording site. Routine clinical ECS mapping used a subset of the same naming stimuli at each cortical site. If ECS disrupted mouth-related motor function, i.e. if it affected the mouth, lips or tongue, naming could not be tested with ECS at the same cortical site. Because naming during ECoG involved these muscles of articulation, the sensitivity and specificity of ECoG HGA were estimated relative to both ECS-induced impairments of naming and ECS disruption of mouth-related motor function. When these estimates were made separately for 12 electrode sites per patient (the average number with significant HGA), the specificity of ECoG HGA with respect to ECS was 78% for naming and 81% for mouth-related motor function, and equivalent sensitivities were 38% and 46%, respectively. When ECS maps of naming and mouth-related motor function were combined, the specificity and sensitivity of ECoG HGA with respect to ECS were 84% and 43%, respectively. This study indicates that event-related ECoG HGA during confrontation naming predicts ECS interference with naming and mouth-related motor function with good specificity but relatively low sensitivity. Its favourable specificity suggests that ECoG HGA can be used to construct a preliminary functional map that may help identify cortical sites of lower priority for ECS mapping. Passive recordings of ECoG gamma activity may be done simultaneously at all electrode sites without the risk of after-discharges associated with ECS mapping, which must be done sequentially at pairs of electrodes. We discuss the relative merits of these two functional mapping techniques.

Molecular architectures of benzoic acid-specific type III polyketide synthases

PubMed Central

Stewart, Charles; Woods, Kate; Macias, Greg; Allan, Andrew C.; Noel, Joseph P.

2017-01-01

Biphenyl synthase and benzophenone synthase constitute an evolutionarily distinct clade of type III polyketide synthases (PKSs) that use benzoic acid-derived substrates to produce defense metabolites in plants. The use of benzoyl-CoA as an endogenous substrate is unusual for type III PKSs. Moreover, sequence analyses indicate that the residues responsible for the functional diversification of type III PKSs are mutated in benzoic acid-specific type III PKSs. In order to gain a better understanding of structure–function relationships within the type III PKS family, the crystal structures of biphenyl synthase from Malus × domestica and benzophenone synthase from Hypericum androsaemum were compared with the structure of an archetypal type III PKS: chalcone synthase from Malus × domestica. Both biphenyl synthase and benzophenone synthase contain mutations that reshape their active-site cavities to prevent the binding of 4-coumaroyl-CoA and to favor the binding of small hydrophobic substrates. The active-site cavities of biphenyl synthase and benzophenone synthase also contain a novel pocket associated with their chain-elongation and cyclization reactions. Collectively, these results illuminate structural determinants of benzoic acid-specific type III PKSs and expand the understanding of the evolution of specialized metabolic pathways in plants. PMID:29199980

The structure of the Tiam1 PDZ domain/ phospho-syndecan1 complex reveals a ligand conformation that modulates protein dynamics.

PubMed

Liu, Xu; Shepherd, Tyson R; Murray, Ann M; Xu, Zhen; Fuentes, Ernesto J

2013-03-05

PDZ (PSD-95/Dlg/ZO-1) domains are protein-protein interaction modules often regulated by ligand phosphorylation. Here, we investigated the specificity, structure, and dynamics of Tiam1 PDZ domain/ligand interactions. We show that the PDZ domain specifically binds syndecan1 (SDC1), phosphorylated SDC1 (pSDC1), and SDC3 but not other syndecan isoforms. The crystal structure of the PDZ/SDC1 complex indicates that syndecan affinity is derived from amino acids beyond the four C-terminal residues. Remarkably, the crystal structure of the PDZ/pSDC1 complex reveals a binding pocket that accommodates the phosphoryl group. Methyl relaxation experiments of PDZ/SCD1 and PDZ/pSDC1 complexes reveal that PDZ-phosphoryl interactions dampen dynamic motions in a distal region of the PDZ domain by decoupling them from the ligand-binding site. Our data are consistent with a selection model by which specificity and phosphorylation regulate PDZ/syndecan interactions and signaling events. Importantly, our relaxation data demonstrate that PDZ/phospho-ligand interactions regulate protein dynamics and their coupling to distal sites. Copyright © 2013 Elsevier Ltd. All rights reserved.

Submolecular Gates Self-Assemble for Hot-Electron Transfer in Proteins.

PubMed

Filip-Granit, Neta; Goldberg, Eran; Samish, Ilan; Ashur, Idan; van der Boom, Milko E; Cohen, Hagai; Scherz, Avigdor

2017-07-27

Redox reactions play key roles in fundamental biological processes. The related spatial organization of donors and acceptors is assumed to undergo evolutionary optimization facilitating charge mobilization within the relevant biological context. Experimental information from submolecular functional sites is needed to understand the organization strategies and driving forces involved in the self-development of structure-function relationships. Here we exploit chemically resolved electrical measurements (CREM) to probe the atom-specific electrostatic potentials (ESPs) in artificial arrays of bacteriochlorophyll (BChl) derivatives that provide model systems for photoexcited (hot) electron donation and withdrawal. On the basis of computations we show that native BChl's in the photosynthetic reaction center (RC) self-assemble at their ground-state as aligned gates for functional charge transfer. The combined computational and experimental results further reveal how site-specific polarizability perpendicular to the molecular plane enhances the hot-electron transport. Maximal transport efficiency is predicted for a specific, ∼5 Å, distance above the center of the metalized BChl, which is in remarkably close agreement with the distance and mutual orientation of corresponding native cofactors. These findings provide new metrics and guidelines for analysis of biological redox centers and for designing charge mobilizing machines such as artificial photosynthesis.

HRP2 determines the efficiency and specificity of HIV-1 integration in LEDGF/p75 knockout cells but does not contribute to the antiviral activity of a potent LEDGF/p75-binding site integrase inhibitor.

PubMed

Wang, Hao; Jurado, Kellie A; Wu, Xiaolin; Shun, Ming-Chieh; Li, Xiang; Ferris, Andrea L; Smith, Steven J; Patel, Pratiq A; Fuchs, James R; Cherepanov, Peter; Kvaratskhelia, Mamuka; Hughes, Stephen H; Engelman, Alan

2012-12-01

The binding of integrase (IN) to lens epithelium-derived growth factor (LEDGF)/p75 in large part determines the efficiency and specificity of HIV-1 integration. However, a significant residual preference for integration into active genes persists in Psip1 (the gene that encodes for LEDGF/p75) knockout (KO) cells. One other cellular protein, HRP2, harbors both the PWWP and IN-binding domains that are important for LEDGF/p75 co-factor function. To assess the role of HRP2 in HIV-1 integration, cells generated from Hdgfrp2 (the gene that encodes for HRP2) and Psip1/Hdgfrp2 KO mice were infected alongside matched control cells. HRP2 depleted cells supported normal infection, while disruption of Hdgfrp2 in Psip1 KO cells yielded additional defects in the efficiency and specificity of integration. These deficits were largely restored by ectopic expression of either LEDGF/p75 or HRP2. The double-KO cells nevertheless supported residual integration into genes, indicating that IN and/or other host factors contribute to integration specificity in the absence of LEDGF/p75 and HRP2. Psip1 KO significantly increased the potency of an allosteric inhibitor that binds the LEDGF/p75 binding site on IN, a result that was not significantly altered by Hdgfrp2 disruption. These findings help to rule out the host factor-IN interactions as the primary antiviral targets of LEDGF/p75-binding site IN inhibitors.

Normalized difference vegetation index as an estimator for abundance and quality of avian herbivore forage in arctic Alaska

USGS Publications Warehouse

Hogrefe, Kyle R.; Patil, Vijay; Ruthrauff, Daniel R.; Meixell, Brandt W.; Budde, Michael E.; Hupp, Jerry W.; Ward, David H.

2017-01-01

Tools that can monitor biomass and nutritional quality of forage plants are needed to understand how arctic herbivores may respond to the rapidly changing environment at high latitudes. The Normalized Difference Vegetation Index (NDVI) has been widely used to assess changes in abundance and distribution of terrestrial vegetative communities. However, the efficacy of NDVI to measure seasonal changes in biomass and nutritional quality of forage plants in the Arctic remains largely un-evaluated at landscape and fine-scale levels. We modeled the relationships between NDVI and seasonal changes in aboveground biomass and nitrogen concentration in halophytic graminoids, a key food source for arctic-nesting geese. The model was calibrated based on data collected at one site and validated using data from another site. Effects of spatial scale on model accuracy were determined by comparing model predictions between NDVI derived from moderate resolution (250 × 250 m pixels) satellite data and high resolution (20 cm diameter area) handheld spectrometer data. NDVI derived from the handheld spectrometer was a superior estimator (R2 ≥ 0.67) of seasonal changes in aboveground biomass compared to satellite-derived NDVI (R2 ≤ 0.40). The addition of temperature and precipitation variables to the model for biomass improved fit, but provided minor gains in predictive power beyond that of the NDVI-only model. This model, however, was only a moderately accurate estimator of biomass in an ecologically-similar halophytic graminoid wetland located 100 km away, indicating the necessity for site-specific validation. In contrast to assessments of biomass, satellite-derived NDVI was a better estimator for the timing of peak percent of nitrogen than NDVI derived from the handheld spectrometer. We confirmed that the date when NDVI reached 50% of its seasonal maximum was a reasonable approximation of the period of peak spring vegetative green-up and peak percent nitrogen. This study demonstrates the importance of matching the scale of NDVI measurements to the vegetation properties of biomass and nitrogen phenology.

Physical locations of 5S and 18S-25S rDNA in Asian and American diploid Hordeum species with the I genome.

PubMed

Taketa, S; Ando, H; Takeda, K; von Bothmer, R

2001-05-01

The physical locations of 5S and 18S-25S rDNA sequences in 15 diploid Hordeum species with the I genome were examined by double-target in situ hybridization with pTa71 (18S-25S rDNA) and pTa794 (5S rDNA) clones as probes. All the three Asian species had a species-specific rDNA pattern. In 12 American species studied, eight different rDNA types were found. The type reported previously in H. chilense (the 'chilense' type) was observed in eight American species. The chilense type had double 5S rDNA sites - two sites on one chromosome arm separated by a short distance - and two pairs of major 18S-25S rDNA sites on two pairs of satellite chromosomes. The other seven types found in American species were similar to the chilense type and could be derived from the chilense type through deletion, reduction or addition of a rDNA site. Intraspecific polymorphisms were observed in three American species. The overall similarity in rDNA patterns among American species indicates the close relationships between North and South American species and their derivation from a single ancestral source. The differences in the distribution patterns of 5S and 18S-25S rDNA between Asian and American species suggest differentiation between the I genomes of Asian and American species. The 5S and 18S-25S rDNA sites are useful chromosome markers for delimiting Asian species, but have limited value as a taxonomic character in American species. On the basis of rDNA patterns, karyotype evolution and phylogeny of the I-genome diploid species are discussed.

Prediction of Protein Modification Sites of Pyrrolidone Carboxylic Acid Using mRMR Feature Selection and Analysis

PubMed Central

Zheng, Lu-Lu; Niu, Shen; Hao, Pei; Feng, KaiYan; Cai, Yu-Dong; Li, Yixue

2011-01-01

Pyrrolidone carboxylic acid (PCA) is formed during a common post-translational modification (PTM) of extracellular and multi-pass membrane proteins. In this study, we developed a new predictor to predict the modification sites of PCA based on maximum relevance minimum redundancy (mRMR) and incremental feature selection (IFS). We incorporated 727 features that belonged to 7 kinds of protein properties to predict the modification sites, including sequence conservation, residual disorder, amino acid factor, secondary structure and solvent accessibility, gain/loss of amino acid during evolution, propensity of amino acid to be conserved at protein-protein interface and protein surface, and deviation of side chain carbon atom number. Among these 727 features, 244 features were selected by mRMR and IFS as the optimized features for the prediction, with which the prediction model achieved a maximum of MCC of 0.7812. Feature analysis showed that all feature types contributed to the modification process. Further site-specific feature analysis showed that the features derived from PCA's surrounding sites contributed more to the determination of PCA sites than other sites. The detailed feature analysis in this paper might provide important clues for understanding the mechanism of the PCA formation and guide relevant experimental validations. PMID:22174779

Prediction of lysine ubiquitylation with ensemble classifier and feature selection.

PubMed

Zhao, Xiaowei; Li, Xiangtao; Ma, Zhiqiang; Yin, Minghao

2011-01-01

Ubiquitylation is an important process of post-translational modification. Correct identification of protein lysine ubiquitylation sites is of fundamental importance to understand the molecular mechanism of lysine ubiquitylation in biological systems. This paper develops a novel computational method to effectively identify the lysine ubiquitylation sites based on the ensemble approach. In the proposed method, 468 ubiquitylation sites from 323 proteins retrieved from the Swiss-Prot database were encoded into feature vectors by using four kinds of protein sequences information. An effective feature selection method was then applied to extract informative feature subsets. After different feature subsets were obtained by setting different starting points in the search procedure, they were used to train multiple random forests classifiers and then aggregated into a consensus classifier by majority voting. Evaluated by jackknife tests and independent tests respectively, the accuracy of the proposed predictor reached 76.82% for the training dataset and 79.16% for the test dataset, indicating that this predictor is a useful tool to predict lysine ubiquitylation sites. Furthermore, site-specific feature analysis was performed and it was shown that ubiquitylation is intimately correlated with the features of its surrounding sites in addition to features derived from the lysine site itself. The feature selection method is available upon request.

Myeloid-derived suppressor cell functionality and interaction with Leishmania major parasites differ in C57BL/6 and BALB/c mice.

PubMed

Schmid, Maximilian; Zimara, Nicole; Wege, Anja Kathrin; Ritter, Uwe

2014-11-01

Myeloid-derived suppressor cells (MDSCs) represent a heterogeneous population of CD11b+ cells. According to the surface molecules Ly6G and Ly6C (where Ly6G and Ly6C are lymphocyte antigen 6, locus G and C, respectively), MDSCs are further divided into monocytic (Mo-MDSCs, CD11b+ /Ly6C(high) /Ly6G-) and polymorphonucleated suppressor cells (PMN-MDSCs, CD11b+ /Ly6C(int) /Ly6G+). Most published manuscripts focus on the suppressive role of MDSCs in cancer, whereas their impact on adaptive immunity against obligatory intracellular parasites is not well understood. Furthermore, it is not clear how the genetic background of mice influences MDSC functionality. Therefore, we implemented an experimental model of leishmaniasis, and analyzed MDSC maturation and the impact of MDSCs on the parasite-specific T-cell responses in resistant C57BL/6 and susceptible BALB/c mice. This experimental setup demonstrated the impaired ability of BALB/c mice to produce Mo-MDSCs when compared with C57BL/6 mice. This phenotype is detectable after subcutaneous infection with parasites and is specifically represented by a reduced accumulation of Mo-MDSCs at the site of infection in BALB/c mice. Moreover, infected C57BL/6-derived MDSCs were able to suppress Leishmania-specific CD4+ -cell proliferation, whereas BALB/c-derived MDSCs harboring parasites lost this suppressive function. In conclusion, we demonstrate that (i) genetic background defines MDSC differentiation; and (ii) Leishmania major parasites are able to modulate the suppressive effect of MDSCs in a strain-dependent manner. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Antibody induced CD4 down-modulation of T cells is site-specifically mediated by CD64+ cells

PubMed Central

Vogel, Stephanie; Grabski, Elena; Buschjäger, Daniela; Klawonn, Frank; Döring, Marius; Wang, Junxi; Fletcher, Erika; Bechmann, Ingo; Witte, Torsten; Durisin, Martin; Schraven, Burkhart; Mangsbo, Sara M.; Schönfeld, Kurt; Czeloth, Niklas; Kalinke, Ulrich

2015-01-01

Treatment of PBMC with the CD4-specific mAb BT-061 induces CD4 down-modulation of T cells. Here we report that addition of BT-061 to purified T cells did not confer this effect, whereas incubation of T cells in BT-061 coated wells restored CD4 down-modulation. These results implied that Fcγ receptor mediated cell-cell interactions played a role. In consistence with this hypothesis PBMC depleted of CD64+ monocytes did not confer CD4 down-modulation of BT-061 decorated T cells. Strikingly, CD4 down-modulation was observed in BT-061 treated synovial fluid punctuated from patients’ inflamed joints that comprised enhanced numbers of CD64+ cells. In contrast, in a circulating whole blood system injection of BT-061 did not induce CD4 down-modulation, due to CD64 saturation by serum IgG. Similarly, tonsil derived mononuclear cells devoid of CD64+ cells did not show CD4 down-modulation, whereas addition of blood derived monocytes restored the effect. Thus, the interaction of BT-061 decorated T cells with CD64+ cells is needed for CD4 down-modulation, implying that in patients BT-061 would primarily induce CD4 down-modulation at inflammatory sites. These results highlight the need not only to examine the interaction of a given mAb with single FcγR, but also the immunological environment that is appropriate to support such interactions. PMID:26670584

Computational analysis of protein-protein interfaces involving an alpha helix: insights for terphenyl-like molecules binding.

PubMed

Isvoran, Adriana; Craciun, Dana; Martiny, Virginie; Sperandio, Olivier; Miteva, Maria A

2013-06-14

Protein-Protein Interactions (PPIs) are key for many cellular processes. The characterization of PPI interfaces and the prediction of putative ligand binding sites and hot spot residues are essential to design efficient small-molecule modulators of PPI. Terphenyl and its derivatives are small organic molecules known to mimic one face of protein-binding alpha-helical peptides. In this work we focus on several PPIs mediated by alpha-helical peptides. We performed computational sequence- and structure-based analyses in order to evaluate several key physicochemical and surface properties of proteins known to interact with alpha-helical peptides and/or terphenyl and its derivatives. Sequence-based analysis revealed low sequence identity between some of the analyzed proteins binding alpha-helical peptides. Structure-based analysis was performed to calculate the volume, the fractal dimension roughness and the hydrophobicity of the binding regions. Besides the overall hydrophobic character of the binding pockets, some specificities were detected. We showed that the hydrophobicity is not uniformly distributed in different alpha-helix binding pockets that can help to identify key hydrophobic hot spots. The presence of hydrophobic cavities at the protein surface with a more complex shape than the entire protein surface seems to be an important property related to the ability of proteins to bind alpha-helical peptides and low molecular weight mimetics. Characterization of similarities and specificities of PPI binding sites can be helpful for further development of small molecules targeting alpha-helix binding proteins.

Site-specific differences of insulin action in adipose tissue derived from normal prepubertal children.

PubMed

Grohmann, Malcolm; Stewart, Claire; Welsh, Gavin; Hunt, Linda; Tavaré, Jeremy; Holly, Jeff; Shield, Julian; Sabin, Matt; Crowne, Elizabeth

2005-08-15

Body fat distribution determines obesity-related morbidity in adults but little is known of the aetiology or pathophysiology in children. This study investigates differences in insulin-mediated metabolism in primary cell cultures of subcutaneous and visceral preadipocytes derived from prepubertal children. The impact of differentiation and responses to TNFalpha exposure was also investigated. Proliferation rates were greater in subcutaneous versus visceral preadipocytes (41 h3 versus 69 h4; P=0.008). Insulin caused a dose-dependent increase in GSK-3 phosphorylation and an increase in MAPK phosphorylation over time, with increased sensitivity in subcutaneous preadipocytes. Post-differentiation, dose-dependent increases in GSK-3 phosphorylation were maintained, while MAPK phosphorylation was identical in both subtypes. No changes were observed in insulin receptor abundance pre-/post-differentiation. GLUT4 abundance was significantly increased in visceral versus subcutaneous adipocytes by 76(4)%; P=0.03), coincidental with increased insulin-stimulated 2-deoxy-glucose transport (+150(26)% versus +79(10)%; P=0.014) and further elevated by acute exposure to TNFalpha (+230(52)%; P=0.019 versus +123(24)%; P=0.025, respectively). TNFalpha also significantly increased basal glucose transport rates (+44(14)%; P=0.006 versus +34(11)%; P=0.007) and GLUT1 localisation to the plasma membrane. These data establish site-specific differences in subcutaneous and visceral fat cells from children. Responses to insulin varied with differentiation and TNFalpha exposure in the two depots, consistent with parallel changes in GLUT1/4 abundance and localisation.

Infusion of donor-derived CD19-redirected virus-specific T cells for B-cell malignancies relapsed after allogeneic stem cell transplant: a phase 1 study.

PubMed

Cruz, Conrad Russell Y; Micklethwaite, Kenneth P; Savoldo, Barbara; Ramos, Carlos A; Lam, Sharon; Ku, Stephanie; Diouf, Oumar; Liu, Enli; Barrett, A John; Ito, Sawa; Shpall, Elizabeth J; Krance, Robert A; Kamble, Rammurti T; Carrum, George; Hosing, Chitra M; Gee, Adrian P; Mei, Zhuyong; Grilley, Bambi J; Heslop, Helen E; Rooney, Cliona M; Brenner, Malcolm K; Bollard, Catherine M; Dotti, Gianpietro

2013-10-24

Autologous T cells expressing a CD19-specific chimeric antigen receptor (CD19.CAR) are active against B-cell malignancies, but it is unknown whether allogeneic CD19.CAR T cells are safe or effective. After allogeneic hematopoietic stem cell transplantation (HSCT), infused donor-derived virus-specific T cells (VSTs) expand in vivo, persist long term, and display antiviral activity without inducing graft-vs-host disease; therefore, we determined whether donor VSTs, engineered to express CD19.CAR, retained the characteristics of nonmanipulated allogeneic VSTs while gaining antitumor activity. We treated 8 patients with allogeneic (donor-derived) CD19.CAR-VSTs 3 months to 13 years after HSCT. There were no infusion-related toxicities. VSTs persisted for a median of 8 weeks in blood and up to 9 weeks at disease sites. Objective antitumor activity was evident in 2 of 6 patients with relapsed disease during the period of CD19.CAR-VST persistence, whereas 2 patients who received cells while in remission remain disease free. In 2 of 3 patients with viral reactivation, donor CD19.CAR-VSTs expanded concomitantly with VSTs. Hence CD19.CAR-VSTs display antitumor activity and, because their number may be increased in the presence of viral stimuli, earlier treatment post-HSCT (when lymphodepletion is greater and the incidence of viral infection is higher) or planned vaccination with viral antigens may enhance disease control.

Perfluorocarbon Nanoparticles as Delivery Vehicles for Melittin and Its Protease-Activated Derivatives

NASA Astrophysics Data System (ADS)

Jallouk, Andrew Philip

Melittin is a cytolytic peptide derived from honeybee venom which inserts into lipid membranes and oligomerizes to form membrane pores. While this peptide is an attractive candidate for treatment of cancers and infectious processes, its nonspecific cytotoxicity and hemolytic activity have limited its therapeutic applications. The goal of this dissertation was to enhance the specificity of melittin therapy through the use of perfluorocarbon nanoparticles to minimize nonspecific cytotoxicity and the development of melittin prodrugs which only exhibit cytolytic activity following activation by site-specific proteases. Although previous studies have characterized the biological effects of melittin-loaded nanoparticles following intravenous administration, we first investigated their use as topical agents for prevention of HIV infection. We found that incorporation of native melittin onto perfluorocarbon nanoparticles maintained antiviral activity while substantially reducing contact toxicity to sperm and vaginal epithelium. These results demonstrated the potential utility of melittin-loaded nanoparticles as a topical vaginal virucide. To further enhance melittin specificity, we developed melittin derivatives which could be activated by matrix metalloproteinase-9, a protease which is overexpressed in many tumors and which plays a critical role in cancer invasion and metastasis. We then characterized the interactions of these peptides with perfluorocarbon nanoparticles and demonstrated the safety and efficacy of intravenous prodrug-loaded nanoparticle therapy in a mouse model of melanoma. The versatility of this platform could facilitate development of personalized cancer therapies directed towards a patient's individual protease expression profile.

Conservation planning for offsetting the impacts of development: a case study of biodiversity and renewable energy in the Mojave Desert

USGS Publications Warehouse

Kreitler, Jason R.; Schloss, Carrie A.; Soong, Oliver; Lee Hannah,; Davis, Frank W.

2015-01-01

Balancing society’s competing needs of development and conservation requires careful consideration of tradeoffs. Renewable energy development and biodiversity conservation are often considered beneficial environmental goals. The direct footprint and disturbance of renewable energy, however, can displace species’ habitat and negatively impact populations and natural communities if sited without ecological consideration. Offsets have emerged as a potentially useful tool to mitigate residual impacts after trying to avoid, minimize, or restore affected sites. Yet the problem of efficiently designing a set of offset sites becomes increasingly complex where many species or many sites are involved. Spatial conservation prioritization tools are designed to handle this problem, but have seen little application to offset siting and analysis. To address this need we designed an offset siting support tool for the Desert Renewable Energy Conservation Plan (DRECP) of California, and present a case study of hypothetical impacts from solar development in the Western Mojave subsection. We compare two offset scenarios designed to mitigate a hypothetical 15,331 ha derived from proposed utility-scale solar energy development (USSED) projects. The first scenario prioritizes offsets based precisely on impacted features, while the second scenario offsets impacts to maximize biodiversity conservation gains in the region. The two methods only agree on 28% of their prioritized sites and differ in meeting species-specific offset goals. Differences between the two scenarios highlight the importance of clearly specifying choices and priorities for offset siting and mitigation in general. Similarly, the effects of background climate and land use change may lessen the durability or effectiveness of offsets if not considered. Our offset siting support tool was designed specifically for the DRECP area, but with minor code modification could work well in other offset analyses, and could provide continuing support for a potentially innovative mitigation solution to environmental impacts.

Conservation Planning for Offsetting the Impacts of Development: A Case Study of Biodiversity and Renewable Energy in the Mojave Desert.

PubMed

Kreitler, Jason; Schloss, Carrie A; Soong, Oliver; Hannah, Lee; Davis, Frank W

2015-01-01

Balancing society's competing needs of development and conservation requires careful consideration of tradeoffs. Renewable energy development and biodiversity conservation are often considered beneficial environmental goals. The direct footprint and disturbance of renewable energy, however, can displace species' habitat and negatively impact populations and natural communities if sited without ecological consideration. Offsets have emerged as a potentially useful tool to mitigate residual impacts after trying to avoid, minimize, or restore affected sites. Yet the problem of efficiently designing a set of offset sites becomes increasingly complex where many species or many sites are involved. Spatial conservation prioritization tools are designed to handle this problem, but have seen little application to offset siting and analysis. To address this need we designed an offset siting support tool for the Desert Renewable Energy Conservation Plan (DRECP) of California, and present a case study of hypothetical impacts from solar development in the Western Mojave subsection. We compare two offset scenarios designed to mitigate a hypothetical 15,331 ha derived from proposed utility-scale solar energy development (USSED) projects. The first scenario prioritizes offsets based precisely on impacted features, while the second scenario offsets impacts to maximize biodiversity conservation gains in the region. The two methods only agree on 28% of their prioritized sites and differ in meeting species-specific offset goals. Differences between the two scenarios highlight the importance of clearly specifying choices and priorities for offset siting and mitigation in general. Similarly, the effects of background climate and land use change may lessen the durability or effectiveness of offsets if not considered. Our offset siting support tool was designed specifically for the DRECP area, but with minor code modification could work well in other offset analyses, and could provide continuing support for a potentially innovative mitigation solution to environmental impacts.

Conservation Planning for Offsetting the Impacts of Development: A Case Study of Biodiversity and Renewable Energy in the Mojave Desert

PubMed Central

Kreitler, Jason; Schloss, Carrie A.; Soong, Oliver; Hannah, Lee; Davis, Frank W.

2015-01-01

Balancing society’s competing needs of development and conservation requires careful consideration of tradeoffs. Renewable energy development and biodiversity conservation are often considered beneficial environmental goals. The direct footprint and disturbance of renewable energy, however, can displace species’ habitat and negatively impact populations and natural communities if sited without ecological consideration. Offsets have emerged as a potentially useful tool to mitigate residual impacts after trying to avoid, minimize, or restore affected sites. Yet the problem of efficiently designing a set of offset sites becomes increasingly complex where many species or many sites are involved. Spatial conservation prioritization tools are designed to handle this problem, but have seen little application to offset siting and analysis. To address this need we designed an offset siting support tool for the Desert Renewable Energy Conservation Plan (DRECP) of California, and present a case study of hypothetical impacts from solar development in the Western Mojave subsection. We compare two offset scenarios designed to mitigate a hypothetical 15,331 ha derived from proposed utility-scale solar energy development (USSED) projects. The first scenario prioritizes offsets based precisely on impacted features, while the second scenario offsets impacts to maximize biodiversity conservation gains in the region. The two methods only agree on 28% of their prioritized sites and differ in meeting species-specific offset goals. Differences between the two scenarios highlight the importance of clearly specifying choices and priorities for offset siting and mitigation in general. Similarly, the effects of background climate and land use change may lessen the durability or effectiveness of offsets if not considered. Our offset siting support tool was designed specifically for the DRECP area, but with minor code modification could work well in other offset analyses, and could provide continuing support for a potentially innovative mitigation solution to environmental impacts. PMID:26529595

Heat, chloride, and specific conductance as ground water tracers near streams

USGS Publications Warehouse

Cox, M.H.; Su, G.W.; Constantz, J.

2007-01-01

Commonly measured water quality parameters were compared to heat as tracers of stream water exchange with ground water. Temperature, specific conductance, and chloride were sampled at various frequencies in the stream and adjacent wells over a 2-year period. Strong seasonal variations in stream water were observed for temperature and specific conductance. In observation wells where the temperature response correlated to stream water, chloride and specific conductance values were similar to stream water values as well, indicating significant stream water exchange with ground water. At sites where ground water temperature fluctuations were negligible, chloride and/or specific conductance values did not correlate to stream water values, indicating that ground water was not significantly influenced by exchange with stream water. Best-fit simulation modeling was performed at two sites to derive temperature-based estimates of hydraulic conductivities of the alluvial sediments between the stream and wells. These estimates were used in solute transport simulations for a comparison of measured and simulated values for chloride and specific conductance. Simulation results showed that hydraulic conductivities vary seasonally and annually. This variability was a result of seasonal changes in temperature-dependent hydraulic conductivity and scouring or clogging of the streambed. Specific conductance fits were good, while chloride data were difficult to fit due to the infrequent (quarterly) stream water chloride measurements during the study period. Combined analyses of temperature, chloride, and specific conductance led to improved quantification of the spatial and temporal variability of stream water exchange with shallow ground water in an alluvial system. ?? 2007 National Ground Water Association.

An evaluation of terrain-based downscaling of fractional snow covered area data sets based on LiDAR-derived snow data and orthoimagery

NASA Astrophysics Data System (ADS)

Cristea, Nicoleta C.; Breckheimer, Ian; Raleigh, Mark S.; HilleRisLambers, Janneke; Lundquist, Jessica D.

2017-08-01

Reliable maps of snow-covered areas at scales of meters to tens of meters, with daily temporal resolution, are essential to understanding snow heterogeneity, melt runoff, energy exchange, and ecological processes. Here we develop a parsimonious downscaling routine that can be applied to fractional snow covered area (fSCA) products from satellite platforms such as the Moderate Resolution Imaging Spectroradiometer (MODIS) that provide daily ˜500 m data, to derive higher-resolution snow presence/absence grids. The method uses a composite index combining both the topographic position index (TPI) to represent accumulation effects and the diurnal anisotropic heat (DAH, sun exposure) index to represent ablation effects. The procedure is evaluated and calibrated using airborne-derived high-resolution data sets across the Tuolumne watershed, CA using 11 scenes in 2014 to downscale to 30 m resolution. The average matching F score was 0.83. We then tested our method's transferability in time and space by comparing against the Tuolumne watershed in water years 2013 and 2015, and over an entirely different site, Mt. Rainier, WA in 2009 and 2011, to assess applicability to other topographic and climatic conditions. For application to sites without validation data, we recommend equal weights for the TPI and DAH indices and close TPI neighborhoods (60 and 27 m for downscaling to 30 and 3 m, respectively), which worked well in both our study areas. The method is less effective in forested areas, which still requires site-specific treatment. We demonstrate that the procedure can even be applied to downscale to 3 m resolution, a very fine scale relevant to alpine ecohydrology research.

Computer-aided active-site-directed modeling of the Herpes Simplex Virus 1 and human thymidine kinase

NASA Astrophysics Data System (ADS)

Folkers, Gerd; Trumpp-Kallmeyer, Susanne; Gutbrod, Oliver; Krickl, Sabine; Fetzer, Jürgen; Keil, Günther M.

1991-10-01

Thymidine kinase (TK), which is induced by Herpes Simplex Virus 1 (HSV1), plays a key role in the antiviral activity of guanine derivatives such as aciclovir (ACV). In contrast, ACV shows only low affinity to the corresponding host cell enzyme. In order to define the differences in substrate binding of the two enzymes on molecular level, models for the three-dimensional (3-D) structures of the active sites of HSV1-TK and human TK were developed. The reconstruction of the active sites started from primary and secondary structure analysis of various kinases. The results were validated to homologous enzymes with known 3-D structures. The models predict that both enzymes consist of a central core β-sheet structure, connected by loops and α-helices very similar to the overall structure of other nucleotide binding enzymes. The phosphate binding is made up of a highly conserved glycine-rich loop at the N-terminus of the proteins and a conserved region at the C-terminus. The thymidine recognition site was found about 100 amino acids downstream from the phosphate binding loop. The differing substrate specificity of human and HSV1-TK can be explained by amino-acid substitutions in the homologous regions. To achieve a better understanding of the structure of the active site and how the thymidine kinase proteins interact with their substrates, the corresponding complexes of thymidine and dihydroxypropoxyguanine (DHPG) with HSV1 and human TK were built. For the docking of the guanine derivative, the X-ray structure of Elongation Factor Tu (EF-Tu), co-crystallized with guanosine diphosphate, was taken as reference. Fitting of thymidine into the active sites was done with respect to similar interactions found in thymidylate kinase. To complement the analysis of the 3-D structures of the two kinases and the substrate enzyme interactions, site-directed mutagenesis of the thymidine recognition site of HSV1-TK has been undertaken, changing Asp162 in the thymidine recognition site into Asn. First investigations reveal that the enzymatic activity of the mutant protein is destroyed.

Quantification by qPCR of Pathobionts in Chronic Periodontitis: Development of Predictive Models of Disease Severity at Site-Specific Level.

PubMed

Tomás, Inmaculada; Regueira-Iglesias, Alba; López, Maria; Arias-Bujanda, Nora; Novoa, Lourdes; Balsa-Castro, Carlos; Tomás, Maria

2017-01-01

Currently, there is little evidence available on the development of predictive models for the diagnosis or prognosis of chronic periodontitis based on the qPCR quantification of subgingival pathobionts. Our objectives were to: (1) analyze and internally validate pathobiont-based models that could be used to distinguish different periodontal conditions at site-specific level within the same patient with chronic periodontitis; (2) develop nomograms derived from predictive models. Subgingival plaque samples were obtained from control and periodontal sites (probing pocket depth and clinical attachment loss <4 mm and >4 mm, respectively) from 40 patients with moderate-severe generalized chronic periodontitis. The samples were analyzed by qPCR using TaqMan probes and specific primers to determine the concentrations of Actinobacillus actinomycetemcomitans (Aa) , Fusobacterium nucleatum (Fn) , Parvimonas micra (Pm) , Porphyromonas gingivalis (Pg) , Prevotella intermedia (Pi) , Tannerella forsythia (Tf) , and Treponema denticola (Td) . The pathobiont-based models were obtained using multivariate binary logistic regression. The best models were selected according to specified criteria. The discrimination was assessed using receiver operating characteristic curves and numerous classification measures were thus obtained. The nomograms were built based on the best predictive models. Eight bacterial cluster-based models showed an area under the curve (AUC) ≥0.760 and a sensitivity and specificity ≥75.0%. The PiTfFn cluster showed an AUC of 0.773 (sensitivity and specificity = 75.0%). When Pm and AaPm were incorporated in the TdPiTfFn cluster, we detected the two best predictive models with an AUC of 0.788 and 0.789, respectively (sensitivity and specificity = 77.5%). The TdPiTfAa cluster had an AUC of 0.785 (sensitivity and specificity = 75.0%). When Pm was incorporated in this cluster, a new predictive model appeared with better AUC and specificity values (0.787 and 80.0%, respectively). Distinct clusters formed by species with different etiopathogenic role (belonging to different Socransky's complexes) had a good predictive accuracy for distinguishing a site with periodontal destruction in a periodontal patient. The predictive clusters with the lowest number of bacteria were PiTfFn and TdPiTfAa , while TdPiTfAaFnPm had the highest number. In all the developed nomograms, high concentrations of these clusters were associated with an increased probability of having a periodontal site in a patient with chronic periodontitis.

Quantification by qPCR of Pathobionts in Chronic Periodontitis: Development of Predictive Models of Disease Severity at Site-Specific Level

PubMed Central

Tomás, Inmaculada; Regueira-Iglesias, Alba; López, Maria; Arias-Bujanda, Nora; Novoa, Lourdes; Balsa-Castro, Carlos; Tomás, Maria

2017-01-01

Currently, there is little evidence available on the development of predictive models for the diagnosis or prognosis of chronic periodontitis based on the qPCR quantification of subgingival pathobionts. Our objectives were to: (1) analyze and internally validate pathobiont-based models that could be used to distinguish different periodontal conditions at site-specific level within the same patient with chronic periodontitis; (2) develop nomograms derived from predictive models. Subgingival plaque samples were obtained from control and periodontal sites (probing pocket depth and clinical attachment loss <4 mm and >4 mm, respectively) from 40 patients with moderate-severe generalized chronic periodontitis. The samples were analyzed by qPCR using TaqMan probes and specific primers to determine the concentrations of Actinobacillus actinomycetemcomitans (Aa), Fusobacterium nucleatum (Fn), Parvimonas micra (Pm), Porphyromonas gingivalis (Pg), Prevotella intermedia (Pi), Tannerella forsythia (Tf), and Treponema denticola (Td). The pathobiont-based models were obtained using multivariate binary logistic regression. The best models were selected according to specified criteria. The discrimination was assessed using receiver operating characteristic curves and numerous classification measures were thus obtained. The nomograms were built based on the best predictive models. Eight bacterial cluster-based models showed an area under the curve (AUC) ≥0.760 and a sensitivity and specificity ≥75.0%. The PiTfFn cluster showed an AUC of 0.773 (sensitivity and specificity = 75.0%). When Pm and AaPm were incorporated in the TdPiTfFn cluster, we detected the two best predictive models with an AUC of 0.788 and 0.789, respectively (sensitivity and specificity = 77.5%). The TdPiTfAa cluster had an AUC of 0.785 (sensitivity and specificity = 75.0%). When Pm was incorporated in this cluster, a new predictive model appeared with better AUC and specificity values (0.787 and 80.0%, respectively). Distinct clusters formed by species with different etiopathogenic role (belonging to different Socransky’s complexes) had a good predictive accuracy for distinguishing a site with periodontal destruction in a periodontal patient. The predictive clusters with the lowest number of bacteria were PiTfFn and TdPiTfAa, while TdPiTfAaFnPm had the highest number. In all the developed nomograms, high concentrations of these clusters were associated with an increased probability of having a periodontal site in a patient with chronic periodontitis. PMID:28848499

Variable promoter methylation contributes to differential expression of key genes in human placenta-derived venous and arterial endothelial cells.

PubMed

Joo, Jihoon E; Hiden, Ursula; Lassance, Luciana; Gordon, Lavinia; Martino, David J; Desoye, Gernot; Saffery, Richard

2013-07-15

The endothelial compartment, comprising arterial, venous and lymphatic cell types, is established prenatally in association with rapid phenotypic and functional changes. The molecular mechanisms underpinning this process in utero have yet to be fully elucidated. The aim of this study was to investigate the potential for DNA methylation to act as a driver of the specific gene expression profiles of arterial and venous endothelial cells. Placenta-derived venous and arterial endothelial cells were collected at birth prior to culturing. DNA methylation was measured at >450,000 CpG sites in parallel with expression measurements taken from 25,000 annotated genes. A consistent set of genomic loci was found to show coordinate differential methylation between the arterial and venous cell types. This included many loci previously not investigated in relation to endothelial function. An inverse relationship was observed between gene expression and promoter methylation levels for a limited subset of genes implicated in endothelial function, including NOS3, encoding endothelial Nitric Oxide Synthase. Endothelial cells derived from the placental vasculature at birth contain widespread methylation of key regulatory genes. These are candidates involved in the specification of different endothelial cell types and represent potential target genes for environmentally mediated epigenetic disruption in utero in association with cardiovascular disease risk later in life.

Seamless correction of the sickle cell disease mutation of the HBB gene in human induced pluripotent stem cells using TALENs.

PubMed

Sun, Ning; Zhao, Huimin

2014-05-01

Sickle cell disease (SCD) is the most common human genetic disease which is caused by a single mutation of human β-globin (HBB) gene. The lack of long-term treatment makes the development of reliable cell and gene therapies highly desirable. Disease-specific patient-derived human induced pluripotent stem cells (hiPSCs) have great potential for developing novel cell and gene therapies. With the disease-causing mutations corrected in situ, patient-derived hiPSCs can restore normal cell functions and serve as a renewable autologous cell source for the treatment of genetic disorders. Here we successfully utilized transcription activator-like effector nucleases (TALENs), a recently emerged novel genome editing tool, to correct the SCD mutation in patient-derived hiPSCs. The TALENs we have engineered are highly specific and generate minimal off-target effects. In combination with piggyBac transposon, TALEN-mediated gene targeting leaves no residual ectopic sequences at the site of correction and the corrected hiPSCs retain full pluripotency and a normal karyotype. Our study demonstrates an important first step of using TALENs for the treatment of genetic diseases such as SCD, which represents a significant advance toward hiPSC-based cell and gene therapies. © 2013 Wiley Periodicals, Inc.

Synthesis and olfactory activity of unnatural, sulfated 5β-bile acid derivatives in the sea lamprey (Petromyzon marinus)

PubMed Central

Burns, Aaron C.; Sorensen, Peter W.

2011-01-01

A variety of unnatural bile acid derivatives (9a–9f) were synthesized and used to examine the specificity with which the sea lamprey (Petromyzon marinus) olfactory system detects these compounds. These compounds are analogs of petromyzonol sulfate (PS, 1), a component of the sea lamprey migratory pheromone. Both the stereochemical configuration at C5 (i.e., 5α vs. 5β) and the extent and sites of oxygenation (hydroxylation or ketonization) of the bile acid derived steroid skeleton were evaluated by screening the compounds for olfactory activity using electro-olfactogram recording. 5β-Petromyzonol sulfate (9a) elicited a considerable olfactory response at sub-nanomolar concentration. In addition, less oxygenated systems (i.e., 9b–9e) elicited olfactory responses, albeit with less potency. The sea lamprey sex pheromone mimic 9f (5β-3-ketopetromyzonol sulfate) was also examined and found to produce a much lower olfactory response. Mixture studies conducted with 9a and PS (1) suggest that stimulation is occurring via similar modes of activation, demonstrating a relative lack of specificity for recognition of the allo-configuration (i.e., 5α) in sea lamprey olfaction. This attribute could facilitate design of pheromone analogs to control this invasive species. PMID:21145335

New ΦBT1 site-specific integrative vectors with neutral phenotype in Streptomyces.

PubMed

Gonzalez-Quiñonez, Nathaly; López-García, María Teresa; Yagüe, Paula; Rioseras, Beatriz; Pisciotta, Annalisa; Alduina, Rosa; Manteca, Ángel

2016-03-01

Integrative plasmids are one of the best options to introduce genes in low copy and in a stable form into bacteria. The ΦC31-derived plasmids constitute the most common integrative vectors used in Streptomyces. They integrate at different positions (attB and pseudo-attB sites) generating different mutations. The less common ΦBT1-derived vectors integrate at the unique attB site localized in the SCO4848 gene (S. coelicolor genome) or their orthologues in other streptomycetes. This work demonstrates that disruption of SCO4848 generates a delay in spore germination. SCO4848 is co-transcribed with SCO4849, and the spore germination phenotype is complemented by SCO4849. Plasmids pNG1-4 were created by modifying the ΦBT1 integrative vector pMS82 by introducing a copy of SCO4849 under the control of the promoter region of SCO4848. pNG2 and pNG4 also included a copy of the P ermE * in order to facilitate gene overexpression. pNG3 and pNG4 harboured a copy of the bla gene (ampicillin resistance) to facilitate selection in E. coli. pNG1-4 are the only integrative vectors designed to produce a neutral phenotype when they are integrated into the Streptomyces genome. The experimental approach developed in this work can be applied to create phenotypically neutral integrative plasmids in other bacteria.

Detection analysis of surface hydroxyl active sites and simulation calculation of the surface dissociation constants of aqueous diatomite suspensions

NASA Astrophysics Data System (ADS)

Ma, Shu-Cui; Wang, Zhi-Gang; Zhang, Ji-Lin; Sun, De-Hui; Liu, Gui-Xia

2015-02-01

The surface properties of the diatomite were investigated using nitrogen adsorption/deadsorption isotherms, TG-DSC, FTIR, and XPS, and surface protonation-deprotonation behavior was determined by continuous acid-base potentiometric titration technique. The diatomite sample with porous honeycomb structure has a BET specific surface area of 10.21 m2/g and large numbers of surface hydroxyl functional groups (i.e. tbnd Si-OH, tbnd Fe-OH, and tbnd Al-OH). These surface hydroxyls can be protonated or deprotonated depending on the pH of the suspension. The experimental potentiometric data in two different ionic strength solutions (0.1 and 0.05 mol/L NaCl) were fitted using ProtoFit GUI V2.1 program by applying diffuse double layer model (DLM) with three amphoteric sites and minimizing the sum of squares between a dataset derivative function and a model derivative function. The optimized surface parameters (i.e. surface dissociation constants (log K1, log K2) and surface site concentrations (log C)) of the sample were obtained. Based on the optimized surface parameters, the surface species distribution was calculated using Program-free PHREEQC 3.1.2. Thus, this work reveals considerable new information about surface protonation-deprotonation processes and surface adsorptive behaviors of the diatomite, which helps us to effectively use the cheap and cheerful diatomite clay adsorbent.

Coupled motions in the SH2 and kinase domains of Csk control Src phosphorylation.

PubMed

Wong, Lilly; Lieser, Scot A; Miyashita, Osamu; Miller, Meghan; Tasken, Kjetil; Onuchic, Josè N; Adams, Joseph A; Woods, Virgil L; Jennings, Patricia A

2005-08-05

The C-terminal Src kinase (Csk) phosphorylates and down-regulates Src family tyrosine kinases. The Csk-binding protein (Cbp) localizes Csk close to its substrates at the plasma membrane, and increases the specific activity of the kinase. To investigate this long-range catalytic effect, the phosphorylation of Src and the conformation of Csk were investigated in the presence of a high-affinity phosphopeptide derived from Cbp. This peptide binds tightly to the SH2 domain and enhances Src recognition (lowers K(m)) by increasing the apparent phosphoryl transfer rate in the Csk active site, a phenomenon detected in rapid quench flow experiments. Previous studies demonstrated that the regulation of Csk activity is linked to conformational changes in the enzyme that can be probed with hydrogen-deuterium exchange methods. We show that the Cbp peptide impacts deuterium incorporation into its binding partner (the SH2 domain), and into the SH2-kinase linker and several sequences in the kinase domain, including the glycine-rich loop in the active site. These findings, along with computational data from normal mode analyses, suggest that the SH2 domain moves in a cantilever fashion with respect to the small lobe of the kinase domain, ordering the active site for catalysis. The binding of a small Cbp-derived peptide to the SH2 domain of Csk modifies these motions, enhancing Src recognition.

Thyroid nodules in the population living around Semipalatinsk nuclear test site: possible implications for dose-response relationships study.

PubMed

Zhumadilov, Zhaxybay

2006-02-01

The risk of radiation-induced nodules is higher than the risk for radiation-induced cancer. Risk factors and specific modifiers of the dose-response relationship may vary among different populations and not be well recognized. Many thyroid studies have considered thyroid nodularity itself, but not specific morphological types of thyroid nodules. There are many specific types of thyroid nodules which follow a morphological classification of thyroid lesions, including some congenital and tumor-like conditions. Modern equipment and technique can help us to identify particular specific types of thyroid nodules. In this study we report some results of a clinically applicable approach to materials derived from three studies. From 1999 through 2002, we have screened 571 current residents from 4 exposed and 1 control village near the Semipalatinsk Nuclear Test Site area, who were of similar ages (<20) at the time of major radiation fallout events at the SNTS. Prevalent nodules were identified by ultrasound and fine-needle aspiration biopsy, cytopathology results. Analysis of ultrasound images and cytopathology of thyroid lesions among exposed and non-exposed population allowed us to distinguish some interesting ultrasound features for specific types of thyroid nodules. We believe that it would be interesting and possibly more informative for thyroid dosimetry studies to consider specific morphological types of thyroid nodules. We need more detailed research to clarify the feasibility of applying these findings for study of the dose-response relationship.

Energy and greenhouse gas profiles of polyhydroxybutyrates derived from corn grain: a life cycle perspective.

PubMed

Kim, Seungdo; Dale, Bruce E

2008-10-15

Polyhydroxybutyrates (PHB) are well-known biopolymers derived from sugars orvegetable oils. Cradle-to-gate environmental performance of PHB derived from corn grain is evaluated through life cycle assessment (LCA), particularly nonrenewable energy consumption and greenhouse gas emissions. Site-specific process information on the corn wet milling and PHB fermentation and recovery processes was obtained from Telles. Most of energy used in the corn wet milling and PHB fermentation and recovery processes is generated in a cogeneration power plant in which corn stover, assumed to be representative of a variety of biomass sources that could be used, is burned to generate electricity and steam. County level agricultural information is used in estimating the environmental burdens associated with both corn grain and corn stover production. Results show that PHB derived from corn grain offers environmental advantages over petroleum-derived polymers in terms of nonrenewable energy consumption and greenhouse gas emissions. Furthermore, PHB provides greenhouse gas credits, and thus PHB use reduces greenhouse gas emissions compared to petroleum-derived polymers. Corn cultivation is one of the environmentally sensitive areas in the PHB production system. More sustainable practices in corn cultivation (e.g., using no-tillage and winter cover crops) could reduce the environmental impacts of PHB by up to 72%.

When and where does preferential flow matter - from observation to large scale modelling

NASA Astrophysics Data System (ADS)

Weiler, Markus; Leistert, Hannes; Steinbrich, Andreas

2017-04-01

Preferential flow can be of relevance in a wide range of soils and the interaction of different processes and factors are still difficult to assess. As most studies (including our own studies) focusing on the effect of preferential flow are based on relatively high precipitation rates, there is always the question how relevant preferential flow is under natural conditions, considering the site specific precipitation characteristics, the effect of the drying and wetting cycle on the initial soil water condition and shrinkage cracks, the site specific soil properties, soil structure and rock fragments, and the effect of plant roots and soil fauna (e.g. earthworm channels). In order to assess this question, we developed the distributed, process-based model RoGeR (Runoff Generation Research) to include a large number relevant features and processes of preferential flow in soils. The model was developed from a large number of process based research and experiments and includes preferential flow in roots, earthworm channels, along rock fragments and shrinkage cracks. We parameterized the uncalibrated model at a high spatial resolution of 5x5m for the whole state of Baden-Württemberg in Germany using LiDAR data, degree of sealing, landuse, soil properties and geology. As the model is an event based model, we derived typical event based precipitation characteristics based on rainfall duration, mean intensity and amount. Using the site-specific variability of initial soil moisture derived from a water balance model based on the same dataset, we simulated the infiltration and recharge amounts of all event classes derived from the event precipitation characteristics and initial soil moisture conditions. The analysis of the simulation results allowed us to extracts the relevance of preferential flow for infiltration and recharge considering all factors above. We could clearly see a strong effect of the soil properties and land-use, but also, particular for clay rich soils a strong effect of the initial conditions due to the development of soil cracks. Not too surprisingly, the relevance of preferential flow was much lower when considering the whole range of precipitation events as only considering events with a high rainfall intensity. Also, the influence on infiltration and recharge were different. Despite the model can still be improved in particular considering more realistic information about the spatial and temporal variability of preferential flow by soil fauna and plants, the model already shows under what situation we need to be very careful when predicting infiltration and recharge with models considering only longer time steps (daily) or only matrix flow.

Bathymetry (Part I), sedimentary regimes (Part II), and abyssal waste-disposal potential near the conterminous United States

NASA Astrophysics Data System (ADS)

Bowles, Frederick A.; Vogt, Peter R.; Jung, Woo-Yeol

1998-05-01

Placing waste on the seafloor, with the intention that it remain in place and isolated from mankind, requires a knowledge of the environmental factors that may be applicable to a specific seafloor area. DBDB5 (Digital Bathymetric Database gridded at 5' latitude by 5' longitude cell dimension) is used here for regional assessments of seafloor depth, slope, and relief at five surrogate abyssal waste sites; two each in the western Atlantic and eastern Pacific, and one in the Gulf of Mexico. Only Pacific-1 exhibits a `high' slope (2°) by DBDB5 standards, whereas the remaining sites are located on almost level seafloor. Detailed examination of the sites using multibeam-based contour sheets show the area around Atlantic-1 to be a featureless plain. Atlantic-2 and both Pacific sites are surrounded by abyssal hill topography, with local slopes ranging from greater than 6° at all sites to above 15° at Pacific-2. Neither Pacific site features a seafloor as `flat' as at Atlantic-1 or at the Gulf of Mexico site. Locating waste sites on sedimented slopes could have serious consequences due to catastrophic slope failure and downslope displacement of waste by mass sediment-transport processes. Neither slumping nor sliding are perceived as critical processes affecting the surrogate sites because of their locations on negligibly sloping seafloors. However, debris flows and turbidity currents are capable of transporting large volumes of sediment for long distances over low gradients and, in the case of turbidity currents, at great speed. Dispersal of loose waste material by these processes is virtually assured, but less likely if the waste is bagged. The turbidity current problem is alleviated (but not eliminated) by locating waste sites on distal portions of abyssal plains. Both Pacific sites are surrounded by abyssal hills and, in the case of Pacific-2, far beyond the reach of land-derived turbidity currents. Thin sediment cover and low rates of sedimentation have also resulted in highly stable slope (abyssal hill) deposits. Hence, the probability of locally derived, small-volume flows is low at these sites. Existing high sea levels have also resulted in a worldwide decrease in turbidity current activity relative to glacial times when sea levels were much lower.

NASA Remote Sensing Applications for Archaeology and Cultural Resources Management

NASA Technical Reports Server (NTRS)

Giardino, Marco J.

2008-01-01

NASA's Earth Science Mission Directorate recently completed the deployment of the Earth Observation System (EOS) which is a coordinated series of polar-orbiting and low inclination satellites for long-term global observations of the land surface, biosphere, solid Earth, atmosphere, and oceans. One of the many applications derived from EOS is the advancement of archaeological research and applications. Using satellites, manned and unmanned airborne platform, NASA scientists and their partners have conducted archaeological research using both active and passive sensors. The NASA Stennis Space Center (SSC) located in south Mississippi, near New Orleans, has been a leader in space archaeology since the mid-1970s. Remote sensing is useful in a wide range of archaeological research applications from landscape classification and predictive modeling to site discovery and mapping. Remote sensing technology and image analysis are currently undergoing a profound shift in emphasis from broad classification to detection, identification and condition of specific materials, both organic and inorganic. In the last few years, remote sensing platforms have grown increasingly capable and sophisticated. Sensors currently in use, including commercial instruments, offer significantly improved spatial and spectral resolutions. Paired with new techniques of image analysis, this technology provides for the direct detection of archaeological sites. As in all archaeological research, the application of remote sensing to archaeology requires a priori development of specific research designs and objectives. Initially targeted at broad archaeological issues, NASA space archaeology has progressed toward developing practical applications for cultural resources management (CRM). These efforts culminated with the Biloxi Workshop held by NASA and the University of Mississippi in 2002. The workshop and resulting publication specifically address the requirements of cultural resource managers through the use of remote sensing. In 2007, NASA awarded six competitively chosen projects in Space Archaeology through an open solicitation whose purpose, among several, was to addresses the potential benefits to modern society that can be derived through a better understanding of how past cultures succeeded or failed to adapt to local, regional, and global change. A further objective of NASA's space archaeology is the protection and preservation of cultural heritage sites while planning for the sustainable development of cultural resources. NASA s archaeological approach through remote sensing builds on traditional methods of aerial archaeology (i.e. crop marks) and utilizes advanced technologies for collecting and analyzing archaeological data from digital imagery. NASA s archaeological research and application projects using remote sensing have been conducted throughout the world. In North America, NASA has imaged prehistoric mound sites in Mississippi; prehistoric shell middens in Louisiana, Puebloan sites in New Mexico and more recently the sites associated with the Lewis and Clark Corps of Discovery Expedition (1804-1806). In Central America, NASA archaeologists have researched Mayan sites throughout the region, including the Yucatan and Costa Rica, as well as Olmec localities in Veracruz. Other data has been collected over Angkor, Cambodia, Giza in Egypt, the lost city of Ubar on the Arabian Peninsula.

Site-specific quantitative analysis of cardiac mitochondrial protein phosphorylation.

PubMed

Lam, Maggie P Y; Lau, Edward; Scruggs, Sarah B; Wang, Ding; Kim, Tae-Young; Liem, David A; Zhang, Jun; Ryan, Christopher M; Faull, Kym F; Ping, Peipei

2013-04-09

We report the development of a multiple-reaction monitoring (MRM) strategy specifically tailored to the detection and quantification of mitochondrial protein phosphorylation. We recently derived 68 MRM transitions specific to protein modifications in the respiratory chain, voltage-dependent anion channel, and adenine nucleotide translocase. Here, we have now expanded the total number of MRM transitions to 176 to cover proteins from the tricarboxylic acid cycle, pyruvate dehydrogenase complex, and branched-chain alpha-keto acid dehydrogenase complex. We utilized the transition set to analyze endogenous protein phosphorylation in human heart, mouse heart, and mouse liver. The data demonstrate the potential utility of the MRM workflow for studying the functional details of mitochondrial phosphorylation signaling. This article is part of a Special Issue entitled: From protein structures to clinical applications. Copyright © 2012 Elsevier B.V. All rights reserved.

Concentration-Dependent Multiple Binding Sites on Saliva-Treated Hydroxyapatite for Streptococcus sanguis

PubMed Central

Gibbons, R. J.; Moreno, E. C.; Etherden, I.

1983-01-01

The influence of bacterial cell concentration on estimates of the number of binding sites and the affinity for the adsorption of a strain of Streptococcus sanguis to saliva-treated hydroxyapatite was determined, and the possible presence of multiple binding sites for this organism was tested. The range of concentrations of available bacteria varied from 4.7 × 106 to 5,960 × 106 cells per ml. The numbers of adsorbed bacteria increased over the entire range tested, but a suggestion of a break in an otherwise smooth adsorption isotherm was evident. Values for the number of binding sites and the affinity varied considerably depending upon the range of available bacterial concentrations used to estimate them; high correlation coefficients were obtained in all cases. The use of low bacterial cell concentrations yielded lower values for the number of sites and much higher values for the affinity constant than did the use of high bacterial cell concentrations. When data covering the entire range of bacterial concentrations were employed, values for the number of sites and the affinity were similar to those obtained by using only high bacterial cell concentrations. The simplest explanation for these results is that there are multiple binding sites for S. sanguis on saliva-treated hydroxyapatite surfaces. When present in low concentration, the streptococci evidently attach to more specific high-affinity sites which become saturated when higher bacterial concentrations are employed. The possibility of multiple binding sites was substantiated by comparing estimates of the adsorption parameters from a computer-simulated isotherm with those derived from the experimentally generated isotherm. A mathematical model describing bacterial adsorption to binary binding sites was further evidence for the existence of at least two classes of binding sites for S. sanguis. Far fewer streptococci adsorbed to experimental pellicles prepared from saliva depleted of bacterial aggregating activity when low numbers of streptococci were used, but the magnitude of this difference was considerably less when high streptococcal concentrations were employed. This suggests an association between salivary components which possess bacterial-aggregating activity and bacterial adsorption to high-affinity specific binding sites on saliva-treated hydroxyapatite surfaces. PMID:6822416

Specific binding of nicergoline on an alpha1-like adrenoreceptor in the rat retina.

PubMed

Lograno, M D; Tricarico, D; Masciopinto, V; Scuderl, A C

2000-02-01

Systemic treatment with nicergoline, an ergoline derivative showing alpha1-antagonist properties, causes vasodilatation in the eye without apparent untoward cardiovascular effects. In the present work we investigated the ability of nicergoline to inhibit the binding of radiolabelled prazosin in the rat retina and cortex. We found that nicergoline inhibited [3H]prazosin binding in both tissues, being more potent than unlabelled prazosin in the retinal tissue. The competition curves of the ergoline derivative were well fitted by a one-site model in the cortical tissue, with an IC50 (concentration of the drugs needed to inhibit the binding of labelled prazosin by 50%) of 2.54 x 10(-8) M, and by a two-site model in the retinal tissue, with IC50 values of 7.08 x 10(-12) M and 1.82 x 10(-5) M. 2-(2,6 dimetoxyphenoxyethyl) aminomethyl-1,4-benzodioxane hydrochloride (WB4101) and phentolamine, selective ligands for the high-affinity binding site for prazosin, in particular the alpha1A-site, fully inhibited prazosin binding in the cortex but only partially inhibited prazosin binding in the retina, being less potent in this tissue than either nicergoline or prazosin. Our results suggest that a binding component of alpha1-adrenoreceptors is expressed to a lesser extent in the retina than the cortex, leading to a reduced response of the retinal tissue to prazosin, and more particularly to WB4101 and phentolamine. The selective binding of the nicergoline on this retinal adrenoreceptor may explain the peculiar efficacy of the drug in ocular pathophysiology.

Frataxin Directly Stimulates Mitochondrial Cysteine Desulfurase by Exposing Substrate-binding Sites, and a Mutant Fe-S Cluster Scaffold Protein with Frataxin-bypassing Ability Acts Similarly*♦

PubMed Central

Pandey, Alok; Gordon, Donna M.; Pain, Jayashree; Stemmler, Timothy L.; Dancis, Andrew; Pain, Debkumar

2013-01-01

For iron-sulfur (Fe-S) cluster synthesis in mitochondria, the sulfur is derived from the amino acid cysteine by the cysteine desulfurase activity of Nfs1. The enzyme binds the substrate cysteine in the pyridoxal phosphate-containing site, and a persulfide is formed on the active site cysteine in a manner depending on the accessory protein Isd11. The persulfide is then transferred to the scaffold Isu, where it combines with iron to form the Fe-S cluster intermediate. Frataxin is implicated in the process, although it is unclear where and how, and deficiency causes Friedreich ataxia. Using purified proteins and isolated mitochondria, we show here that the yeast frataxin homolog (Yfh1) directly and specifically stimulates cysteine binding to Nfs1 by exposing substrate-binding sites. This novel function of frataxin does not require iron, Isu1, or Isd11. Once bound to Nfs1, the substrate cysteine is the source of the Nfs1 persulfide, but this step is independent of frataxin and strictly dependent on Isd11. Recently, a point mutation in Isu1 was found to bypass many frataxin functions. The data presented here show that the Isu1 suppressor mimics the frataxin effects on Nfs1, explaining the bypassing activity. We propose a regulatory mechanism for the Nfs1 persulfide-forming activity. Specifically, at least two separate conformational changes must occur in the enzyme for optimum activity as follows: one is mediated by frataxin interaction that exposes the “buried” substrate-binding sites, and the other is mediated by Isd11 interaction that brings the bound substrate cysteine and the active site cysteine in proximity for persulfide formation. PMID:24217246

Tetrazepam: a benzodiazepine which dissociates sedation from other benzodiazepine activities. II. In vitro and in vivo interactions with benzodiazepine binding sites

DOE Office of Scientific and Technical Information (OSTI.GOV)

Keane, P.E.; Bachy, A.; Morre, M.

1988-05-01

Tetrazepam is a 1,4-benzodiazepine (BZD) derivative which, in rodents, appears to have very little sedative and ataxic effects. In an attempt to identify the molecular mechanisms underlying this particular pharmacological profile we examined the interaction of tetrazepam with BZD binding sites. Tetrazepam interacted competitively with central and peripheral BZD binding sites and exhibited comparable affinities for both sites. Tetrazepam was approximately one-seventh as potent as diazepam at the central receptor and as potent as diazepam at the peripheral binding site. Tetrazepam did not distinguish type I from type II central BZD receptors, as evidenced by comparable affinities for the cerebellarmore » and hippocampal receptors. In vitro autoradiographic studies showed that tetrazepam displaced (3H)flunitrazepam from rat brain membranes without any clear regional specificity. Like all BZD receptor agonists, tetrazepam exhibited a gamma-aminobutyric acid shift, a photoaffinity shift and potentiated the binding of 35S-t-butyl-bicyclophosphorothionate to rat brain membranes. However, the latter effect was observed at relatively high concentrations of tetrazepam. In vivo, tetrazepam displaced specifically bound (3H)flunitrazepam from mouse brain (ID50, 37 mg/kg p.o. vs 3.5 mg/kg p.o. for diazepam) and from mouse kidney (ID50, 38 mg/kg p.o. vs. 21 mg/kg p.o. for diazepam). It is concluded that tetrazepam is a BZD receptor agonist; the molecular mechanisms which underly the low sedative potential of the drug cannot at present be explained by a particular interaction with either central or peripheral BZD binding sites, but may be related to the drug's relatively weak effect on 35S-t-butyl-bicyclophosphorothionate binding.« less

Kinetics of heavy metal adsorption and desorption in soil: Developing a unified model based on chemical speciation

NASA Astrophysics Data System (ADS)

Peng, Lanfang; Liu, Paiyu; Feng, Xionghan; Wang, Zimeng; Cheng, Tao; Liang, Yuzhen; Lin, Zhang; Shi, Zhenqing

2018-03-01

Predicting the kinetics of heavy metal adsorption and desorption in soil requires consideration of multiple heterogeneous soil binding sites and variations of reaction chemistry conditions. Although chemical speciation models have been developed for predicting the equilibrium of metal adsorption on soil organic matter (SOM) and important mineral phases (e.g. Fe and Al (hydr)oxides), there is still a lack of modeling tools for predicting the kinetics of metal adsorption and desorption reactions in soil. In this study, we developed a unified model for the kinetics of heavy metal adsorption and desorption in soil based on the equilibrium models WHAM 7 and CD-MUSIC, which specifically consider metal kinetic reactions with multiple binding sites of SOM and soil minerals simultaneously. For each specific binding site, metal adsorption and desorption rate coefficients were constrained by the local equilibrium partition coefficients predicted by WHAM 7 or CD-MUSIC, and, for each metal, the desorption rate coefficients of various binding sites were constrained by their metal binding constants with those sites. The model had only one fitting parameter for each soil binding phase, and all other parameters were derived from WHAM 7 and CD-MUSIC. A stirred-flow method was used to study the kinetics of Cd, Cu, Ni, Pb, and Zn adsorption and desorption in multiple soils under various pH and metal concentrations, and the model successfully reproduced most of the kinetic data. We quantitatively elucidated the significance of different soil components and important soil binding sites during the adsorption and desorption kinetic processes. Our model has provided a theoretical framework to predict metal adsorption and desorption kinetics, which can be further used to predict the dynamic behavior of heavy metals in soil under various natural conditions by coupling other important soil processes.

Frataxin directly stimulates mitochondrial cysteine desulfurase by exposing substrate-binding sites, and a mutant Fe-S cluster scaffold protein with frataxin-bypassing ability acts similarly.

PubMed

Pandey, Alok; Gordon, Donna M; Pain, Jayashree; Stemmler, Timothy L; Dancis, Andrew; Pain, Debkumar

2013-12-27

For iron-sulfur (Fe-S) cluster synthesis in mitochondria, the sulfur is derived from the amino acid cysteine by the cysteine desulfurase activity of Nfs1. The enzyme binds the substrate cysteine in the pyridoxal phosphate-containing site, and a persulfide is formed on the active site cysteine in a manner depending on the accessory protein Isd11. The persulfide is then transferred to the scaffold Isu, where it combines with iron to form the Fe-S cluster intermediate. Frataxin is implicated in the process, although it is unclear where and how, and deficiency causes Friedreich ataxia. Using purified proteins and isolated mitochondria, we show here that the yeast frataxin homolog (Yfh1) directly and specifically stimulates cysteine binding to Nfs1 by exposing substrate-binding sites. This novel function of frataxin does not require iron, Isu1, or Isd11. Once bound to Nfs1, the substrate cysteine is the source of the Nfs1 persulfide, but this step is independent of frataxin and strictly dependent on Isd11. Recently, a point mutation in Isu1 was found to bypass many frataxin functions. The data presented here show that the Isu1 suppressor mimics the frataxin effects on Nfs1, explaining the bypassing activity. We propose a regulatory mechanism for the Nfs1 persulfide-forming activity. Specifically, at least two separate conformational changes must occur in the enzyme for optimum activity as follows: one is mediated by frataxin interaction that exposes the "buried" substrate-binding sites, and the other is mediated by Isd11 interaction that brings the bound substrate cysteine and the active site cysteine in proximity for persulfide formation.

Evolution of divergent DNA recognition specificities in VDE homing endonucleases from two yeast species

PubMed Central

Posey, Karen L.; Koufopanou, Vassiliki; Burt, Austin; Gimble, Frederick S.

2004-01-01

Homing endonuclease genes (HEGs) are mobile DNA elements that are thought to confer no benefit to their host. They encode site-specific DNA endonucleases that perpetuate the element within a species population by homing and disseminate it between species by horizontal transfer. Several yeast species contain the VMA1 HEG that encodes the intein-associated VMA1-derived endonuclease (VDE). The evolutionary state of VDEs from 12 species was assessed by assaying their endonuclease activities. Only two enzymes are active, PI-ZbaI from Zygosaccharomyces bailii and PI-ScaI from Saccharomyces cariocanus. PI-ZbaI cleaves the Z.bailii recognition sequence significantly faster than the Saccharomyces cerevisiae site, which differs at six nucleotide positions. A mutational analysis indicates that PI-ZbaI cleaves the S.cerevisiae substrate poorly due to the absence of a contact that is analogous to one made in PI-SceI between Gln-55 and nucleotides +9/+10. PI-ZbaI cleaves the Z.bailii substrate primarily due to a single base-pair substitution (A/T+5 → T/A+5). Structural modeling of the PI-ZbaI/DNA complex suggests that Arg-331, which is absent in PI-SceI, contacts T/A+5, and the reduced activity observed in a PI-ZbaI R331A mutant provides evidence for this interaction. These data illustrate that homing endonucleases evolve altered specificity as they adapt to recognize alternative target sites. PMID:15280510

Evolution of divergent DNA recognition specificities in VDE homing endonucleases from two yeast species.

PubMed

Posey, Karen L; Koufopanou, Vassiliki; Burt, Austin; Gimble, Frederick S

2004-01-01

Homing endonuclease genes (HEGs) are mobile DNA elements that are thought to confer no benefit to their host. They encode site-specific DNA endonucleases that perpetuate the element within a species population by homing and disseminate it between species by horizontal transfer. Several yeast species contain the VMA1 HEG that encodes the intein-associated VMA1-derived endonuclease (VDE). The evolutionary state of VDEs from 12 species was assessed by assaying their endonuclease activities. Only two enzymes are active, PI-ZbaI from Zygosaccharomyces bailii and PI-ScaI from Saccharomyces cariocanus. PI-ZbaI cleaves the Z.bailii recognition sequence significantly faster than the Saccharomyces cerevisiae site, which differs at six nucleotide positions. A mutational analysis indicates that PI-ZbaI cleaves the S.cerevisiae substrate poorly due to the absence of a contact that is analogous to one made in PI-SceI between Gln-55 and nucleotides +9/+10. PI-ZbaI cleaves the Z.bailii substrate primarily due to a single base-pair substitution (A/T+5 --> T/A+5). Structural modeling of the PI-ZbaI/DNA complex suggests that Arg-331, which is absent in PI-SceI, contacts T/A+5, and the reduced activity observed in a PI-ZbaI R331A mutant provides evidence for this interaction. These data illustrate that homing endonucleases evolve altered specificity as they adapt to recognize alternative target sites.

Molecular mechanisms of substrate recognition and specificity of botulinum neurotoxin serotype F.

PubMed

Chen, Sheng; Wan, Hoi Ying

2011-01-15

BoNTs (botulinum neurotoxins) are both deadly neurotoxins and natural toxins that are widely used in protein therapies to treat numerous neurological disorders of dystonia and spinal spasticity. Understanding the mechanism of action and substrate specificity of BoNTs is a prerequisite to develop antitoxin and novel BoNT-derived protein therapy. To date, there is a lack of detailed information with regard to how BoNTs recognize and hydrolyse the substrate VAMP-2 (vesicle-associated membrane protein 2), even though it is known to be cleaved by four of the seven BoNT serotypes, B, D, F, G and TeNT (tetanus neurotoxin). In the present study we dissected the molecular mechanisms of VAMP-2 recognition by BoNT serotype F for the first time. The initial substrate recognition was mediated through sequential binding of VAMP-2 to the B1, B2 and B3 pockets in LC/F (light chain of BoNT serotype F), which directed VAMP-2 to the active site of LC/F and stabilized the active site substrate recognition, where the P2, P1' and P2' sites of VAMP-2 were specifically recognized by the S2, S1' and S2' pockets of LC/F to promote substrate hydrolysis. The understanding of the molecular mechanisms of LC/F substrate recognition provides insights into the development of antitoxins and engineering novel BoNTs to optimize current therapy and extend therapeutic interventions.

Global Velocities from VLBI

NASA Technical Reports Server (NTRS)

Ma, Chopo; Gordon, David; MacMillan, Daniel

1999-01-01

Precise geodetic Very Long Baseline Interferometry (VLBI) measurements have been made since 1979 at about 130 points on all major tectonic plates, including stable interiors and deformation zones. From the data set of about 2900 observing sessions and about 2.3 million observations, useful three-dimensional velocities can be derived for about 80 sites using an incremental least-squares adjustment of terrestrial, celestial, Earth rotation and site/session-specific parameters. The long history and high precision of the data yield formal errors for horizontal velocity as low as 0.1 mm/yr, but the limitation on the interpretation of individual site velocities is the tie to the terrestrial reference frame. Our studies indicate that the effect of converting precise relative VLBI velocities to individual site velocities is an error floor of about 0.4 mm/yr. Most VLBI horizontal velocities in stable plate interiors agree with the NUVEL-1A model, but there are significant departures in Africa and the Pacific. Vertical precision is worse by a factor of 2-3, and there are significant non-zero values that can be interpreted as post-glacial rebound, regional effects, and local disturbances.

Errant processing and structural alterations of genomes present in a varicella-zoster virus vaccine.

PubMed Central

Vlazny, D A; Hyman, R W

1985-01-01

Five minority populations of aberrant, varicella-zoster virus (VZV)-derived genomes were identified among the encapsidated DNAs obtained from the nuclear and cytoplasmic fractions of an in vitro infection initiated with a lyophilized sample of the BIKEN VZV vaccine (strain Oka). These were (i) VZV genomes, present within nuclear but not cytoplasmic viral capsids, which had been cleaved at a specific site within the short segment and which were, therefore, 3.15 megadaltons (approximately 4% of the VZV genome length) short of full length; (ii) highly deleted, repetitive VZV genomes which contained the errant cleavage site but not the usual VZV genome terminal sequences; (iii) VZV genomes into which multiples of 1 through 5 defective genome repeat units had been inserted into a homologous site; (iv) VZV genomes with additions of 0.1 or 0.18 megadaltons of DNA at both the terminal and internal ends of the short segment; and (v) VZV DNA which had lost the HindIII restriction site at map position 0.11. Images PMID:2993670

The liver but not the skin is the site for conversion of a red carotenoid in a passerine bird

NASA Astrophysics Data System (ADS)

Del Val, Esther; Senar, Juan Carlos; Garrido-Fernández, Juan; Jarén, Manuel; Borràs, Antoni; Cabrera, Josep; Negro, Juan José

2009-07-01

Carotenoids may provide numerous health benefits and are also responsible for the integumentary coloration of many bird species. Despite their importance, many aspects of their metabolism are still poorly known, and even basic issues such as the anatomical sites of conversion remain controversial. Recent studies suggest that the transformation of carotenoid pigments takes place directly in the follicles during feather growth, even though the liver has been previously recognised as a storing organ for these pigments with a certain potential for conversion. In this context, we analysed the carotenoid profile of plasma, liver, skin and feathers of male Common Crossbills ( Loxia curvirostra). Interestingly, the derivative feather pigment 3-hydroxy-echinenone was detected in the liver and in the bloodstream (i.e. the necessary vehicle to transport metabolites to colourful peripheral tissues). Our results demonstrate for the first time with empirical data that the liver may act as the main site for the synthesis of integumentary carotenoids. This finding contradicts previous assumptions and raises the question of possible inter-specific differences in the site of carotenoid conversion in birds.

Site specific oxidation of amino acid residues in rat lens γ-crystallin induced by low-dose γ-irradiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Ingu; Saito, Takeshi; Research Reactor Institute, Kyoto University, Kumatori, Osaka 590-0494

Although cataracts are a well-known age-related disease, the mechanism of their formation is not well understood. It is currently thought that eye lens proteins become abnormally aggregated, initially causing clumping that scatters the light and interferes with focusing on the retina, and ultimately resulting in a cataract. The abnormal aggregation of lens proteins is considered to be triggered by various post-translational modifications, such as oxidation, deamidation, truncation and isomerization, that occur during the aging process. Such modifications, which are also generated by free radical and reactive oxygen species derived from γ-irradiation, decrease crystallin solubility and lens transparency, and ultimately leadmore » to the development of a cataract. In this study, we irradiated young rat lenses with low-dose γ-rays and extracted the water-soluble and insoluble protein fractions. The water-soluble and water-insoluble lens proteins were digested with trypsin, and the resulting peptides were analyzed by LC-MS. Specific oxidation sites of methionine, cysteine and tryptophan in rat water-soluble and -insoluble γE and γF-crystallin were determined by one-shot analysis. The oxidation sites in rat γE and γF-crystallin resemble those previously identified in γC and γD-crystallin from human age-related cataracts. Our study on modifications of crystallins induced by ionizing irradiation may provide useful information relevant to human senile cataract formation. - Highlights: • Low-dose γ-rays induced oxidation at specific residues in γE- and γF-crystallin. • The number of oxidation sites was higher in insoluble than soluble crystallins. • γ-Irradiation closely mimics the oxidation that occur in senile human cataracts.« less

Tropospheric delay ray tracing applied in VLBI analysis

NASA Astrophysics Data System (ADS)

Eriksson, David; MacMillan, D. S.; Gipson, John M.

2014-12-01

Tropospheric delay modeling error continues to be one of the largest sources of error in VLBI (very long baseline interferometry) analysis. For standard operational solutions, we use the VMF1 elevation-dependent mapping functions derived from European Centre for Medium-Range Weather Forecasts data. These mapping functions assume that tropospheric delay at a site is azimuthally symmetric. As this assumption is not true, we have instead determined the ray trace delay along the signal path through the troposphere for each VLBI quasar observation. We determined the troposphere refractivity fields from the pressure, temperature, specific humidity, and geopotential height fields of the NASA Goddard Space Flight Center Goddard Earth Observing System version 5 numerical weather model. When applied in VLBI analysis, baseline length repeatabilities were improved compared with using the VMF1 mapping function model for 72% of the baselines and site vertical repeatabilities were better for 11 of 13 sites during the 2 week CONT11 observing period in September 2011. When applied to a larger data set (2011-2013), we see a similar improvement in baseline length and also in site position repeatabilities for about two thirds of the stations in each of the site topocentric components.

Characteristics of the early immune response following transplantation of mouse ES cell derived insulin-producing cell clusters.

PubMed

Boyd, Ashleigh S; Wood, Kathryn J

2010-06-04

The fully differentiated progeny of ES cells (ESC) may eventually be used for cell replacement therapy (CRT). However, elements of the innate immune system may contribute to damage or destruction of these tissues when transplanted. Herein, we assessed the hitherto ill-defined contribution of the early innate immune response in CRT after transplantation of either ESC derived insulin producing cell clusters (IPCCs) or adult pancreatic islets. Ingress of neutrophil or macrophage cells was noted immediately at the site of IPCC transplantation, but this infiltration was attenuated by day three. Gene profiling identified specific inflammatory cytokines and chemokines that were either absent or sharply reduced by three days after IPCC transplantation. Thus, IPCC transplantation provoked less of an early immune response than pancreatic islet transplantation. Our study offers insights into the characteristics of the immune response of an ESC derived tissue in the incipient stages following transplantation and suggests potential strategies to inhibit cell damage to ensure their long-term perpetuation and functionality in CRT.

Synthesis, alkaline phosphatase inhibition studies and molecular docking of novel derivatives of 4-quinolones.

PubMed

Miliutina, Mariia; Ejaz, Syeda Abida; Khan, Shafi Ullah; Iaroshenko, Viktor O; Villinger, Alexander; Iqbal, Jamshed; Langer, Peter

2017-01-27

New and convenient methods for the functionalization of the 4-quinolone scaffold at positions C-1, C-3 and C-6 were developed. The 4-quinolone derivatives were evaluated for their inhibitory potential on alkaline phosphatase isozymes. Most of the compounds exhibit excellent inhibitory activity and moderate selectivity. The IC 50 values on tissue non-specific alkaline phosphatase (TNAP) were in the range of 1.34 ± 0.11 to 44.80 ± 2.34 μM, while the values on intestinal alkaline phosphatase (IAP) were in the range of 1.06 ± 0.32 to 192.10 ± 3.78 μM. The most active derivative exhibits a potent inhibition on IAP with a ≈14 fold higher selectivity as compared to TNAP. Furthermore, molecular docking calculations were performed for the most potent inhibitors to show their binding interactions within the active site of the respective enzymes. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Decellularized Tissue and Cell-Derived Extracellular Matrices as Scaffolds for Orthopaedic Tissue Engineering

PubMed Central

Cheng, Christina W.; Solorio, Loran D.; Alsberg, Eben

2014-01-01

The reconstruction of musculoskeletal defects is a constant challenge for orthopaedic surgeons. Musculoskeletal injuries such as fractures, chondral lesions, infections and tumor debulking can often lead to large tissue voids requiring reconstruction with tissue grafts. Autografts are currently the gold standard in orthopaedic tissue reconstruction; however, there is a limit to the amount of tissue that can be harvested before compromising the donor site. Tissue engineering strategies using allogeneic or xenogeneic decellularized bone, cartilage, skeletal muscle, tendon and ligament have emerged as promising potential alternative treatment. The extracellular matrix provides a natural scaffold for cell attachment, proliferation and differentiation. Decellularization of in vitro cell-derived matrices can also enable the generation of autologous constructs from tissue specific cells or progenitor cells. Although decellularized bone tissue is widely used clinically in orthopaedic applications, the exciting potential of decellularized cartilage, skeletal muscle, tendon and ligament cell-derived matrices has only recently begun to be explored for ultimate translation to the orthopaedic clinic. PMID:24417915

Production of Mutated Porcine Embryos Using Zinc Finger Nucleases and a Reporter-based Cell Enrichment System.

PubMed

Koo, Ok Jae; Park, Sol Ji; Lee, Choongil; Kang, Jung Taek; Kim, Sujin; Moon, Joon Ho; Choi, Ji Yei; Kim, Hyojin; Jang, Goo; Kim, Jin-Soo; Kim, Seokjoong; Lee, Byeong-Chun

2014-03-01

To facilitate the construction of genetically-modified pigs, we produced cloned embryos derived from porcine fibroblasts transfected with a pair of engineered zinc finger nuclease (ZFN) plasmids to create targeted mutations and enriched using a reporter plasmid system. The reporter expresses RFP and eGFP simultaneously when ZFN-mediated site-specific mutations occur. Thus, double positive cells (RFP(+)/eGFP(+)) were selected and used for somatic cell nuclear transfer. Two types of reporter based enrichment systems were used in this study; the cloned embryos derived from cells enriched using a magnetic sorting-based system showed better developmental competence than did those derived from cells enriched by flow cytometry. Mutated sequences, such as insertions, deletions, or substitutions, together with the wild-type sequence, were found in the cloned porcine blastocysts. Therefore, genetic mutations can be achieved in cloned porcine embryos reconstructed with ZFN-treated cells that were enriched by a reporter-based system.

Solution NMR studies provide structural basis for endotoxin pattern recognition by the innate immune receptor CD14

DOE Office of Scientific and Technical Information (OSTI.GOV)

Albright, Seth; Chen Bin; Holbrook, Kristen

CD14 functions as a key pattern recognition receptor for a diverse array of Gram-negative and Gram-positive cell-wall components in the host innate immune response by binding to pathogen-associated molecular patterns (PAMPs) at partially overlapping binding site(s). To determine the potential contribution of CD14 residues in this pattern recognition, we have examined using solution NMR spectroscopy, the binding of three different endotoxin ligands, lipopolysaccharide, lipoteichoic acid, and a PGN-derived compound, muramyl dipeptide to a {sup 15}N isotopically labeled 152-residue N-terminal fragment of sCD14 expressed in Pichia pastoris. Mapping of NMR spectral changes upon addition of ligands revealed that the pattern ofmore » residues affected by binding of each ligand is partially similar and partially different. This first direct structural observation of the ability of specific residue combinations of CD14 to differentially affect endotoxin binding may help explain the broad specificity of CD14 in ligand recognition and provide a structural basis for pattern recognition. Another interesting finding from the observed spectral changes is that the mode of binding may be dynamically modulated and could provide a mechanism for binding endotoxins with structural diversity through a common binding site.« less

N- and O-Glycosylation in the Murine Synaptosome*

PubMed Central

Trinidad, Jonathan C.; Schoepfer, Ralf; Burlingame, Alma L.; Medzihradszky, Katalin F.

2013-01-01

We present the first large scale study characterizing both N- and O-linked glycosylation in a site-specific manner on hundreds of proteins. We demonstrate that a lectin-affinity fractionation step using wheat germ agglutinin enriches not only peptides carrying intracellular O-GlcNAc, but also those bearing ER/Golgi-derived N- and O-linked carbohydrate structures. Liquid chromatography-MS (LC/MS) analysis with high accuracy precursor mass measurements and high sensitivity ion trap electron-transfer dissociation (ETD) were utilized for structural characterization of glycopeptides. Our results reveal both the identity of the precise sites of glycosylation and information on the oligosaccharide structures possible on these proteins. We report a novel iterative approach that allowed us to interpret the ETD data set directly without making prior assumptions about the nature and distribution of oligosaccharides present in our glycopeptide mixture. Over 2500 unique N- and O-linked glycopeptides were identified on 453 proteins. The extent of microheterogeneity varied extensively, and up to 19 different oligosaccharides were attached at a given site. We describe the presence of the well-known mucin-type structures for O-glycosylation, an EGF-domain-specific fucosylation and a rare O-mannosylation on the transmembrane phosphatase Ptprz1. Finally, we identified three examples of O-glycosylation on tyrosine residues. PMID:23816992

N- and O-glycosylation in the murine synaptosome.

PubMed

Trinidad, Jonathan C; Schoepfer, Ralf; Burlingame, Alma L; Medzihradszky, Katalin F

2013-12-01

We present the first large scale study characterizing both N- and O-linked glycosylation in a site-specific manner on hundreds of proteins. We demonstrate that a lectin-affinity fractionation step using wheat germ agglutinin enriches not only peptides carrying intracellular O-GlcNAc, but also those bearing ER/Golgi-derived N- and O-linked carbohydrate structures. Liquid chromatography-MS (LC/MS) analysis with high accuracy precursor mass measurements and high sensitivity ion trap electron-transfer dissociation (ETD) were utilized for structural characterization of glycopeptides. Our results reveal both the identity of the precise sites of glycosylation and information on the oligosaccharide structures possible on these proteins. We report a novel iterative approach that allowed us to interpret the ETD data set directly without making prior assumptions about the nature and distribution of oligosaccharides present in our glycopeptide mixture. Over 2500 unique N- and O-linked glycopeptides were identified on 453 proteins. The extent of microheterogeneity varied extensively, and up to 19 different oligosaccharides were attached at a given site. We describe the presence of the well-known mucin-type structures for O-glycosylation, an EGF-domain-specific fucosylation and a rare O-mannosylation on the transmembrane phosphatase Ptprz1. Finally, we identified three examples of O-glycosylation on tyrosine residues.

Structure, Function, and Evolution of Biogenic Amine-binding Proteins in Soft Ticks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mans, Ben J.; Ribeiro, Jose M.C.; Andersen, John F.

2008-08-19

Two highly abundant lipocalins, monomine and monotonin, have been isolated from the salivary gland of the soft tick Argas monolakensis and shown to bind histamine and 5-hydroxytryptamine (5-HT), respectively. The crystal structures of monomine and a paralog of monotonin were determined in the presence of ligands to compare the determinants of ligand binding. Both the structures and binding measurements indicate that the proteins have a single binding site rather than the two sites previously described for the female-specific histamine-binding protein (FS-HBP), the histamine-binding lipocalin of the tick Rhipicephalus appendiculatus. The binding sites of monomine and monotonin are similar to themore » lower, low affinity site of FS-HBP. The interaction of the protein with the aliphatic amine group of the ligand is very similar for the all of the proteins, whereas specificity is determined by interactions with the aromatic portion of the ligand. Interestingly, protein interaction with the imidazole ring of histamine differs significantly between the low affinity binding site of FS-HBP and monomine, suggesting that histamine binding has evolved independently in the two lineages. From the conserved features of these proteins, a tick lipocalin biogenic amine-binding motif could be derived that was used to predict biogenic amine-binding function in other tick lipocalins. Heterologous expression of genes from salivary gland libraries led to the discovery of biogenic amine-binding proteins in soft (Ornithodoros) and hard (Ixodes) tick genera. The data generated were used to reconstruct the most probable evolutionary pathway for the evolution of biogenic amine-binding in tick lipocalins.« less

Skeletal dosimetry for external exposure to photons based on µCT images of spongiosa from different bone sites

NASA Astrophysics Data System (ADS)

Kramer, R.; Khoury, H. J.; Vieira, J. W.; Kawrakow, I.

2007-11-01

Micro computed tomography (µCT) images of human spongiosa have recently been used for skeletal dosimetry with respect to external exposure to photon radiation. In this previous investigation, the calculation of equivalent dose to the red bone marrow (RBM) and to the bone surface cells (BSC) was based on five different clusters of micro matrices derived from µCT images of vertebrae, and the BSC equivalent dose for 10 µm thickness of the BSC layer was determined using an extrapolation method. The purpose of this study is to extend the earlier investigation by using µCT images from eight different bone sites and by introducing an algorithm for the direct calculation of the BSC equivalent dose with sub-micro voxel resolution. The results show that for given trabecular bone volume fractions (TBVFs) the whole-body RBM equivalent dose does not depend on bone site-specific properties or imaging parameters. However, this study demonstrates that apart from the TBVF and the BSC layer thickness, the BSC equivalent dose additionally depends on a so-called trabecular bone structure (TBS) effect, i.e. that the contribution of photo-electrons released in trabecular bone to the BSC equivalent dose also depends on the bone site-specific structure of the trabeculae. For a given bone site, the TBS effect is also a function of the thickness of the BSC layer, and it could be shown that this effect would disappear almost completely, should the BSC layer thickness be raised from 10 to 50 µm, according to new radiobiological findings.

Assessing the impact of anthropogenic pollution on isoprene-derived secondary organic aerosol formation in PM2.5 collected from the Birmingham, Alabama, ground site during the 2013 Southern Oxidant and Aerosol Study

NASA Astrophysics Data System (ADS)

Rattanavaraha, Weruka; Chu, Kevin; Hapsari Budisulistiorini, Sri; Riva, Matthieu; Lin, Ying-Hsuan; Edgerton, Eric S.; Baumann, Karsten; Shaw, Stephanie L.; Guo, Hongyu; King, Laura; Weber, Rodney J.; Neff, Miranda E.; Stone, Elizabeth A.; Offenberg, John H.; Zhang, Zhenfa; Gold, Avram; Surratt, Jason D.

2016-04-01

In the southeastern US, substantial emissions of isoprene from deciduous trees undergo atmospheric oxidation to form secondary organic aerosol (SOA) that contributes to fine particulate matter (PM2.5). Laboratory studies have revealed that anthropogenic pollutants, such as sulfur dioxide (SO2), oxides of nitrogen (NOx), and aerosol acidity, can enhance SOA formation from the hydroxyl radical (OH)-initiated oxidation of isoprene; however, the mechanisms by which specific pollutants enhance isoprene SOA in ambient PM2.5 remain unclear. As one aspect of an investigation to examine how anthropogenic pollutants influence isoprene-derived SOA formation, high-volume PM2.5 filter samples were collected at the Birmingham, Alabama (BHM), ground site during the 2013 Southern Oxidant and Aerosol Study (SOAS). Sample extracts were analyzed by gas chromatography-electron ionization-mass spectrometry (GC/EI-MS) with prior trimethylsilylation and ultra performance liquid chromatography coupled to electrospray ionization high-resolution quadrupole time-of-flight mass spectrometry (UPLC/ESI-HR-QTOFMS) to identify known isoprene SOA tracers. Tracers quantified using both surrogate and authentic standards were compared with collocated gas- and particle-phase data as well as meteorological data provided by the Southeastern Aerosol Research and Characterization (SEARCH) network to assess the impact of anthropogenic pollution on isoprene-derived SOA formation. Results of this study reveal that isoprene-derived SOA tracers contribute a substantial mass fraction of organic matter (OM) ( ˜ 7 to ˜ 20 %). Isoprene-derived SOA tracers correlated with sulfate (SO42-) (r2 = 0.34, n = 117) but not with NOx. Moderate correlations between methacrylic acid epoxide and hydroxymethyl-methyl-α-lactone (together abbreviated MAE/HMML)-derived SOA tracers with nitrate radical production (P[NO3]) (r2 = 0.57, n = 40) were observed during nighttime, suggesting a potential role of the NO3 radical in forming this SOA type. However, the nighttime correlation of these tracers with nitrogen dioxide (NO2) (r2 = 0.26, n = 40) was weaker. Ozone (O3) correlated strongly with MAE/HMML-derived tracers (r2 = 0.72, n = 30) and moderately with 2-methyltetrols (r2 = 0.34, n = 15) during daytime only, suggesting that a fraction of SOA formation could occur from isoprene ozonolysis in urban areas. No correlation was observed between aerosol pH and isoprene-derived SOA. Lack of correlation between aerosol acidity and isoprene-derived SOA is consistent with the observation that acidity is not a limiting factor for isoprene SOA formation at the BHM site as aerosols were acidic enough to promote multiphase chemistry of isoprene-derived epoxides throughout the duration of the study. All in all, these results confirm previous studies suggesting that anthropogenic pollutants enhance isoprene-derived SOA formation.

Prediction of citrullination sites by incorporating k-spaced amino acid pairs into Chou's general pseudo amino acid composition.

PubMed

Ju, Zhe; Wang, Shi-Yun

2018-04-22

As one of the most important and common protein post-translational modifications, citrullination plays a key role in regulating various biological processes and is associated with several human diseases. The accurate identification of citrullination sites is crucial for elucidating the underlying molecular mechanisms of citrullination and designing drugs for related human diseases. In this study, a novel bioinformatics tool named CKSAAP_CitrSite is developed for the prediction of citrullination sites. With the assistance of support vector machine algorithm, the highlight of CKSAAP_CitrSite is to adopt the composition of k-spaced amino acid pairs surrounding a query site as input. As illustrated by 10-fold cross-validation, CKSAAP_CitrSite achieves a satisfactory performance with a Sensitivity of 77.59%, a Specificity of 95.26%, an Accuracy of 89.37% and a Matthew's correlation coefficient of 0.7566, which is much better than those of the existing prediction method. Feature analysis shows that the N-terminal space containing pairs may play an important role in the prediction of citrullination sites, and the arginines close to N-terminus tend to be citrullinated. The conclusions derived from this study could offer useful information for elucidating the molecular mechanisms of citrullination and related experimental validations. A user-friendly web-server for CKSAAP_CitrSite is available at 123.206.31.171/CKSAAP_CitrSite/. Copyright © 2017. Published by Elsevier B.V.

Chlorophyll-Derivative Modulation of Rhodopsin Signaling Properties through Evolutionarily Conserved Interaction Pathways

PubMed Central

Woods, Kristina N.; Pfeffer, Jürgen; Klein-Seetharaman, Judith

2017-01-01

Retinal is the light-absorbing chromophore that is responsible for the activation of visual pigments and light-driven ion pumps. Evolutionary changes in the intermolecular interactions of the retinal with specific amino acids allow for adaptation of the spectral characteristics, referred to as spectral tuning. However, it has been proposed that a specific species of dragon fish has bypassed the adaptive evolutionary process of spectral tuning and replaced it with a single evolutionary event: photosensitization of rhodopsin by chlorophyll derivatives. Here, by using a combination of experimental measurements and computational modeling to probe retinal-receptor interactions in rhodopsin, we show how the binding of the chlorophyll derivative, chlorin-e6 (Ce6) in the intracellular domain (ICD) of the receptor allosterically excites G-protein coupled receptor class A (GPCR-A) conserved long-range correlated fluctuations that connect distant parts of the receptor. These long-range correlated motions are associated with regulating the dynamics and intermolecular interactions of specific amino acids in the retinal ligand-binding pocket that have been associated with shifts in the absorbance peak maximum (λmax) and hence, spectral sensitivity of the visual system. Moreover, the binding of Ce6 affects the overall global properties of the receptor. Specifically, we find that Ce6-induced dynamics alter the thermal stability of rhodopsin by adjusting hydrogen-bonding interactions near the receptor active-site that consequently also influences the intrinsic conformational equilibrium of the receptor. Due to the conservation of the ICD residues amongst different receptors in this class and the fact that all GPCR-A receptors share a common mechanism of activation, it is possible that the allosteric associations excited in rhodopsin with Ce6 binding are a common feature in all class A GPCRs. PMID:29312953

Development of DCGLs by using both probabilistic and deterministic analyses in RESRAD (onsite) and RESRAD-OFFSITE codes.

PubMed

Kamboj, Sunita; Yu, Charley; Johnson, Robert

2013-05-01

The Derived Concentration Guideline Levels for two building areas previously used in waste processing and storage at Argonne National Laboratory were developed using both probabilistic and deterministic radiological environmental pathway analysis. Four scenarios were considered. The two current uses considered were on-site industrial use and off-site residential use with farming. The two future uses (i.e., after an institutional control period of 100 y) were on-site recreational use and on-site residential use with farming. The RESRAD-OFFSITE code was used for the current-use off-site residential/farming scenario and RESRAD (onsite) was used for the other three scenarios. Contaminants of concern were identified from the past operations conducted in the buildings and the actual characterization done at the site. Derived Concentration Guideline Levels were developed for all four scenarios using deterministic and probabilistic approaches, which include both "peak-of-the-means" and "mean-of-the-peaks" analyses. The future-use on-site residential/farming scenario resulted in the most restrictive Derived Concentration Guideline Levels for most radionuclides.

Intestinal ABCA1 directly contributes to HDL biogenesis in vivo

PubMed Central

Brunham, Liam R.; Kruit, Janine K.; Iqbal, Jahangir; Fievet, Catherine; Timmins, Jenelle M.; Pape, Terry D.; Coburn, Bryan A.; Bissada, Nagat; Staels, Bart; Groen, Albert K.; Hussain, M. Mahmood; Parks, John S.; Kuipers, Folkert; Hayden, Michael R.

2006-01-01

Plasma HDL cholesterol levels are inversely related to risk for atherosclerosis. The ATP-binding cassette, subfamily A, member 1 (ABCA1) mediates the rate-controlling step in HDL particle formation, the assembly of free cholesterol and phospholipids with apoA-I. ABCA1 is expressed in many tissues; however, the physiological functions of ABCA1 in specific tissues and organs are still elusive. The liver is known to be the major source of plasma HDL, but it is likely that there are other important sites of HDL biogenesis. To assess the contribution of intestinal ABCA1 to plasma HDL levels in vivo, we generated mice that specifically lack ABCA1 in the intestine. Our results indicate that approximately 30% of the steady-state plasma HDL pool is contributed by intestinal ABCA1 in mice. In addition, our data suggest that HDL derived from intestinal ABCA1 is secreted directly into the circulation and that HDL in lymph is predominantly derived from the plasma compartment. These data establish a critical role for intestinal ABCA1 in plasma HDL biogenesis in vivo. PMID:16543947

Hydroxypyridinone Derivatives: A Fascinating Class of Chelators with Therapeutic Applications - An Update.

PubMed

Chaves, Sílvia; Piemontese, Luca; Hiremathad, Asha; Santos, Maria A

2018-01-01

Hydroxypyridinones (HPs) are a family of N-heterocyclic metal chelators, which have been an attractive target in the development of a variety of new pharmaceutical drugs, due to their high metal chelating efficacy/specificity and easy derivatization to tune the desired biological properties. In fact, along the last decades, hydroxypyridinone derivatives, but mostly 3-hydroxy-4-pyridinone (3,4-HP), have been intensively used in drug design, following either a multitarget approach, in which one chelating unity is extrafunctionalized (hybridized) to enable the interaction with other important specific biological sites, or a polydenticity approach, in which more than one chelating moiety is conveniently attached to one scaffold, to increase the metal chelating efficacy. This review represents an update of the most recent publications (2014-2016) in mono-HP hybrids, namely as potential anti-Alzheimer's drugs, inhibitors of metalloenzymes and anti-microbials, and also polychelating compounds (poly- HP), in view of potential application, such as anti-microbial/biostatic agents, luminescent biosensors or diagnostic agents. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Energy metabolism of synaptosomes from different neuronal systems of rat cerebellum during aging: a functional proteomic characterization.

PubMed

Ferrari, Federica; Gorini, Antonella; Villa, Roberto Federico

2015-01-01

Functional proteomics was used to characterize age-related changes in energy metabolism of different neuronal pathways within the cerebellar cortex of Wistar rats aged 2, 6, 12, 18, and 24 months. The "large" synaptosomes, derived from the glutamatergic mossy fibre endings which make synaptic contact with the granule cells of the granular layer, and the "small" synaptosomes, derived from the pre-synaptic terminals of granule cells making synaptic contact with the dendrites of Purkinje cells, were isolated by a combined differential/gradient centrifugation technique. Because most brain disorders are associated with bioenergetic changes, the maximum rate (Vmax) of selected enzymes of glycolysis, Krebs' cycle, glutamate and amino acids metabolism, and acetylcholine catabolism were evaluated. The results show that "large" and "small" synaptosomes possess specific and independent metabolic features. This study represents a reliable model to study in vivo (1) the physiopathological molecular mechanisms of some brain diseases dependent on energy metabolism, (2) the responsiveness to noxious stimuli, and (3) the effects of drugs, discriminating their action sites at subcellular level on specific neuronal pathways.

Onion-derived N, S self-doped carbon materials as highly efficient metal-free electrocatalysts for the oxygen reduction reaction

NASA Astrophysics Data System (ADS)

Yang, Shuting; Mao, Xinxin; Cao, Zhaoxia; Yin, Yanhong; Wang, Zhichao; Shi, Mengjiao; Dong, Hongyu

2018-01-01

Onion-derived nitrogen, sulfur self-doped nanoporous carbon spheres (NSC) as efficient metal-free electrocatalyst were synthesized via a facile hydrothermal and subsequent pyrolysis process. The typical NSC with a high BET specific surface area of 1558 m2 g-1, contains 6.23 at.% N and 0.36 at.% S, and possesses high concentration of pyridinic and graphitic nitrogen species. Experimentally, the best performance was the NSC-A2 which showed excellent catalytic activity to oxygen reduction reaction via a 4 electron mechanism with an onset potential of 0.88 V (vs. RHE), and a superior stability comparable to commercial Pt/C catalyst. The high electrocatalytic activity is attributed to not only the synergistic effect of N and S dual doping in carbon and the sufficient active sites, but also its high BET specific surface area and suitable microporous structure. The results demonstrate that it is a simple and scalable approach for preparing efficient and low-cost carbon-based electrocatalysts for oxygen reduction reaction.

Transgenic Citrus Expressing an Arabidopsis NPR1 Gene Exhibit Enhanced Resistance against Huanglongbing (HLB; Citrus Greening).

PubMed

Dutt, Manjul; Barthe, Gary; Irey, Michael; Grosser, Jude

2015-01-01

Commercial sweet orange cultivars lack resistance to Huanglongbing (HLB), a serious phloem limited bacterial disease that is usually fatal. In order to develop sustained disease resistance to HLB, transgenic sweet orange cultivars 'Hamlin' and 'Valencia' expressing an Arabidopsis thaliana NPR1 gene under the control of a constitutive CaMV 35S promoter or a phloem specific Arabidopsis SUC2 (AtSUC2) promoter were produced. Overexpression of AtNPR1 resulted in trees with normal phenotypes that exhibited enhanced resistance to HLB. Phloem specific expression of NPR1 was equally effective for enhancing disease resistance. Transgenic trees exhibited reduced diseased severity and a few lines remained disease-free even after 36 months of planting in a high-disease pressure field site. Expression of the NPR1 gene induced expression of several native genes involved in the plant defense signaling pathways. The AtNPR1 gene being plant derived can serve as a component for the development of an all plant T-DNA derived consumer friendly GM tree.

When seconds count: A study of communication variables in the opening segment of emergency calls.

PubMed

Penn, Claire; Koole, Tom; Nattrass, Rhona

2017-09-01

The opening sequence of an emergency call influences the efficiency of the ambulance dispatch time. The greeting sequences in 105 calls to a South African emergency service were analysed. Initial results suggested the advantage of a specific two-part opening sequence. An on-site experiment aimed at improving call efficiency was conducted during one shift (1100 calls). Results indicated reduced conversational repairs and a significant reduction of 4 seconds in mean call length. Implications for systems and training are derived.

Biogenesis of the multifunctional lipid droplet: Lipids, proteins, and sites

PubMed Central

Gross, Steven P.

2014-01-01

Lipid droplets (LDs) are ubiquitous dynamic organelles that store and supply lipids in all eukaryotic and some prokaryotic cells for energy metabolism, membrane synthesis, and production of essential lipid-derived molecules. Interest in the organelle’s cell biology has exponentially increased over the last decade due to the link between LDs and prevalent human diseases and the discovery of new and unexpected functions of LDs. As a result, there has been significant recent progress toward understanding where and how LDs are formed, and the specific lipid pathways that coordinate LD biogenesis. PMID:24590170

Status of Progress Made Toward Safety Analysis and Technical Site Evaluations for DOE Managed HLW and SNF.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sevougian, S. David; Stein, Emily; Gross, Michael B

The Spent Fuel and Waste Science and Technology (SFWST) Campaign of the U.S. Department of Energy (DOE) Office of Nuclear Energy (NE) is conducting research and development (R&D) on generic deep geologic disposal systems (i.e., repositories). This report describes specific activities in FY 2016 associated with the development of a Defense Waste Repository (DWR)a for the permanent disposal of a portion of the HLW and SNF derived from national defense and research and development (R&D) activities of the DOE.

Replacement of RNA hairpins by in vitro selected tetranucleotides.

PubMed Central

Dichtl, B; Pan, T; DiRenzo, A B; Uhlenbeck, O C

1993-01-01

An in vitro selection method based on the autolytic cleavage of yeast tRNA(Phe) by Pb2+ was applied to obtain tRNA derivatives with the anticodon hairpin replaced by four single-stranded nucleotides. Based on the rates of the site-specific cleavage by Pb2+ and the presence of a specific UV-induced crosslink, certain tetranucleotide sequences allow proper folding of the rest of the tRNA molecule, whereas others do not. One such successful tetramer sequence was also used to replace the acceptor stem of yeast tRNA(Phe) and the anticodon hairpin of E.coli tRNA(Phe) without disrupting folding. These experiments suggest that certain tetramers may be able to replace structurally nonessential hairpins in any RNA. Images PMID:7680121

Tissue-specific contribution of macrophages to wound healing.

PubMed

Minutti, Carlos M; Knipper, Johanna A; Allen, Judith E; Zaiss, Dietmar M W

2017-01-01

Macrophages are present in all tissues, either as resident cells or monocyte-derived cells that infiltrate into tissues. The tissue site largely determines the phenotype of tissue-resident cells, which help to maintain tissue homeostasis and act as sentinels of injury. Both tissue resident and recruited macrophages make a substantial contribution to wound healing following injury. In this review, we evaluate how macrophages in two fundamentally distinct tissues, i.e. the lung and the skin, differentially contribute to the process of wound healing. We highlight the commonalities of macrophage functions during repair and contrast them with distinct, tissue-specific functions that macrophages fulfill during the different stages of wound healing. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

Virtual Institute of Microbial Stress and Survival: Deduction of Stress Response Pathways in Metal and Radionuclide Reducing Microorganisms

DOE Office of Scientific and Technical Information (OSTI.GOV)

None

2004-04-17

The projects application goals are to: (1) To understand bacterial stress-response to the unique stressors in metal/radionuclide contamination sites; (2) To turn this understanding into a quantitative, data-driven model for exploring policies for natural and biostimulatory bioremediation; (3) To implement proposed policies in the field and compare results to model predictions; and (4) Close the experimental/computation cycle by using discrepancies between models and predictions to drive new measurements and construction of new models. The projects science goals are to: (1) Compare physiological and molecular response of three target microorganisms to environmental perturbation; (2) Deduce the underlying regulatory pathways that controlmore » these responses through analysis of phenotype, functional genomic, and molecular interaction data; (3) Use differences in the cellular responses among the target organisms to understand niche specific adaptations of the stress and metal reduction pathways; (4) From this analysis derive an understanding of the mechanisms of pathway evolution in the environment; and (5) Ultimately, derive dynamical models for the control of these pathways to predict how natural stimulation can optimize growth and metal reduction efficiency at field sites.« less

Digitalis metabolism and human liver alcohol dehydrogenase.

PubMed Central

Frey, W A; Vallee, B L

1980-01-01

Human liver alcohol dehydrogenase (alcohol: NAD" oxidoreductase, EC 1.1.1.1) catalyzes the oxidation of the 3 beta-OH group of digitoxigenin, digoxigenin, and gitoxigenin to their 3-keto derivatives, which have been characterized by high performance liquid chromatography and mass spectrometry. These studies have identified human liver alcohol dehydrogenase as the unknown NAD(H)-dependent liver enzyme specific for the free hydroxyl group at C3 of the cardiac genins; this hydroxyl is the critical site of the genins' enzymatic oxidation and concomitant pharmacological inactivation in humans. Several kinetic approaches have demonstrated that ethanol and the pharmacologically active components of the digitalis glycosides are oxidized with closely similar kcat/Km values at the same site on human liver alcohol dehydrogenase, for which they compete. Human liver alcohol dehydrogenase thereby becomes an important biochemical link in the metabolism, pharmacology, and toxicology of ethanol and these glycosides, structurally unrelated agents that are both used widely. Both the competition of ethanol with these cardiac sterols and the narrow margin of safety in the therapeutic use of digitalis derivatives would seem to place at increased risk those individuals who receive digitalis and simultaneously consume large amounts of ethanol or whose alcohol dehydrogenase function is impaired. PMID:6987673

Sediment data sources and estimated annual suspended-sediment loads of rivers and streams in Colorado

USGS Publications Warehouse

Elliott, J.G.; DeFeyter, K.L.

1986-01-01

Sources of sediment data collected by several government agencies through water year 1984 are summarized for Colorado. The U.S. Geological Survey has collected suspended-sediment data at 243 sites; these data are stored in the U.S. Geological Survey 's water data storage and retrieval system. The U.S. Forest Service has collected suspended-sediment and bedload data at an additional 225 sites, and most of these data are stored in the U.S. Environmental Protection Agency 's water-quality-control information system. Additional unpublished sediment data are in the possession of the collecting entities. Annual suspended-sediment loads were computed for 133 U.S. Geological Survey sediment-data-collection sites using the daily mean water-discharge/sediment-transport-curve method. Sediment-transport curves were derived for each site by one of three techniques: (1) Least-squares linear regression of all pairs of suspended-sediment and corresponding water-discharge data, (2) least-squares linear regression of data sets subdivided on the basis of hydrograph season; and (3) graphical fit to a logarithm-logarithm plot of data. The curve-fitting technique used for each site depended on site-specific characteristics. Sediment-data sources and estimates of annual loads of suspended, bed, and total sediment from several other reports also are summarized. (USGS)

Molecular field coefficients and cation distribution of substituted yttrium iron garnets

NASA Astrophysics Data System (ADS)

Röschmann, P.; Hansen, P.

1981-10-01

The saturation magnetization Ms(T) of Ga, Al, Sc, and CaVBi substituted Y3Fe5O12 (YIG) single crystals and of polycrystalline Ca/Ge and Ca/Ti substituted YIG has been investigated for 4.2 K ⩽T⩽TC. The samples were repeatedly annealed and quenched at different equilibrium temperatures 773 K⩽Te ⩽1523 K. The attained site exchange of Fe and the substituents between the a and d sites resulted in considerable changes of Ms(T). From a fit of the Néel molecular field theory to the Ms(T) data the dependence of the magnetic moments at T = 0 K and of the molecular field coefficients on the amount of nonmagnetic substitutions on the a and d sites were determined. It turned out that ion-specific sets of equations are required accounting for the ''particular ion effect'' of different cation species. The cation distributions inferred from the magnetic data have been analyzed along with a thermodynamic equilibrium model. The derived site stabilizing energies for the mixed Fe-Ga and Fe-Al garnets agree well with recently reported data. New results are presented for the site stabilizing energies in Ca/Ge:YIG and for the substituents Sc and Ti with octahedral site preference.

Prediction of lysine glutarylation sites by maximum relevance minimum redundancy feature selection.

PubMed

Ju, Zhe; He, Jian-Jun

2018-06-01

Lysine glutarylation is new type of protein acylation modification in both prokaryotes and eukaryotes. To better understand the molecular mechanism of glutarylation, it is important to identify glutarylated substrates and their corresponding glutarylation sites accurately. In this study, a novel bioinformatics tool named GlutPred is developed to predict glutarylation sites by using multiple feature extraction and maximum relevance minimum redundancy feature selection. On the one hand, amino acid factors, binary encoding, and the composition of k-spaced amino acid pairs features are incorporated to encode glutarylation sites. And the maximum relevance minimum redundancy method and the incremental feature selection algorithm are adopted to remove the redundant features. On the other hand, a biased support vector machine algorithm is used to handle the imbalanced problem in glutarylation sites training dataset. As illustrated by 10-fold cross-validation, the performance of GlutPred achieves a satisfactory performance with a Sensitivity of 64.80%, a Specificity of 76.60%, an Accuracy of 74.90% and a Matthew's correlation coefficient of 0.3194. Feature analysis shows that some k-spaced amino acid pair features play the most important roles in the prediction of glutarylation sites. The conclusions derived from this study might provide some clues for understanding the molecular mechanisms of glutarylation. Copyright © 2018 Elsevier Inc. All rights reserved.

Incorporating evolution of transcription factor binding sites into annotated alignments.

PubMed

Bais, Abha S; Grossmann, Stefen; Vingron, Martin

2007-08-01

Identifying transcription factor binding sites (TFBSs) is essential to elucidate putative regulatory mechanisms. A common strategy is to combine cross-species conservation with single sequence TFBS annotation to yield "conserved TFBSs". Most current methods in this field adopt a multi-step approach that segregates the two aspects. Again, it is widely accepted that the evolutionary dynamics of binding sites differ from those of the surrounding sequence. Hence, it is desirable to have an approach that explicitly takes this factor into account. Although a plethora of approaches have been proposed for the prediction of conserved TFBSs, very few explicitly model TFBS evolutionary properties, while additionally being multi-step. Recently, we introduced a novel approach to simultaneously align and annotate conserved TFBSs in a pair of sequences. Building upon the standard Smith-Waterman algorithm for local alignments, SimAnn introduces additional states for profiles to output extended alignments or annotated alignments. That is, alignments with parts annotated as gaplessly aligned TFBSs (pair-profile hits)are generated. Moreover,the pair- profile related parameters are derived in a sound statistical framework. In this article, we extend this approach to explicitly incorporate evolution of binding sites in the SimAnn framework. We demonstrate the extension in the theoretical derivations through two position-specific evolutionary models, previously used for modelling TFBS evolution. In a simulated setting, we provide a proof of concept that the approach works given the underlying assumptions,as compared to the original work. Finally, using a real dataset of experimentally verified binding sites in human-mouse sequence pairs,we compare the new approach (eSimAnn) to an existing multi-step tool that also considers TFBS evolution. Although it is widely accepted that binding sites evolve differently from the surrounding sequences, most comparative TFBS identification methods do not explicitly consider this.Additionally, prediction of conserved binding sites is carried out in a multi-step approach that segregates alignment from TFBS annotation. In this paper, we demonstrate how the simultaneous alignment and annotation approach of SimAnn can be further extended to incorporate TFBS evolutionary relationships. We study how alignments and binding site predictions interplay at varying evolutionary distances and for various profile qualities.

One- and two-photon states for quantum information

NASA Astrophysics Data System (ADS)

Peters, Nicholas A.

To find expression stability among transgenic lines, the Recombinase Mediated Transgene Integration (RMTI) technology using the Cre/ lox-mediated site-specific gene integration system was used. The objectives were to develop an efficient method of site-specific transgene integration and to test the effectiveness of this method by assaying transgene expression in the RMTI lines. The RMTI technology allows the precise integration of a transgene in a previously placed target genomic location containing a lox site. The efficiency of CRE-mediated site-specific integration in rice by particle bombardment was found to vary from 3 to 28% in nine different experiments. Some hemizygous site-specific integration plants that were derived from homozygous target locus were found to undergo CRE-mediated reversion of the integration locus. No reversion was observed in callus; however, reverting cells may have been excluded due to selection pressure. The expression of the transgene gus was studied in all 40 callus lines, 12 regenerated T0 plants and the T1 and T2 progenies of 5 lines. The isogenic SC lines had an average expression level based on the activity of beta-glucuronidase of 158 +/- 9 units/mg protein (mean +/- SEM; n=3; variance within SC lines are expressed as standard error of the mean SEM) indicating a significantly higher level of expression, as compared to MC lines that had a much lower expression level 44 +/- 8 units/mg protein (mean +/- SEM; n=3) and the imprecise lines that had 22 +/- 8 units/mg protein (mean +/- SEM; n=3). Transgene expression in the callus cells of precise single copy lines varied by ˜3 fold, whereas that in multi-copy lines varied by ˜30 fold. Furthermore, precise single copy lines, on an average, contained ˜3.5 fold higher expression than multi-copy lines. Transgene expression in the plants of precise single-copy lines was highly variable, which was found to be due to the loss of the integration because of CRE-mediated reversion in the locus. (Abstract shortened by UMI.)

Performance assessment of FY-3C/MERSI on early orbit

NASA Astrophysics Data System (ADS)

Hu, Xiuqing; Xu, Na; Wu, Ronghua; Chen, Lin; Min, Min; Wang, Ling; Xu, Hanlie; Sun, Ling; Yang, Zhongdong; Zhang, Peng

2014-11-01

FY-3C/MERSI has some remarkable improvements compared to the previous MERSIs including better spectral response function (SRF) consistency of different detectors within one band, increasing the capability of lunar observation by space view (SV) and the improvement of radiometric response stability of solar bands. During the In-orbit verification (IOV) commissioning phase, early results that indicate the MERSI representative performance were derived, including the signal noise ratio (SNR), dynamic range, MTF, B2B registration, calibration bias and instrument stability. The SNRs at the solar bands (Bands 1-4 and 6-20) was largely beyond the specifications except for two NIR bands. The in-flight calibration and verification for these bands are also heavily relied on the vicarious techniques such as China radiometric calibration sites(CRCS), cross-calibration, lunar calibration, DCC calibration, stability monitoring using Pseudo Invariant Calibration Sites (PICS) and multi-site radiance simulation. This paper will give the results of the above several calibration methods and monitoring the instrument degradation in early on-orbit time.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kersten, Roland D.; Diedrich, Jolene K.; Yates, III, John R.

Terpenes are ubiquitous natural chemicals with diverse biological functions spanning all three domains of life. In specialized metabolism, the active sites of terpene synthases (TPSs) evolve in shape and reactivity to direct the biosynthesis of a myriad of chemotypes for organismal fitness. As most terpene biosynthesis mechanistically involves highly reactive carbocationic intermediates, the protein surfaces catalyzing these cascade reactions possess reactive regions possibly prone to premature carbocation capture and potentially enzyme inactivation. Here, we show using proteomic and X-ray crystallographic analyses that cationic intermediates undergo capture by conserved active site residues leading to inhibitory self-alkylation. Furthermore, the level of cation-mediatedmore » inactivation increases with mutation of the active site, upon changes in the size and structure of isoprenoid diphosphate substrates, and alongside increases in reaction temperatures. TPSs that individually synthesize multiple products are less prone to self-alkylation then TPSs possessing relatively high product specificity. In total, the results presented suggest that mechanism-based alkylation represents an overlooked mechanistic pressure during the evolution of cation-derived terpene biosynthesis.« less

Mass absorption efficiency of elemental carbon over Van Vihar National Park, Bhopal, India: Temporal variability and implications to estimates of black carbon radiative forcing

NASA Astrophysics Data System (ADS)

Samiksha, S.; Raman, R. S.; Singh, A.

2016-12-01

It is now well recognized that black carbon (a component of aerosols that is similar but not identical to elemental carbon) is an important contributor to global warming, second only to CO2.However, the most popular methods for estimation of black carbon rely on accurate estimates of its mass absorption efficiency (MAE) to convert optical attenuation measurements to black carbon concentrations. Often a constant manufacturer specified MAE is used for this purposes. Recent literature has unequivocally established that MAE shows large spatio-temporal heterogeneities. This is so because MAE depends on emission sources, chemical composition, and mixing state of aerosols. In this study, ambient PM2.5 samples were collected over an ecologically sensitive zone (Van Vihar National Park) in Bhopal, Central India for two years (01 January, 2012 to 31 December, 2013). Samples were collected on Teflon, Nylon, and Tissue quartz filter substrates. Punches of quartz fibre filter were analysed for organic and elemental carbon (OC/EC) by a thermal-optical-transmittance/reflectance (TOT-TOR) analyser operating with a 632 nm laser diode. Teflon filters were also used to interdependently measure PM2.5 attenuation (at 370 nm and 800 nm) by transmissometry. Site-specific mass absorption efficiency (MAE) for elemental carbon over the study site will be derived using a combination of measurements from the TOT/TOR analyser and transmissometer. An assessment of site-specific MAE values, its temporal variability and implications to black carbon radiative forcing will be discussed. It is now well recognized that black carbon (a component of aerosols that is similar but not identical to elemental carbon) is an important contributor to global warming, second only to CO2. However, the most popular methods for estimation of black carbon rely on accurate estimates of its mass absorption efficiency (MAE) to convert optical attenuation measurements to black carbon concentrations. Often a constant manufacturer specified MAE is used for this purposes. Recent literature has unequivocally established that MAE shows large spatio-temporal heterogeneities. This is so because MAE depends on emission sources, chemical composition, and mixing state of aerosols. In this study, ambient PM2.5 samples were collected over an ecologically sensitive zone (Van Vihar National Park) in Bhopal, Central India for two years (01 January, 2012 to 31 December, 2013). Samples were collected on Teflon, Nylon, and Tissue quartz filter substrates. Punches of quartz fibre filter were analysed for organic and elemental carbon (OC/EC) by a thermal-optical-transmittance/reflectance (TOT-TOR) analyser operating with a 632 nm laser diode. Teflon filters were also used to interdependently measure PM2.5 attenuation (at 370 nm and 800 nm) by transmissometry. Site-specific mass absorption efficiency (MAE) for elemental carbon over the study site will be derived using a combination of measurements from the TOT/TOR analyser and transmissometer. An assessment of site-specific MAE values, its temporal variability and implications to black carbon radiative forcing will be discussed.

Site-specific Isopeptide Bridge Tethering of Chimeric gp41 N-terminal Heptad Repeat Helical Trimers for the Treatment of HIV-1 Infection.

PubMed

Wang, Chao; Li, Xue; Yu, Fei; Lu, Lu; Jiang, Xifeng; Xu, Xiaoyu; Wang, Huixin; Lai, Wenqing; Zhang, Tianhong; Zhang, Zhenqing; Ye, Ling; Jiang, Shibo; Liu, Keliang

2016-08-26

Peptides derived from the N-terminal heptad repeat (NHR) of HIV-1 gp41 can be potent inhibitors against viral entry when presented in a nonaggregating trimeric coiled-coil conformation via the introduction of exogenous trimerization motifs and intermolecular disulfide bonds. We recently discovered that crosslinking isopeptide bridges within the de novo helical trimers added exceptional resistance to unfolding. Herein, we attempted to optimize (CCIZN17)3, a representative disulfide bond-stabilized chimeric NHR-trimer, by incorporating site-specific interhelical isopeptide bonds as the redox-sensitive disulfide surrogate. In this process, we systematically examined the effect of isopeptide bond position and molecular sizes of auxiliary trimeric coiled-coil motif and NHR fragments on the antiviral potency of these NHR-trimers. Pleasingly, (IZ14N24N)3 possessed promising inhibitory activity against HIV-1 infection and markedly increased proteolytic stability relative to its disulfide-tethered counterpart, suggesting good potential for further development as an effective antiviral agent for treatment of HIV-1 infection.

Pressure for Pattern-Specific Intertypic Recombination between Sabin Polioviruses: Evolutionary Implications.

PubMed

Korotkova, Ekaterina; Laassri, Majid; Zagorodnyaya, Tatiana; Petrovskaya, Svetlana; Rodionova, Elvira; Cherkasova, Elena; Gmyl, Anatoly; Ivanova, Olga E; Eremeeva, Tatyana P; Lipskaya, Galina Y; Agol, Vadim I; Chumakov, Konstantin

2017-11-22

Complete genomic sequences of a non-redundant set of 70 recombinants between three serotypes of attenuated Sabin polioviruses as well as location (based on partial sequencing) of crossover sites of 28 additional recombinants were determined and compared with the previously published data. It is demonstrated that the genomes of Sabin viruses contain distinct strain-specific segments that are eliminated by recombination. The presumed low fitness of these segments could be linked to mutations acquired upon derivation of the vaccine strains and/or may have been present in wild-type parents of Sabin viruses. These "weak" segments contribute to the propensity of these viruses to recombine with each other and with other enteroviruses as well as determine the choice of crossover sites. The knowledge of location of such segments opens additional possibilities for the design of more genetically stable and/or more attenuated variants, i.e., candidates for new oral polio vaccines. The results also suggest that the genome of wild polioviruses, and, by generalization, of other RNA viruses, may harbor hidden low-fitness segments that can be readily eliminated only by recombination.

Bispecific small molecule–antibody conjugate targeting prostate cancer

PubMed Central

Kim, Chan Hyuk; Axup, Jun Y.; Lawson, Brian R.; Yun, Hwayoung; Tardif, Virginie; Choi, Sei Hyun; Zhou, Quan; Dubrovska, Anna; Biroc, Sandra L.; Marsden, Robin; Pinstaff, Jason; Smider, Vaughn V.; Schultz, Peter G.

2013-01-01

Bispecific antibodies, which simultaneously target CD3 on T cells and tumor-associated antigens to recruit cytotoxic T cells to cancer cells, are a promising new approach to the treatment of hormone-refractory prostate cancer. Here we report a site-specific, semisynthetic method for the production of bispecific antibody-like therapeutics in which a derivative of the prostate-specific membrane antigen-binding small molecule DUPA was selectively conjugated to a mutant αCD3 Fab containing the unnatural amino acid, p-acetylphenylalanine, at a defined site. Homogeneous conjugates were generated in excellent yields and had good solubility. The efficacy of the conjugate was optimized by modifying the linker structure, relative binding orientation, and stoichiometry of the ligand. The optimized conjugate showed potent and selective in vitro activity (EC50 ∼100 pM), good serum half-life, and potent in vivo activity in prophylactic and treatment xenograft mouse models. This semisynthetic approach is likely to be applicable to the generation of additional bispecific agents using drug-like ligands selective for other cell-surface receptors. PMID:24127589

Site-specific Isopeptide Bridge Tethering of Chimeric gp41 N-terminal Heptad Repeat Helical Trimers for the Treatment of HIV-1 Infection

PubMed Central

Wang, Chao; Li, Xue; Yu, Fei; Lu, Lu; Jiang, Xifeng; Xu, Xiaoyu; Wang, Huixin; Lai, Wenqing; Zhang, Tianhong; Zhang, Zhenqing; Ye, Ling; Jiang, Shibo; Liu, Keliang

2016-01-01

Peptides derived from the N-terminal heptad repeat (NHR) of HIV-1 gp41 can be potent inhibitors against viral entry when presented in a nonaggregating trimeric coiled-coil conformation via the introduction of exogenous trimerization motifs and intermolecular disulfide bonds. We recently discovered that crosslinking isopeptide bridges within the de novo helical trimers added exceptional resistance to unfolding. Herein, we attempted to optimize (CCIZN17)3, a representative disulfide bond-stabilized chimeric NHR-trimer, by incorporating site-specific interhelical isopeptide bonds as the redox-sensitive disulfide surrogate. In this process, we systematically examined the effect of isopeptide bond position and molecular sizes of auxiliary trimeric coiled-coil motif and NHR fragments on the antiviral potency of these NHR-trimers. Pleasingly, (IZ14N24N)3 possessed promising inhibitory activity against HIV-1 infection and markedly increased proteolytic stability relative to its disulfide-tethered counterpart, suggesting good potential for further development as an effective antiviral agent for treatment of HIV-1 infection. PMID:27562370

Evaluation of vegetation indices for rangeland biomass estimation in the Kimberley area of Western Australia

NASA Astrophysics Data System (ADS)

Mundava, C.; Helmholz, P.; Schut, A. G. T.; Corner, R.; McAtee, B.; Lamb, D. W.

2014-09-01

The objective of this paper is to test the relationships between Above Ground Biomass (AGB) and remotely sensed vegetation indices for AGB assessments in the Kimberley area in Western Australia. For 19 different sites, vegetation indices were derived from eight Landsat ETM+ scenes over a period of two years (2011-2013). The sites were divided into three groups (Open plains, Bunch grasses and Spinifex) based on similarities in dominant vegetation types. Dry and green biomass fractions were measured at these sites. Single and multiple regression relationships between vegetation indices and green and total AGB were calibrated and validated using a "leave site out" cross validation. Four tests were compared: (1) relationships between AGB and vegetation indices combining all sites; (2) separate relationships per site group; (3) multiple regressions including selected vegetation indices per site group; and (4) as in 3 but including rainfall and elevation data. Results indicate that relationships based on single vegetation indices are moderately accurate for green biomass in wide open plains covered with annual grasses. The cross-validation results for green AGB improved for a combination of indices for the Open plains and Bunch grasses sites, but not for Spinifex sites. When rainfall and elevation data are included, cross validation improved slightly with a Q2 of 0.49-0.72 for Open plains and Bunch grasses sites respectively. Cross validation results for total AGB were moderately accurate (Q2 of 0.41) for Open plains but weak or absent for other site groups despite good calibration results, indicating strong influence of site-specific factors.

Development of a simulated earthworm gut for determining bioaccessible arsenic, copper, and zinc from soil.

PubMed

Ma, Wai K; Smith, Ben A; Stephenson, Gladys L; Siciliano, Steven D

2009-07-01

Soil physicochemical characteristics and contamination levels alter the bioavailability of metals to terrestrial invertebrates. Current laboratory-derived benchmark concentrations used to estimate risk do not take into account site-specific conditions, such as contaminant sequestration, and site-specific risk assessment requires a battery of time-consuming and costly toxicity tests. The development of an in vitro simulator for earthworm bioaccessibility would significantly shorten analytical time and enable site managers to focus on areas of greatest concern. The simulated earthworm gut (SEG) was developed to measure the bioaccessibility of metals in soil to earthworms by mimicking the gastrointestinal fluid composition of earthworms. Three formulations of the SEG (enzymes, microbial culture, enzymes and microbial culture) were developed and used to digest field soils from a former industrial site with varying physicochemical characteristics and contamination levels. Formulations containing enzymes released between two to 10 times more arsenic, copper, and zinc from contaminated soils compared with control and 0.01 M CaCl2 extractions. Metal concentrations in extracts from SEG formulation with microbial culture alone were not different from values for chemical extractions. The mechanism for greater bioaccessible metal concentrations from enzyme-treated soils is uncertain, but it is postulated that enzymatic digestion of soil organic matter might release sequestered metal. The relevance of these SEG results will need validation through further comparison and correlation with bioaccumulation tests, alternative chemical extraction tests, and a battery of chronic toxicity tests with invertebrates and plants.

Developing ecologically based PCB, pesticide, and metal remedial goals for an impacted northeast wooded swamp

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rury, P.M.; Turton, D.J.

Historically, remedial goals at hazardous waste sites have been developed based on human health risk estimates. As the disciplines of remedial investigation, risk assessment, and remedial design have evolved, there has been a shift toward the development of remedial goals that are protective of both human health and the environment. This has increased the need for sound quantitative ecological risk methodologies from which to derive ecologically protective remedial goals. The foundation of many ecological risk assessment models is the bioconcentration or bioaccumulation factor that estimates the partitioning of the compound of concern between the media (e.g., water, soil, or food)more » and the organism. Simple dietary food-chain models are then used to estimate the dose and resulting risk to higher trophic levels. For a Superfund site that encompassed a northeastern wooded swamp, a PCB pesticide and metal uptake and toxicity study was conducted on the earthworm commonly known as the red wiggler (Eisenea foetida). The study resulted in site-specific sediment to earthworm bioconcentration factors for PCBs and a range of pesticides and metals. In addition, largemouth bass and yellow perch were collected from an impacted pond to identify PCB and pesticide concentrations in mink (Mustela vison) prey. Utilizing the empirical data and site-specific bioconcentration factors in food-chain models, potential risks to the American woodcock (Scolopax minor) and mink were assessed, and ecologically protective PCB, pesticide, and metal remedial goals for the sediments of the wooded swamp were developed.« less

Bacterial biomass and activity in the deep waters of the eastern Atlantic—evidence of a barophilic community

NASA Astrophysics Data System (ADS)

Patching, J. W.; Eardly, D.

1997-09-01

Bacterial biomass and activity were investigated in deep waters at two sites in the eastern Atlantic, of similar depth (4560-4800 m), but varying in their nutritional status. The Northern (N) site was eutrophic and subject to a strong seasonal input of surface derived organic matter (phytodetritus) to the sediment. The Southern (S) site was oligotrophic. Deep water at this site does not appear to receive any strong seasonal input. Bacterial numbers in the deep water column at the N site showed no significant seasonal variation but were greater than those at the S site. Deep water bacteria were typically small and free-living. From biovolume determinations, it was estimated that mean concentrations of bacterial organic carbon at depths greater than 500 m were 0.12 (0.03-0.29) μg C 1 -1 and 0.02 (0.01-0.04) μg C 1 -1 at the N and S sites, respectively. Rates of thymidine and leucine incorporation were used as indicators of bacterial activity. Bacterial communities in water in contact with the sediment (SCW; sediment contact water) at both sites (but especially at the S site) were strongly barophilic at in situ temperatures (2.5-4.1°C). The barophilic response of thymidine incorporation was enhanced when SCW samples from the N site were incubated at 11.5°C. It is proposed that this result indicated an elevating effect of pressure on cardinal temperatures and that the SCW community was obligately psychrophilic when unpressurised. Comparison of cell-specific incorporation rates determined under in situ conditions showed bacteria in the SCW to have levels of activity comparable with bacteria from a depth of 150 m. Thymidine incorporation rates were highest in SCW samples taken at the N site in May 1988 and September 1989. Thymidine incorporation by SCW samples taken immediately before (10 April 1994) the main spring-bloom-associated deposition of phytodetritus was significantly lower and comparable with that determined for the oligotrophic S site. The attributes exhibited by the SCW community appeared to be highly localised. We conclude that the bacterial communities of the SCW are active and adapted to their environment. Activity is influenced by the trophic nature of the site and may show temporal changes linked with episodic food supply. We postulate that the existence of such communities is linked to the role of the sediment-water interface as the initial site of deposition of sea-surface derived labile organic material.

Preparation and preliminary characterization of crystallizing fluorescent derivatives of chicken egg white lysozyme

NASA Astrophysics Data System (ADS)

Sumida, John P.; Forsythe, Elizabeth L.; Pusey, Marc L.

2001-11-01

Fluorescence is one of the most versatile and powerful tools for the study of macromolecules. While most proteins are intrinsically fluorescent, working at crystallization concentrations require the use of covalently prepared derivatives added as tracers. This approach requires derivatives that do not markedly affect the crystal packing. We have prepared fluorescent derivatives of chicken egg white lysozyme with probes bound to one of two different sites on the protein molecule. Lucifer yellow and 5-(2-aminoethyl)aminonapthalene-1-sulfonic acid (EDANS) have been attached to the side chain carboxyl of Asp 101 using a carbodiimide coupling procedure. Asp 101 lies within the active site cleft, and it is believed that the probes are "buried" within that cleft. Lucifer yellow and EDANS probes with iodoacetamide reactive groups have been bound to His 15, located on the "back side" of the molecule relative to the active site. All the derivatives fluoresce in the solution and the crystalline states. Fluorescence characterization has focused on determination of binding effects on the probe quantum yield, lifetime, absorption and emission spectra, and quenching by added solutes. Quenching studies show that, as postulated, the Asp 101-bound probes are partially sheltered from the bulk solution by their location within the active site cleft. Probes bound to His 15 have quenching constants about equal to those for the free probes, indicating that this site is highly exposed to the bulk solution.

Transport and fate of radionuclides in aquatic environments--the use of ecosystem modelling for exposure assessments of nuclear facilities.

PubMed

Kumblad, L; Kautsky, U; Naeslund, B

2006-01-01

In safety assessments of nuclear facilities, a wide range of radioactive isotopes and their potential hazard to a large assortment of organisms and ecosystem types over long time scales need to be considered. Models used for these purposes have typically employed approaches based on generic reference organisms, stylised environments and transfer functions for biological uptake exclusively based on bioconcentration factors (BCFs). These models are of non-mechanistic nature and involve no understanding of uptake and transport processes in the environment, which is a severe limitation when assessing real ecosystems. In this paper, ecosystem models are suggested as a method to include site-specific data and to facilitate the modelling of dynamic systems. An aquatic ecosystem model for the environmental transport of radionuclides is presented and discussed. With this model, driven and constrained by site-specific carbon dynamics and three radionuclide specific mechanisms: (i) radionuclide uptake by plants, (ii) excretion by animals, and (iii) adsorption to organic surfaces, it was possible to estimate the radionuclide concentrations in all components of the modelled ecosystem with only two radionuclide specific input parameters (BCF for plants and Kd). The importance of radionuclide specific mechanisms for the exposure to organisms was examined, and probabilistic and sensitivity analyses to assess the uncertainties related to ecosystem input parameters were performed. Verification of the model suggests that this model produces analogous results to empirically derived data for more than 20 different radionuclides.

Preliminary survey on site-adaptation techniques for satellite-derived and reanalysis solar radiation datasets

DOE Office of Scientific and Technical Information (OSTI.GOV)

Polo, J.; Wilbert, S.; Ruiz-Arias, J. A.

2016-07-01

At any site, the bankability of a projected solar power plant largely depends on the accuracy and general quality of the solar radiation data generated during the solar resource assessment phase. The term 'site adaptation' has recently started to be used in the framework of solar energy projects to refer to the improvement that can be achieved in satellite-derived solar irradiance and model data when short-term local ground measurements are used to correct systematic errors and bias in the original dataset. This contribution presents a preliminary survey of different possible techniques that can improve long-term satellite-derived and model-derived solar radiationmore » data through the use of short-term on-site ground measurements. The possible approaches that are reported here may be applied in different ways, depending on the origin and characteristics of the uncertainties in the modeled data. This work, which is the first step of a forthcoming in-depth assessment of methodologies for site adaptation, has been done within the framework of the International Energy Agency Solar Heating and Cooling Programme Task 46 'Solar Resource Assessment and Forecasting.'« less

Insights into Methyltransferase Specificity and Bioactivity of Derivatives of the Antibiotic Plantazolicin

DOE PAGES

Hao, Yue; Blair, Patricia M.; Sharma, Abhishek; ...

2015-01-30

Peptide antibiotics represent a class of conformationally-constrained natural products of growing pharmaceutical interest. Plantazolicin (PZN) is a linear, polyheterocyclic natural product with highly selective and potent activity against the anthrax-causing bacterium, Bacillus anthracis. The bioactivity of PZN is contingent on dimethylation of its N-terminal Arg residue by an S-adenosylmethionine-dependent methyltransferase. Here in this paper, we explore the substrate tolerances of two homologous PZN methyltransferases by carrying out kinetic analyses of the enzymes against a synthetic panel of truncated PZN analogs containing the N-terminal Arg residue. X-ray cocrystal structures of the PZN methyltransferases with each of these heterocycle-containing substrates provide amore » rationale for understanding the strict substrate specificity of these enzymes. Kinetic studies of structure-guided, site-specific variants allowed for the assignment of residues governing catalysis and substrate scope. Microbiological testing further revealed that upon dimethylation of the N-terminal Arg, a pentaheterocyclized PZN analog retained potent anti-B. anthracis activity, nearly equal to that of full-length PZN. These studies may be useful in the biosynthetic engineering of natural product analogs with different bioactivity profiles, as demonstrated by our identification of a truncated plantazolicin derivative that is active against methicillin-resistant Staphylococcus aureus (MRSA).« less

Insights into Methyltransferase Specificity and Bioactivity of Derivatives of the Antibiotic Plantazolicin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hao, Yue; Blair, Patricia M.; Sharma, Abhishek

Peptide antibiotics represent a class of conformationally-constrained natural products of growing pharmaceutical interest. Plantazolicin (PZN) is a linear, polyheterocyclic natural product with highly selective and potent activity against the anthrax-causing bacterium, Bacillus anthracis. The bioactivity of PZN is contingent on dimethylation of its N-terminal Arg residue by an S-adenosylmethionine-dependent methyltransferase. Here in this paper, we explore the substrate tolerances of two homologous PZN methyltransferases by carrying out kinetic analyses of the enzymes against a synthetic panel of truncated PZN analogs containing the N-terminal Arg residue. X-ray cocrystal structures of the PZN methyltransferases with each of these heterocycle-containing substrates provide amore » rationale for understanding the strict substrate specificity of these enzymes. Kinetic studies of structure-guided, site-specific variants allowed for the assignment of residues governing catalysis and substrate scope. Microbiological testing further revealed that upon dimethylation of the N-terminal Arg, a pentaheterocyclized PZN analog retained potent anti-B. anthracis activity, nearly equal to that of full-length PZN. These studies may be useful in the biosynthetic engineering of natural product analogs with different bioactivity profiles, as demonstrated by our identification of a truncated plantazolicin derivative that is active against methicillin-resistant Staphylococcus aureus (MRSA).« less

PEGylated DX-1000: pharmacokinetics and antineoplastic activity of a specific plasmin inhibitor.

PubMed

Devy, Laetitia; Rabbani, Shafaat A; Stochl, Mark; Ruskowski, Mary; Mackie, Ian; Naa, Laurent; Toews, Mark; van Gool, Reinoud; Chen, Jie; Ley, Art; Ladner, Robert C; Dransfield, Daniel T; Henderikx, Paula

2007-11-01

Novel inhibitors of the urokinase-mediated plasminogen (plg) activation system are potentially of great clinical benefit as anticancer treatments. Using phage display, we identified DX-1000 a tissue factor pathway inhibitor-derived Kunitz domain protein which is a specific high-affinity inhibitor of plasmin (pln) (K(i) = 99 pM). When tested in vitro, DX-1000 blocks plasmin-mediated pro-matrix metalloproteinase-9 (proMMP-9) activation on cells and dose-dependently inhibits tube formation, while not significantly affecting hemostasis and coagulation. However, this low-molecular weight protein inhibitor ( approximately 7 kDa) exhibits rapid plasma clearance in mice and rabbits, limiting its potential clinical use in chronic diseases. After site-specific PEGylation, DX-1000 retains its activity and exhibits a decreased plasma clearance. This PEGylated derivative is effective in vitro, as well as potent in inhibiting tumor growth of green fluorescent protein (GFP)-labeled MDA-MB-231 cells. 4PEG-DX-1000 treatment causes a significant reduction of urokinase-type plasminogen activator (uPA) and plasminogen expressions, a reduction of tumor proliferation, and vascularization. 4PEG-DX-1000 treatment significantly decreases the level of active mitogen-activated protein kinase (MAPK) in the primary tumors and reduces metastasis incidence. Together, our results demonstrate the potential value of plasmin inhibitors as therapeutic agents for blocking breast cancer growth and metastasis.

Comparison of hip geometry, strength, and estimated fracture risk in women with anorexia nervosa and overweight/obese women.

PubMed

Bachmann, Katherine Neubecker; Fazeli, Pouneh K; Lawson, Elizabeth A; Russell, Brian M; Riccio, Ariana D; Meenaghan, Erinne; Gerweck, Anu V; Eddy, Kamryn; Holmes, Tara; Goldstein, Mark; Weigel, Thomas; Ebrahimi, Seda; Mickley, Diane; Gleysteen, Suzanne; Bredella, Miriam A; Klibanski, Anne; Miller, Karen K

2014-12-01

Data suggest that anorexia nervosa (AN) and obesity are complicated by elevated fracture risk, but skeletal site-specific data are lacking. Traditional bone mineral density (BMD) measurements are unsatisfactory at both weight extremes. Hip structural analysis (HSA) uses dual-energy X-ray absorptiometry data to estimate hip geometry and femoral strength. Factor of risk (φ) is the ratio of force applied to the hip from a fall with respect to femoral strength; higher values indicate higher hip fracture risk. The objective of the study was to investigate hip fracture risk in AN and overweight/obese women. This was a cross-sectional study. The study was conducted at a Clinical Research Center. PATIENTS included 368 women (aged 19-45 y): 246 AN, 53 overweight/obese, and 69 lean controls. HSA-derived femoral geometry, peak factor of risk for hip fracture, and factor of risk for hip fracture attenuated by trochanteric soft tissue (φ(attenuated)) were measured. Most HSA-derived parameters were impaired in AN and superior in obese/overweight women vs controls at the narrow neck, intertrochanteric, and femoral shaft (P ≤ .03). The φ(attenuated) was highest in AN and lowest in overweight/obese women (P < .0001). Lean mass was associated with superior, and duration of amenorrhea with inferior, HSA-derived parameters and φ(attenuated) (P < .05). Mean φ(attenuated) (P = .036), but not femoral neck BMD or HSA-estimated geometry, was impaired in women who had experienced fragility fractures. Femoral geometry by HSA, hip BMD, and factor of risk for hip fracture attenuated by soft tissue are impaired in AN and superior in obesity, suggesting higher and lower hip fracture risk, respectively. Only attenuated factor of risk was associated with fragility fracture prevalence, suggesting that variability in soft tissue padding may help explain site-specific fracture risk not captured by BMD.

Characterization of air manganese exposure estimates for residents in two Ohio towns

PubMed Central

Colledge, Michelle A.; Julian, Jaime R.; Gocheva, Vihra V.; Beseler, Cheryl L.; Roels, Harry A.; Lobdell, Danelle T.; Bowler, Rosemarie M.

2016-01-01

This study was conducted to derive receptor-specific outdoor exposure concentrations of total suspended particulate (TSP) and respirable (dae ≤ 10 μm) air manganese (air-Mn) for East Liverpool and Marietta (Ohio) in the absence of facility emissions data, but where long-term air measurements were available. Our “site-surface area emissions method” used U.S. Environmental Protection Agency’s (EPA) AERMOD (AMS/EPA Regulatory Model) dispersion model and air measurement data to estimate concentrations for residential receptor sites in the two communities. Modeled concentrations were used to create ratios between receptor points and calibrated using measured data from local air monitoring stations. Estimated outdoor air-Mn concentrations were derived for individual study subjects in both towns. The mean estimated long-term air-Mn exposure levels for total suspended particulate were 0.35 μg/m3 (geometric mean [GM]) and 0.88 μg/m3 (arithmetic mean [AM]) in East Liverpool (range: 0.014–6.32 μg/m3) and 0.17 μg/m3 (GM) and 0.21 μg/m3 (AM) in Marietta (range: 0.03–1.61 μg/m3). Modeled results compared well with averaged ambient air measurements from local air monitoring stations. Exposure to respirable Mn particulate matter (PM10; PM <10 μm) was higher in Marietta residents. Implications Few available studies evaluate long-term health outcomes from inhalational manganese (Mn) exposure in residential populations, due in part to challenges in measuring individual exposures. Local long-term air measurements provide the means to calibrate models used in estimating long-term exposures. Furthermore, this combination of modeling and ambient air sampling can be used to derive receptor-specific exposure estimates even in the absence of source emissions data for use in human health outcome studies. PMID:26211636

Comparison of Hip Geometry, Strength, and Estimated Fracture Risk in Women With Anorexia Nervosa and Overweight/Obese Women

PubMed Central

Bachmann, Katherine Neubecker; Fazeli, Pouneh K.; Lawson, Elizabeth A.; Russell, Brian M.; Riccio, Ariana D.; Meenaghan, Erinne; Gerweck, Anu V.; Eddy, Kamryn; Holmes, Tara; Goldstein, Mark; Weigel, Thomas; Ebrahimi, Seda; Mickley, Diane; Gleysteen, Suzanne; Bredella, Miriam A.; Klibanski, Anne

2014-01-01

Context: Data suggest that anorexia nervosa (AN) and obesity are complicated by elevated fracture risk, but skeletal site-specific data are lacking. Traditional bone mineral density (BMD) measurements are unsatisfactory at both weight extremes. Hip structural analysis (HSA) uses dual-energy X-ray absorptiometry data to estimate hip geometry and femoral strength. Factor of risk (φ) is the ratio of force applied to the hip from a fall with respect to femoral strength; higher values indicate higher hip fracture risk. Objective: The objective of the study was to investigate hip fracture risk in AN and overweight/obese women. Design: This was a cross-sectional study. Setting: The study was conducted at a Clinical Research Center. Patients: Patients included 368 women (aged 19–45 y): 246 AN, 53 overweight/obese, and 69 lean controls. Main Outcome Measures: HSA-derived femoral geometry, peak factor of risk for hip fracture, and factor of risk for hip fracture attenuated by trochanteric soft tissue (φattenuated) were measured. Results: Most HSA-derived parameters were impaired in AN and superior in obese/overweight women vs controls at the narrow neck, intertrochanteric, and femoral shaft (P ≤ .03). The φattenuated was highest in AN and lowest in overweight/obese women (P < .0001). Lean mass was associated with superior, and duration of amenorrhea with inferior, HSA-derived parameters and φattenuated (P < .05). Mean φattenuated (P = .036), but not femoral neck BMD or HSA-estimated geometry, was impaired in women who had experienced fragility fractures. Conclusions: Femoral geometry by HSA, hip BMD, and factor of risk for hip fracture attenuated by soft tissue are impaired in AN and superior in obesity, suggesting higher and lower hip fracture risk, respectively. Only attenuated factor of risk was associated with fragility fracture prevalence, suggesting that variability in soft tissue padding may help explain site-specific fracture risk not captured by BMD. PMID:25062461

Land Surface Albedo from MERIS Reflectances Using MODIS Directional Factors

NASA Technical Reports Server (NTRS)

Schaaf, Crystal L. B.; Gao, Feng; Strahler, Alan H.

2004-01-01

MERIS Level 2 surface reflectance products are now available to the scientific community. This paper demonstrates the production of MERIS-derived surface albedo and Nadir Bidirectional Reflectance Distribution Function (BRDF) adjusted reflectances by coupling the MERIS data with MODIS BRDF products. Initial efforts rely on the specification of surface anisotropy as provided by the global MODIS BRDF product for a first guess of the shape of the BRDF and then make use all of the coincidently available, partially atmospherically corrected, cloud cleared, MERIS observations to generate MERIS-derived BRDF and surface albedo quantities for each location. Comparisons between MODIS (aerosol-corrected) and MERIS (not-yet aerosol-corrected) surface values from April and May 2003 are also presented for case studies in Spain and California as well as preliminary comparisons with field data from the Devil's Rock Surfrad/BSRN site.

Endogenous siRNAs and noncoding RNA-derived small RNAs are expressed in adult mouse hippocampus and are up-regulated in olfactory discrimination training.

PubMed

Smalheiser, Neil R; Lugli, Giovanni; Thimmapuram, Jyothi; Cook, Edwin H; Larson, John

2011-01-01

We previously proposed that endogenous siRNAs may regulate synaptic plasticity and long-term gene expression in the mammalian brain. Here, a hippocampal-dependent task was employed in which adult mice were trained to execute a nose-poke in a port containing one of two simultaneously present odors in order to obtain a reward. Mice demonstrating olfactory discrimination training were compared to pseudo-training and nose-poke control groups; size-selected hippocampal RNA was subjected to Illumina deep sequencing. Sequences that aligned uniquely and exactly to the genome without uncertain nucleotide assignments, within exons or introns of MGI annotated genes, were examined further. The data confirm that small RNAs having features of endogenous siRNAs are expressed in brain; that many of them derive from genes that regulate synaptic plasticity (and have been implicated in neuropsychiatric diseases); and that hairpin-derived endo-siRNAs and the 20- to 23-nt size class of small RNAs show a significant increase during an early stage of training. The most abundant putative siRNAs arose from an intronic inverted repeat within the SynGAP1 locus; this inverted repeat was a substrate for dicer in vitro, and SynGAP1 siRNA was specifically associated with Argonaute proteins in vivo. Unexpectedly, a dramatic increase with training (more than 100-fold) was observed for a class of 25- to 30-nt small RNAs derived from specific sites within snoRNAs and abundant noncoding RNAs (Y1 RNA, RNA component of mitochondrial RNAse P, 28S rRNA, and 18S rRNA). Further studies are warranted to characterize the role(s) played by endogenous siRNAs and noncoding RNA-derived small RNAs in learning and memory.

A tiered approach for the human health risk assessment for consumption of vegetables from with cadmium-contaminated land in urban areas

DOE Office of Scientific and Technical Information (OSTI.GOV)

Swartjes, Frank A., E-mail: frank.swartjes@rivm.nl; Versluijs, Kees W.; Otte, Piet F.

Consumption of vegetables that are grown in urban areas takes place worldwide. In developing countries, vegetables are traditionally grown in urban areas for cheap food supply. In developing and developed countries, urban gardening is gaining momentum. A problem that arises with urban gardening is the presence of contaminants in soil, which can be taken up by vegetables. In this study, a scientifically-based and practical procedure has been developed for assessing the human health risks from the consumption of vegetables from cadmium-contaminated land. Starting from a contaminated site, the procedure follows a tiered approach which is laid out as follows. Inmore » Tier 0, the plausibility of growing vegetables is investigated. In Tier 1 soil concentrations are compared with the human health-based Critical soil concentration. Tier 2 offers the possibility for a detailed site-specific human health risk assessment in which calculated exposure is compared to the toxicological reference dose. In Tier 3, vegetable concentrations are measured and tested following a standardized measurement protocol. To underpin the derivation of the Critical soil concentrations and to develop a tool for site-specific assessment the determination of the representative concentration in vegetables has been evaluated for a range of vegetables. The core of the procedure is based on Freundlich-type plant–soil relations, with the total soil concentration and the soil properties as variables. When a significant plant–soil relation is lacking for a specific vegetable a geometric mean of BioConcentrationFactors (BCF) is used, which is normalized according to soil properties. Subsequently, a ‘conservative’ vegetable-group-consumption-rate-weighted BioConcentrationFactor is calculated as basis for the Critical soil concentration (Tier 1). The tool to perform site-specific human health risk assessment (Tier 2) includes the calculation of a ‘realistic worst case’ site-specific vegetable-group-consumption-rate-weighted BioConcentrationFactor. -- Highlights: • A scientifically-based and practical procedure has been developed for assessing the human health risks from the consumption of vegetables. • Uptake characteristics of cadmium in a series of vegetables is represented by a vegetable-group-consumption-rate-weighted BioConcentrationFactor. • Calculations and measurement steps are combined.« less

Utility theoretic approach to estimating the demand for and benefits from recreational fishing: the impact of acid rain

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shaw, W.D. Jr.

1985-01-01

The Adirondack State Park has been hit especially hard by acid precipitation. Acid precipitation impacts particular species of fish at various high altitudes sites in the park. The author estimates consumer surplus measures for changes in a variable to proxy the stock size of these species at specific sites. To do this he first estimates the individual's demand for a recreation site as a function of site prices and the site's characteristics. The demand function for the individual is derived from a utility function. A travel cost approach is used to estimate an individual's share of total fishing time spentmore » at a five fishing site. The shares are estimated by maximum likelihood and the results indicate that price and the three characteristics do explain the allocation of the individual's time spent at the various sites selected for the analysis. Finally, consumer surplus measures for a reduction in the catch rates of the species most likely to be affected by Acid Precipitation are calculated. The meaning of these measures in the context of a model that assumes weak separability is examined. These reductions in catch rates can be linked to changes in the level of Acid Precipitation in the Park, and this provides us with a method for quantifying the impact of acid precipitation on recreation fishing.« less

Statistical and Economic Techniques for Site-specific Nematode Management.

PubMed

Liu, Zheng; Griffin, Terry; Kirkpatrick, Terrence L

2014-03-01

Recent advances in precision agriculture technologies and spatial statistics allow realistic, site-specific estimation of nematode damage to field crops and provide a platform for the site-specific delivery of nematicides within individual fields. This paper reviews the spatial statistical techniques that model correlations among neighboring observations and develop a spatial economic analysis to determine the potential of site-specific nematicide application. The spatial econometric methodology applied in the context of site-specific crop yield response contributes to closing the gap between data analysis and realistic site-specific nematicide recommendations and helps to provide a practical method of site-specifically controlling nematodes.

Long-period amplification in deep alluvial basins and consequences for site-specific probabilistic seismic-hazard: the case of Castelleone in the Po Plain (Northern Italy)

NASA Astrophysics Data System (ADS)

Barani, S.; Mascandola, C.; Massa, M.; Spallarossa, D.

2017-12-01

The recent Emilia seismic sequence (Northern Italy) occurred at the end of the first half of 2012 with main shock of Mw6.1 highlighted the importance of studying site effects in the Po Plain, the larger and deeper sedimentary basin in Italy. As has long been known, long-period amplification related to deep sedimentary basins can significantly affect the characteristics of the ground-motion induced by strong earthquakes. It follows that the effects of deep sedimentary deposits on ground shaking require special attention during the definition of the design seismic action. The work presented here analyzes the impact of deep-soil discontinuities on ground-motion amplification, with particular focus on long-period probabilistic seismic-hazard assessment. The study focuses on the site of Castelleone, where a seismic station of the Italian National Seismic Network has been recording since 2009. Our study includes both experimental and numerical site response analyses. Specifically, extensive active and passive geophysical measurements were carried out in order to define a detailed shear-wave velocity (VS) model to be used in the numerical analyses. These latter are needed to assess the site-specific ground-motion hazard. Besides classical seismic refraction profiles and multichannel analysis of surface waves, we analyzed ambient vibration measurements in both single and array configurations. The VS profile was determined via joint inversion of the experimental phase-velocity dispersion curve with the ellipticity curve derived from horizontal-to-vertical spectral ratios. The profile shows two main discontinuities at depths of around 160 and 1350 m, respectively. The probabilistic site-specific hazard was assessed in terms of both spectral acceleration and displacement. A partially non-ergodic approach was adopted. We have found that the spectral acceleration hazard is barely sensitive to long-period (up to 10 s) amplification related to the deeper discontinuity whereas the displacement hazard is strongly affected. Our results show that neglecting the effects of the deeper discontinuity implies an underestimation of the hazard of up to about 49% for a mean return period (MRP) of 475 years and 57% for an MRP of 2475 years, with possible consequences on the design of very tall buildings and large bridges.

Probabilistic inference of ecohydrological parameters using observations from point to satellite scales

NASA Astrophysics Data System (ADS)

Bassiouni, Maoya; Higgins, Chad W.; Still, Christopher J.; Good, Stephen P.

2018-06-01

Vegetation controls on soil moisture dynamics are challenging to measure and translate into scale- and site-specific ecohydrological parameters for simple soil water balance models. We hypothesize that empirical probability density functions (pdfs) of relative soil moisture or soil saturation encode sufficient information to determine these ecohydrological parameters. Further, these parameters can be estimated through inverse modeling of the analytical equation for soil saturation pdfs, derived from the commonly used stochastic soil water balance framework. We developed a generalizable Bayesian inference framework to estimate ecohydrological parameters consistent with empirical soil saturation pdfs derived from observations at point, footprint, and satellite scales. We applied the inference method to four sites with different land cover and climate assuming (i) an annual rainfall pattern and (ii) a wet season rainfall pattern with a dry season of negligible rainfall. The Nash-Sutcliffe efficiencies of the analytical model's fit to soil observations ranged from 0.89 to 0.99. The coefficient of variation of posterior parameter distributions ranged from < 1 to 15 %. The parameter identifiability was not significantly improved in the more complex seasonal model; however, small differences in parameter values indicate that the annual model may have absorbed dry season dynamics. Parameter estimates were most constrained for scales and locations at which soil water dynamics are more sensitive to the fitted ecohydrological parameters of interest. In these cases, model inversion converged more slowly but ultimately provided better goodness of fit and lower uncertainty. Results were robust using as few as 100 daily observations randomly sampled from the full records, demonstrating the advantage of analyzing soil saturation pdfs instead of time series to estimate ecohydrological parameters from sparse records. Our work combines modeling and empirical approaches in ecohydrology and provides a simple framework to obtain scale- and site-specific analytical descriptions of soil moisture dynamics consistent with soil moisture observations.

Infusion of donor-derived CD19-redirected virus-specific T cells for B-cell malignancies relapsed after allogeneic stem cell transplant: a phase 1 study

PubMed Central

Cruz, Conrad Russell Y.; Micklethwaite, Kenneth P.; Savoldo, Barbara; Ramos, Carlos A.; Lam, Sharon; Ku, Stephanie; Diouf, Oumar; Liu, Enli; Barrett, A. John; Ito, Sawa; Shpall, Elizabeth J.; Krance, Robert A.; Kamble, Rammurti T.; Carrum, George; Hosing, Chitra M.; Gee, Adrian P.; Mei, Zhuyong; Grilley, Bambi J.; Heslop, Helen E.; Rooney, Cliona M.; Brenner, Malcolm K.; Bollard, Catherine M.

2013-01-01

Autologous T cells expressing a CD19-specific chimeric antigen receptor (CD19.CAR) are active against B-cell malignancies, but it is unknown whether allogeneic CD19.CAR T cells are safe or effective. After allogeneic hematopoietic stem cell transplantation (HSCT), infused donor-derived virus-specific T cells (VSTs) expand in vivo, persist long term, and display antiviral activity without inducing graft-vs-host disease; therefore, we determined whether donor VSTs, engineered to express CD19.CAR, retained the characteristics of nonmanipulated allogeneic VSTs while gaining antitumor activity. We treated 8 patients with allogeneic (donor-derived) CD19.CAR-VSTs 3 months to 13 years after HSCT. There were no infusion-related toxicities. VSTs persisted for a median of 8 weeks in blood and up to 9 weeks at disease sites. Objective antitumor activity was evident in 2 of 6 patients with relapsed disease during the period of CD19.CAR-VST persistence, whereas 2 patients who received cells while in remission remain disease free. In 2 of 3 patients with viral reactivation, donor CD19.CAR-VSTs expanded concomitantly with VSTs. Hence CD19.CAR-VSTs display antitumor activity and, because their number may be increased in the presence of viral stimuli, earlier treatment post-HSCT (when lymphodepletion is greater and the incidence of viral infection is higher) or planned vaccination with viral antigens may enhance disease control. This study is registered at clinicaltrials.gov as #NCT00840853. PMID:24030379

Quantifying tolerance indicator values for common stream fish species of the United States

USGS Publications Warehouse

Meador, M.R.; Carlisle, D.M.

2007-01-01

The classification of fish species tolerance to environmental disturbance is often used as a means to assess ecosystem conditions. Its use, however, may be problematic because the approach to tolerance classification is based on subjective judgment. We analyzed fish and physicochemical data from 773 stream sites collected as part of the U.S. Geological Survey's National Water-Quality Assessment Program to calculate tolerance indicator values for 10 physicochemical variables using weighted averaging. Tolerance indicator values (TIVs) for ammonia, chloride, dissolved oxygen, nitrite plus nitrate, pH, phosphorus, specific conductance, sulfate, suspended sediment, and water temperature were calculated for 105 common fish species of the United States. Tolerance indicator values for specific conductance and sulfate were correlated (rho = 0.87), and thus, fish species may be co-tolerant to these water-quality variables. We integrated TIVs for each species into an overall tolerance classification for comparisons with judgment-based tolerance classifications. Principal components analysis indicated that the distinction between tolerant and intolerant classifications was determined largely by tolerance to suspended sediment, specific conductance, chloride, and total phosphorus. Factors such as water temperature, dissolved oxygen, and pH may not be as important in distinguishing between tolerant and intolerant classifications, but may help to segregate species classified as moderate. Empirically derived tolerance classifications were 58.8% in agreement with judgment-derived tolerance classifications. Canonical discriminant analysis revealed that few TIVs, primarily chloride, could discriminate among judgment-derived tolerance classifications of tolerant, moderate, and intolerant. To our knowledge, this is the first empirically based understanding of fish species tolerance for stream fishes in the United States.

Deciphering common recognition principles of nucleoside mono/di and tri-phosphates binding in diverse proteins via structural matching of their binding sites.

PubMed

Bhagavat, Raghu; Srinivasan, Narayanaswamy; Chandra, Nagasuma

2017-09-01

Nucleoside triphosphate (NTP) ligands are of high biological importance and are essential for all life forms. A pre-requisite for them to participate in diverse biochemical processes is their recognition by diverse proteins. It is thus of great interest to understand the basis for such recognition in different proteins. Towards this, we have used a structural bioinformatics approach and analyze structures of 4677 NTP complexes available in Protein Data Bank (PDB). Binding sites were extracted and compared exhaustively using PocketMatch, a sensitive in-house site comparison algorithm, which resulted in grouping the entire dataset into 27 site-types. Each of these site-types represent a structural motif comprised of two or more residue conservations, derived using another in-house tool for superposing binding sites, PocketAlign. The 27 site-types could be grouped further into 9 super-types by considering partial similarities in the sites, which indicated that the individual site-types comprise different combinations of one or more site features. A scan across PDB using the 27 structural motifs determined the motifs to be specific to NTP binding sites, and a computational alanine mutagenesis indicated that residues identified to be highly conserved in the motifs are also most contributing to binding. Alternate orientations of the ligand in several site-types were observed and rationalized, indicating the possibility of some residues serving as anchors for NTP recognition. The presence of multiple site-types and the grouping of multiple folds into each site-type is strongly suggestive of convergent evolution. Knowledge of determinants obtained from this study will be useful for detecting function in unknown proteins. Proteins 2017; 85:1699-1712. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

Sediment redistribution and grainsize effects on 230Th-normalized mass accumulation rates and focusing factors in the Panama Basin

NASA Astrophysics Data System (ADS)

Loveley, Matthew R.; Marcantonio, Franco; Lyle, Mitchell; Ibrahim, Rami; Hertzberg, Jennifer E.; Schmidt, Matthew W.

2017-12-01

Here, we examine how redistribution of differing grain sizes by sediment focusing processes in Panama Basin sediments affects the use of 230Th as a constant-flux proxy. We study representative sediments of Holocene and Last Glacial Maximum (LGM) time slices from four sediment cores from two different localities close to the ridges that bound the Panama Basin. Each locality contains paired sites that are seismically interpreted to have undergone extremes in sediment redistribution, i.e., focused versus winnowed sites. Both Holocene and LGM samples from sites where winnowing has occurred contain significant amounts (up to 50%) of the 230Th within the >63 μm grain size fraction, which makes up 40-70% of the bulk sediment analyzed. For sites where focusing has occurred, Holocene and LGM samples contain the greatest amounts of 230Th (up to 49%) in the finest grain-sized fraction (<4 μm), which makes up 26-40% of the bulk sediment analyzed. There are slight underestimations of 230Th-derived mass accumulation rates (MARs) and overestimations of 230Th-derived focusing factors at focused sites, while the opposite is true for winnowed sites. Corrections made using a model by Kretschmer et al. (2010) suggest a maximum change of about 30% in 230Th-derived MARs and focusing factors at focused sites, except for our most focused site which requires an approximate 70% correction in one sample. Our 230Th-corrected 232Th flux results suggest that the boundary between hemipelagically- and pelagically-derived sediments falls between 350 and 600 km from the continental margin.

Impact of the Location of CpG Methylation within the GSTP1 Gene on Its Specificity as a DNA Marker for Hepatocellular Carcinoma

PubMed Central

Jain, Surbhi; Boldbaatar, Batbold; Hamilton, James P.; Lin, Selena Y.; Chang, Ting-Tsung; Chen, Shun-Hua; Song, Wei; Meltzer, Stephen J.; Block, Timothy M.; Su, Ying-Hsiu

2012-01-01

Hypermethylation of the glutathione S-transferase π 1 (GSTP1) gene promoter region has been reported to be a potential biomarker to distinguish hepatocellular carcinoma (HCC) from other liver diseases. However, reports regarding how specific a marker it is have ranged from 100% to 0%. We hypothesized that, to a large extent, the variation of specificity depends on the location of the CpG sites analyzed. To test this hypothesis, we compared the methylation status of the GSTP1 promoter region of the DNA isolated from HCC, cirrhosis, hepatitis, and normal liver tissues by bisulfite–PCR sequencing. We found that the 5′ region of the position −48 nt from the transcription start site of the GSTP1 gene is selectively methylated in HCC, whereas the 3′ region is methylated in all liver tissues examined, including normal liver and the HCC tissue. Interestingly, when DNA derived from fetal liver and 11 nonhepatic normal tissue was also examined by bisulfite-PCR sequencing, we found that methylation of the 3′ region of the promoter appeared to be liver-specific. A methylation-specific PCR assay targeting the 5′ region of the promoter was developed and used to quantify the methylated GSTP1 gene in various diseased liver tissues including HCC. When we used an assay targeting the 3′ region, we found that the methylation of the 5′-end of the GSTP1 promoter was significantly more specific than that of the 3′-end (97.1% vs. 60%, p<0.0001 by Fisher's exact test) for distinguishing HCC (n = 120) from hepatitis (n = 35) and cirrhosis (n = 35). Encouragingly, 33.8% of the AFP-negative HCC contained the methylated GSTP1 gene. This study clearly demonstrates the importance of the location of CpG site methylation for HCC specificity and how liver-specific DNA methylation should be considered when an epigenetic DNA marker is studied for detection of HCC. PMID:22536438

Neutrophil trails guide influenza-specific CD8+ T cells in the airways

PubMed Central

Lim, Kihong; Hyun, Young-Min; Lambert-Emo, Kris; Capece, Tara; Bae, Seyeon; Miller, Richard; Topham, David J.; Kim, Minsoo

2016-01-01

During viral infections, chemokines guide activated effector T cells to infection sites. However, the cells responsible for producing these chemokines and how such chemokines recruit T cells is unknown. Here, we show that the early recruitment of neutrophils into influenza-infected trachea is essential for CD8+ T cell-mediated immune protection in mice. We observed that migrating neutrophils leave behind long-lasting trails that are enriched in the chemokine CXCL12. Experiments with granulocyte-specific CXCL12 conditional knock-out mice and a CXCR4 antagonist revealed that CXCL12 derived from neutrophil trails is critical for virus-specific CD8+ T cell recruitment and effector functions. Collectively, these results suggest neutrophils deposit long-lasting, chemokine-containing trails, which may provide both chemotactic and haptotactic cues for efficient CD8+ T cell migration and localization in influenza-infected tissues. PMID:26339033

Neutrophil trails guide influenza-specific CD8⁺ T cells in the airways.

PubMed

Lim, Kihong; Hyun, Young-Min; Lambert-Emo, Kris; Capece, Tara; Bae, Seyeon; Miller, Richard; Topham, David J; Kim, Minsoo

2015-09-04

During viral infections, chemokines guide activated effector T cells to infection sites. However, the cells responsible for producing these chemokines and how such chemokines recruit T cells are unknown. Here, we show that the early recruitment of neutrophils into influenza-infected trachea is essential for CD8(+) T cell-mediated immune protection in mice. We observed that migrating neutrophils leave behind long-lasting trails that are enriched in the chemokine CXCL12. Experiments with granulocyte-specific CXCL12 conditionally depleted mice and a CXCR4 antagonist revealed that CXCL12 derived from neutrophil trails is critical for virus-specific CD8(+) T cell recruitment and effector functions. Collectively, these results suggest that neutrophils deposit long-lasting, chemokine-containing trails, which may provide both chemotactic and haptotactic cues for efficient CD8(+) T cell migration and localization in influenza-infected tissues. Copyright © 2015, American Association for the Advancement of Science.

Chemical probing of the human sirtuin 5 active site reveals its substrate acyl specificity and peptide-based inhibitors.

PubMed

Roessler, Claudia; Nowak, Theresa; Pannek, Martin; Gertz, Melanie; Nguyen, Giang T T; Scharfe, Michael; Born, Ilona; Sippl, Wolfgang; Steegborn, Clemens; Schutkowski, Mike

2014-09-26

Sirtuins are NAD(+)-dependent deacetylases acting as sensors in metabolic pathways and stress response. In mammals there are seven isoforms. The mitochondrial sirtuin 5 is a weak deacetylase but a very efficient demalonylase and desuccinylase; however, its substrate acyl specificity has not been systematically analyzed. Herein, we investigated a carbamoyl phosphate synthetase 1 derived peptide substrate and modified the lysine side chain systematically to determine the acyl specificity of Sirt5. From that point we designed six potent peptide-based inhibitors that interact with the NAD(+) binding pocket. To characterize the interaction details causing the different substrate and inhibition properties we report several X-ray crystal structures of Sirt5 complexed with these peptides. Our results reveal the Sirt5 acyl selectivity and its molecular basis and enable the design of inhibitors for Sirt5. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cellular and molecular mechanisms of HIV-1 integration targeting.

PubMed

Engelman, Alan N; Singh, Parmit K

2018-07-01

Integration is central to HIV-1 replication and helps mold the reservoir of cells that persists in AIDS patients. HIV-1 interacts with specific cellular factors to target integration to interior regions of transcriptionally active genes within gene-dense regions of chromatin. The viral capsid interacts with several proteins that are additionally implicated in virus nuclear import, including cleavage and polyadenylation specificity factor 6, to suppress integration into heterochromatin. The viral integrase protein interacts with transcriptional co-activator lens epithelium-derived growth factor p75 to principally position integration within gene bodies. The integrase additionally senses target DNA distortion and nucleotide sequence to help fine-tune the specific phosphodiester bonds that are cleaved at integration sites. Research into virus-host interactions that underlie HIV-1 integration targeting has aided the development of a novel class of integrase inhibitors and may help to improve the safety of viral-based gene therapy vectors.

Bacteriophage recombination systems and biotechnical applications.

PubMed

Nafissi, Nafiseh; Slavcev, Roderick

2014-04-01

Bacteriophage recombination systems have been widely used in biotechnology for modifying prokaryotic species, for creating transgenic animals and plants, and more recently, for human cell gene manipulation. In contrast to homologous recombination, which benefits from the endogenous recombination machinery of the cell, site-specific recombination requires an exogenous source of recombinase in mammalian cells. The mechanism of bacteriophage evolution and their coexistence with bacterial cells has become a point of interest ever since bacterial viruses' life cycles were first explored. Phage recombinases have already been exploited as valuable genetic tools and new phage enzymes, and their potential application to genetic engineering and genome manipulation, vectorology, and generation of new transgene delivery vectors, and cell therapy are attractive areas of research that continue to be investigated. The significance and role of phage recombination systems in biotechnology is reviewed in this paper, with specific focus on homologous and site-specific recombination conferred by the coli phages, λ, and N15, the integrase from the Streptomyces phage, ΦC31, the recombination system of phage P1, and the recently characterized recombination functions of Yersinia phage, PY54. Key steps of the molecular mechanisms involving phage recombination functions and their application to molecular engineering, our novel exploitations of the PY54-derived recombination system, and its application to the development of new DNA vectors are discussed.

Reclamation of mined lands in the western coal region

USGS Publications Warehouse

Narten, Perry F.; Litner, S.F.; Allingham, J.W.; Foster, Lee; Larsen, D.M.; McWreath, H.C.

1983-01-01

In 1978, a group of scientists from several Federal agencies examined reclamation work at 22 coal mines in seven western States. The results of these examinations were not used to derive quantitative predictions of the outcome of reclamation work but rather to determine the general requirements for revegetation success. Locally, reclamation efforts are affected by climate, especially precipitation; the landform of the restored surface; the nature of the overburden material; the nature of the surface soil; and the natural ecological system. The goals of reclamation efforts are now broader than ever. Regulations call not only for reducing the steepness of the final surface and establishing a cover of mostly perennial native vegetation, but for restoring the land for specific land uses, achieving diversity both in types of plants and in number of species, and reintroduction of biological and ecological processes. If specific sites are monitored over a long enough period of time, quantitative predictions of success for individual mines may be possible, and such predictions can be included in environmental impact statements to help in the decision-making process. The results of this study indicate that current reclamation objectives can be met when the reclamation effort is designed on the basis of site-specific needs and when existing technology is used.

Hydrofluoric Acid-Based Derivatization Strategy To Profile PARP-1 ADP-Ribosylation by LC-MS/MS.

PubMed

Gagné, Jean-Philippe; Langelier, Marie-France; Pascal, John M; Poirier, Guy G

2018-06-11

Despite significant advances in the development of mass spectrometry-based methods for the identification of protein ADP-ribosylation, current protocols suffer from several drawbacks that preclude their widespread applicability. Given the intrinsic heterogeneous nature of poly(ADP-ribose), a number of strategies have been developed to generate simple derivatives for effective interrogation of protein databases and site-specific localization of the modified residues. Currently, the generation of spectral signatures indicative of ADP-ribosylation rely on chemical or enzymatic conversion of the modification to a single mass increment. Still, limitations arise from the lability of the poly(ADP-ribose) remnant during tandem mass spectrometry, the varying susceptibilities of different ADP-ribose-protein bonds to chemical hydrolysis, or the context dependence of enzyme-catalyzed reactions. Here, we present a chemical-based derivatization method applicable to the confident identification of site-specific ADP-ribosylation by conventional mass spectrometry on any targeted amino acid residue. Using PARP-1 as a model protein, we report that treatment of ADP-ribosylated peptides with hydrofluoric acid generates a specific +132 Da mass signature that corresponds to the decomposition of mono- and poly(ADP-ribosylated) peptides into ribose adducts as a consequence of the cleavage of the phosphorus-oxygen bonds.

Emission factors for hydraulically fractured gas wells derived using well- and battery-level reported data for Alberta, Canada.

PubMed

Tyner, David R; Johnson, Matthew R

2014-12-16

A comprehensive technical analysis of available industry-reported well activity and production data for Alberta in 2011 has been used to derive flaring, venting, and diesel combustion greenhouse gas and criteria air contaminant emission factors specifically linked to drilling, completion, and operation of hydraulically fractured natural gas wells. Analysis revealed that in-line ("green") completions were used at approximately 53% of wells completed in 2011, and in other cases the majority (99.5%) of flowback gases were flared rather than vented. Comparisons with limited analogous data available in the literature revealed that reported total flared and vented natural gas volumes attributable to tight gas well-completions were ∼ 6 times larger than Canadian Association of Petroleum Producers (CAPP) estimates for natural gas well-completion based on wells ca. 2000, but 62% less than an equivalent emission factor that can be derived from U.S. EPA data. Newly derived emission factors for diesel combustion during well drilling and completion are thought to be among the first such data available in the open literature, where drilling-related emissions for tight gas wells drilled in Alberta in 2011 were found to have increased by a factor of 2.8 relative to a typical well drilled in Canada in 2000 due to increased drilling lengths. From well-by-well analysis of production phase flared, vented, and fuel usage natural gas volumes reported at 3846 operating tight gas wells in 2011, operational emission factors were developed. Overall results highlight the importance of operational phase GHG emissions at upstream well sites (including on-site natural gas fuel use), and the critical levels of uncertainty in current estimates of liquid unloading emissions.

Hypochlorite-induced damage to DNA, RNA, and polynucleotides: formation of chloramines and nitrogen-centered radicals.

PubMed

Hawkins, Clare L; Davies, Michael J

2002-01-01

Stimulated monocytes and neutrophils generate hypochlorite (HOCl) via the release of the enzyme myeloperoxidase and hydrogen peroxide. HOCl is a key bactericidal agent, but can also damage host tissue. As there is a strong link between chronic inflammation and some cancers, we have investigated HOCl damage to DNA, RNA, and polynucleotides. Reaction of HOCl with these materials is shown to yield multiple semistable chloramines (RNHCl/RR'NCl), which are the major initial products, and account for 50-95% of the added HOCl. These chloramines decay by thermal and metal-ion catalyzed processes, to give nucleoside-derived, nitrogen-centered, radicals. The latter have been characterized by EPR spin trapping. The propensity for radical formation with polynucleotides is cytidine > adenosine = guanosine > uridine = thymidine. The rates of decay, and yield of radicals formed, are dependent on the nature of the nucleobase on which they are formed, with chloramines formed from ring heterocyclic amine groups being less stable than those formed on exocyclic amines (RNH2 groups). Evidence is presented for chlorine transfer from the former, kinetically favored, sites to the more thermodynamically favored exocyclic amines. EPR experiments have also provided evidence for the rapid addition of pyrimidine-derived nitrogen-centered radicals to other nucleobases to give dimers and the oxidation of DNA by radicals derived from preformed nucleoside chloramines. Direct reaction of HOCl with plasmid DNA gives rise to single- and double-strand breaks via chloramine-mediated reactions. Preformed nucleoside chloramines also induce plasmid cleavage, though this only occurs to a significant extent with unstable thymidine- and uridine-derived chloramines, where radical formation is rapid. Overall the data rationalize the preferential formation of chlorinated 2'-deoxycytidine and 2'-deoxyadenosine in DNA and suggest that DNA damage induced by HOCl, and preformed chloramines, occurs at sequence-specific sites.

Arabidopsis ARC6 coordinates the division machineries of the inner and outer chloroplast membranes through interaction with PDV2 in the intermembrane space.

PubMed

Glynn, Jonathan M; Froehlich, John E; Osteryoung, Katherine W

2008-09-01

Chloroplasts arose from a free-living cyanobacterial endosymbiont and divide by binary fission. Division involves the assembly and constriction of the endosymbiont-derived, tubulin-like FtsZ ring on the stromal surface of the inner envelope membrane and the host-derived, dynamin-like ARC5 ring on the cytosolic surface of the outer envelope membrane. Despite the identification of many proteins required for plastid division, the factors coordinating the internal and external division machineries are unknown. Here, we provide evidence that this coordination is mediated in Arabidopsis thaliana by an interaction between ARC6, an FtsZ assembly factor spanning the inner envelope membrane, and PDV2, an ARC5 recruitment factor spanning the outer envelope membrane. ARC6 and PDV2 interact via their C-terminal domains in the intermembrane space, consistent with their in vivo topologies. ARC6 acts upstream of PDV2 to localize PDV2 (and hence ARC5) to the division site. We present a model whereby ARC6 relays information on stromal FtsZ ring positioning through PDV2 to the chloroplast surface to specify the site of ARC5 recruitment. Because orthologs of ARC6 occur in land plants, green algae, and cyanobacteria but PDV2 occurs only in land plants, the connection between ARC6 and PDV2 represents the evolution of a plant-specific adaptation to coordinate the assembly and activity of the endosymbiont- and host-derived plastid division components.

Use of passive ambient ozone (O3) samplers in vegetation effects assessment

USGS Publications Warehouse

Krupa, S.; Nosal, M.; Peterson, D.L.

2001-01-01

A stochastistic, Weibull probability model was developed and verified to simulate the underlying frequency distributions of hourly ozone (O3) concentrations (exposure dynamics) using the single, weekly mean values obtained from a passive (sodium nitrite absorbent) sampler. The simulation was based on the data derived from a co-located continuous monitor. Although at the moment the model output may be considered as being specific to the elevation and location of the study site, the results were extremely good. This effort for the approximation of the O3 exposure dynamics can be extended to other sites with similar data sets and in developing a generalized understanding of the stochastic O3 exposure-plant response relationships, conferring measurable benefits to the future use of passive O3 samplers, in the absence of continuous monitoring. Copyright ?? 2000 Elsevier Science Ltd.

A Comprehensive Systems Biology Approach to Studying Zika Virus.

PubMed

May, Meghan; Relich, Ryan F

2016-01-01

Zika virus (ZIKV) is responsible for an ongoing and intensifying epidemic in the Western Hemisphere. We examined the complete predicted proteomes, glycomes, and selectomes of 33 ZIKV strains representing temporally diverse members of the African lineage, the Asian lineage, and the current outbreak in the Americas. Derivation of the complete selectome is an 'omics' approach to identify distinct evolutionary pressures acting on different features of an organism. Employment of the M8 model did not show evidence of global diversifying selection acting on the ZIKV polyprotein; however, a mixed effect model of evolution showed strong evidence (P<0.05) for episodic diversifying selection acting on specific sites. Single nucleotide polymorphisms (SNPs) were predictably frequent across strains relative to the derived consensus sequence. None of the 9 published detection procedures utilize targets that share 100% identity across the 33 strains examined, indicating that ZIKV escape from molecular detection is predictable. The predicted O-linked glycome showed marked diversity across strains; however, the N-linked glycome was highly stable. All Asian and American strains examined were predicted to include glycosylation of E protein ASN154, a modification proposed to mediate neurotropism, whereas the modification was not predicted for African strains. SNP diversity, episodic diversifying selection, and differential glycosylation, particularly of ASN154, may have major biological implications for ZIKV disease. Taken together, the systems biology perspective of ZIKV indicates: a.) The recently emergent Asian/American N-glycotype is mediating the new and emerging neuropathogenic potential of ZIKV; and b.) further divergence at specific sites is predictable as endemnicity is established in the Americas.

A novel mobile dual-wavelength laser altimetry system for improved site-specific Nitrogen fertilizer applications

NASA Astrophysics Data System (ADS)

Eitel, J.; Magney, T. S.; Vierling, L. A.; Brown, T. T.; Huggins, D. R.

2012-12-01

Reducing fertilizer inputs while maintaining yield would increase farmer's profits and similarly lessen the adverse environmental effects of production agriculture. The development of technologies that allow precise, site-specific application of Nitrogen (N) fertilizer has thus been an important research goal over the past decades. Remote sensing of foliar crop properties and function with tractor-mountable optical sensors has thought to be useful to optimize N fertilizer applications. However, on-the-go sensing of foliar crop properties and function has proven difficult, particularly during early crop growth stages when fertilizer decisions are often made. This difficulty arises from the fact that the spectral signal measured by on-the-go sensors is dominated by soil reflectance during early crop growth stages. Here, we present the basic principles behind a novel, dual-wavelength, tractor mountable laser altimetry system that measures the laser return intensity of the reflected green and red laser light. The green (532 nm) and the red (660 nm) wavelength combination allows calculation of a modified Photochemical Reflectance Index (mPRI) that have shown to be sensitive to both crop function and foliar chemistry. The small field of view of the laser points (diameter: 4 mm) combined with its high sampling rate (1000 points sec-1) allows vegetation returns to be isolated from ground returns by using simple thresholds. First tests relating foliar N of winter wheat (Triticum aestivum L.) with laser derived mPRI are promising (r2 = 0.72). Further research is needed to test the relationship between laser derived spectral indices and crop function.

Identification of the platelet-derived chemokine CXCL4/PF-4 as a broad-spectrum HIV-1 inhibitor

PubMed Central

Auerbach, David J.; Lin, Yin; Miao, Huiyi; Cimbro, Raffaello; DiFiore, Michelle J.; Gianolini, Monica E.; Furci, Lucinda; Biswas, Priscilla; Fauci, Anthony S.; Lusso, Paolo

2012-01-01

The natural history of HIV-1 infection is highly variable in different individuals, spanning from a rapidly progressive course to a long-term asymptomatic infection. A major determinant of the pace of disease progression is the in vivo level of HIV-1 replication, which is regulated by a complex network of cytokines and chemokines expressed by immune and inflammatory cells. The chemokine system is critically involved in the control of HIV-1 replication by virtue of the role played by specific chemokine receptors, most notably CCR5 and CXCR4, as cell-surface coreceptors for HIV-1 entry; hence, the chemokines that naturally bind such coreceptors act as endogenous inhibitors of HIV-1. Here, we show that the CXC chemokine CXCL4 (PF-4), the most abundant protein contained within the α-granules of platelets, is a broad-spectrum inhibitor of HIV-1 infection. Unlike other known HIV-suppressive chemokines, CXCL4 inhibits infection by the majority of primary HIV-1 isolates regardless of their coreceptor-usage phenotype or genetic subtype. Consistent with the lack of viral phenotype specificity, blockade of HIV-1 infection occurs at the level of virus attachment and entry via a unique mechanism that involves direct interaction of CXCL4 with the major viral envelope glycoprotein, gp120. The binding site for CXCL4 was mapped to a region of the gp120 outer domain proximal to the CD4-binding site. The identification of a platelet-derived chemokine as an endogenous antiviral factor may have relevance for the pathogenesis and treatment of HIV-1 infection. PMID:22645343

Identification of the platelet-derived chemokine CXCL4/PF-4 as a broad-spectrum HIV-1 inhibitor.

PubMed

Auerbach, David J; Lin, Yin; Miao, Huiyi; Cimbro, Raffaello; Difiore, Michelle J; Gianolini, Monica E; Furci, Lucinda; Biswas, Priscilla; Fauci, Anthony S; Lusso, Paolo

2012-06-12

The natural history of HIV-1 infection is highly variable in different individuals, spanning from a rapidly progressive course to a long-term asymptomatic infection. A major determinant of the pace of disease progression is the in vivo level of HIV-1 replication, which is regulated by a complex network of cytokines and chemokines expressed by immune and inflammatory cells. The chemokine system is critically involved in the control of HIV-1 replication by virtue of the role played by specific chemokine receptors, most notably CCR5 and CXCR4, as cell-surface coreceptors for HIV-1 entry; hence, the chemokines that naturally bind such coreceptors act as endogenous inhibitors of HIV-1. Here, we show that the CXC chemokine CXCL4 (PF-4), the most abundant protein contained within the α-granules of platelets, is a broad-spectrum inhibitor of HIV-1 infection. Unlike other known HIV-suppressive chemokines, CXCL4 inhibits infection by the majority of primary HIV-1 isolates regardless of their coreceptor-usage phenotype or genetic subtype. Consistent with the lack of viral phenotype specificity, blockade of HIV-1 infection occurs at the level of virus attachment and entry via a unique mechanism that involves direct interaction of CXCL4 with the major viral envelope glycoprotein, gp120. The binding site for CXCL4 was mapped to a region of the gp120 outer domain proximal to the CD4-binding site. The identification of a platelet-derived chemokine as an endogenous antiviral factor may have relevance for the pathogenesis and treatment of HIV-1 infection.

Dissociation free-energy profiles of specific and nonspecific DNA-protein complexes.

PubMed

Yonetani, Yoshiteru; Kono, Hidetoshi

2013-06-27

DNA-binding proteins recognize DNA sequences with at least two different binding modes: specific and nonspecific. Experimental structures of such complexes provide us a static view of the bindings. However, it is difficult to reveal further mechanisms of their target-site search and recognition only from static information because the transition process between the bound and unbound states is not clarified by static information. What is the difference between specific and nonspecific bindings? Here we performed adaptive biasing force molecular dynamics simulations with the specific and nonspecific structures of DNA-Lac repressor complexes to investigate the dissociation process. The resultant free-energy profiles showed that the specific complex has a sharp, deep well consistent with tight binding, whereas the nonspecific complex has a broad, shallow well consistent with loose binding. The difference in the well depth, ~5 kcal/mol, was in fair agreement with the experimentally obtained value and was found to mainly come from the protein conformational difference, particularly in the C-terminal tail. Also, the free-energy profiles were found to be correlated with changes in the number of protein-DNA contacts and that of surface water molecules. The derived protein spatial distributions around the DNA indicate that any large dissociation occurs rarely, regardless of the specific and nonspecific sites. Comparison of the free-energy barrier for sliding [~8.7 kcal/mol; Furini J. Phys. Chem. B 2010, 114, 2238] and that for dissociation (at least ~16 kcal/mol) calculated in this study suggests that sliding is much preferred to dissociation.

Physics-Based Hazard Assessment for Critical Structures Near Large Earthquake Sources

NASA Astrophysics Data System (ADS)

Hutchings, L.; Mert, A.; Fahjan, Y.; Novikova, T.; Golara, A.; Miah, M.; Fergany, E.; Foxall, W.

2017-09-01

We argue that for critical structures near large earthquake sources: (1) the ergodic assumption, recent history, and simplified descriptions of the hazard are not appropriate to rely on for earthquake ground motion prediction and can lead to a mis-estimation of the hazard and risk to structures; (2) a physics-based approach can address these issues; (3) a physics-based source model must be provided to generate realistic phasing effects from finite rupture and model near-source ground motion correctly; (4) wave propagations and site response should be site specific; (5) a much wider search of possible sources of ground motion can be achieved computationally with a physics-based approach; (6) unless one utilizes a physics-based approach, the hazard and risk to structures has unknown uncertainties; (7) uncertainties can be reduced with a physics-based approach, but not with an ergodic approach; (8) computational power and computer codes have advanced to the point that risk to structures can be calculated directly from source and site-specific ground motions. Spanning the variability of potential ground motion in a predictive situation is especially difficult for near-source areas, but that is the distance at which the hazard is the greatest. The basis of a "physical-based" approach is ground-motion syntheses derived from physics and an understanding of the earthquake process. This is an overview paper and results from previous studies are used to make the case for these conclusions. Our premise is that 50 years of strong motion records is insufficient to capture all possible ranges of site and propagation path conditions, rupture processes, and spatial geometric relationships between source and site. Predicting future earthquake scenarios is necessary; models that have little or no physical basis but have been tested and adjusted to fit available observations can only "predict" what happened in the past, which should be considered description as opposed to prediction. We have developed a methodology for synthesizing physics-based broadband ground motion that incorporates the effects of realistic earthquake rupture along specific faults and the actual geology between the source and site.

Temporal change in biological community structure in the Fountain Creek basin, Colorado, 2001-2008

USGS Publications Warehouse

Zuellig, Robert E.; Bruce, James F.; Stogner, Sr., Robert W.

2010-01-01

In 2001, the U.S. Geological Survey, in cooperation with Colorado Springs City Engineering, began a study to better understand the relations between environmental characteristics and biological communities in the Fountain Creek basin in order to aide water-resource management and guide future monitoring activities. To accomplish this task, environmental (streamflow, habitat, and water chemistry) and biological (fish and macroinvertebrate) data were collected annually at 24 sites over a 6- or 8-year period (fish, 2003 to 2008; macroinvertebrates, 2001 to 2008). For this report, these data were first analyzed to determine the presence of temporal change in macroinvertebrate and fish community structure among years using nonparametric multivariate statistics. Where temporal change in the biological communities was found, these data were further analyzed using additional nonparametric multivariate techniques to determine which subset of selected streamflow, habitat, or water-chemistry variables best described site-specific changes in community structure relative to a gradient of urbanization. This study identified significant directional patterns of temporal change in macroinvertebrate and fish community structure at 15 of 24 sites in the Fountain Creek basin. At four of these sites, changes in environmental variables were significantly correlated with the concurrent temporal change identified in macroinvertebrate and fish community structure (Monument Creek above Woodmen Road at Colorado Springs, Colo.; Monument Creek at Bijou Street at Colorado Springs, Colo.; Bear Creek near Colorado Springs, Colo.; Fountain Creek at Security, Colo.). Combinations of environmental variables describing directional temporal change in the biota appeared to be site specific as no single variable dominated the results; however, substrate composition variables (percent substrate composition composed of sand, gravel, or cobble) collectively were present in 80 percent of the environmental variable subsets that were significantly correlated with temporal change in the macroinvertebrate and fish community structure. Other important environmental variables related to temporal change in the biological community structure included those describing channel form (streambank height) and streamflow (normalized annual mean daily flow, high flood-pulse count). Site-specific results from this study were derived from a relatively small number of observations (6 or 8 years of data); therefore, additional years of data may reveal other sites with temporal change in biological community structure, or could define stronger and more consistent linkages between environmental variables and observed temporal change. Likewise current variable subsets could become weaker. Nonetheless, there were several sites where temporal change was detected in this study that could not be explained by the available environmental variables studied herein. Modification of current data-collection activities may be necessary to better understand site-specific temporal relations between biological communities and environmental variables.

NDVI derived from IR-enabled digital cameras: applicability across different plant functional types

NASA Astrophysics Data System (ADS)

Filippa, Gianluca; Cremonese, Edoardo; Galvagno, Marta; Migliavacca, Mirco; Sonnentag, Oliver; Hufkens, Koen; Ryu, Youngryel; Humphreys, Elyn; Morra di Cella, Umberto; Richardson, Andrew D.

2017-04-01

Phenological time-series based on the deployment of radiometric measurements are now being constructed at different spatial and temporal scales ranging from weekly satellite observations to sub-hourly in situ measurements by means of e.g. radiometers or digital cameras. In situ measurements are strongly required to provide high-frequency validation data for satellite-derived vegetation indices. In this study we used a recently developed method to calculate NDVI from NIR-enabled digital cameras (NDVIC) at 17 sites encompassing 6 plant functional types and totalizing 74 year-sites of data from the PHENOCAM network. The seasonality of NDVIC was comparable to both NDVI measured by ground light emitting diode (LED) sensors and by MODIS, whereas site-specific scaling factors are required to compare absolute values of NDVIC to standard NDVI measurements. We also compared green chromatic coordinate (GCC) extracted from RGB-only images to NDVIC and found that the two are characterized by slight different dynamics, dependent on the plant functional type. During senescence, NDVIC lags behind GCC in deciduous broad-leaf forests and grasslands, suggesting that GCC is more sensitive to leaf decoloration and NDVIC to the biomass reduction resulting from leaf abscission and green to dry biomass ratio of the canopy. In evergreen forests, NDVIC peaks later than GCC in spring, likely tracking the processes of shoot elongation and new needle formation. Our findings suggest therefore that NDVIC and GCC can complement each other in describing ecosystem phenology.

DNA Inversion on Conjugative Plasmid pVT745

PubMed Central

Chen, Jinbiao; Leblanc, Donald J.; Galli, Dominique M.

2002-01-01

Plasmid pVT745 from Actinobacillus actinomycetemcomitans strain VT745 can be transferred to other A. actinomycetemcomitans strains at a frequency of 10−6. Screening of transconjugants revealed that the DNA of pDMG21A, a pVT745 derivative containing a kanamycin resistance gene, displayed a structural rearrangement after transfer. A 9-kb segment on the plasmid had switched orientation. The inversion was independent of RecA and required the activity of the pVT745-encoded site-specific recombinase. This recombinase, termed Inv, was highly homologous to invertases of the Din family. Two recombination sites of 22 bp, which are arranged in opposite orientation and which function as DNA crossover sequences, were identified on pVT745. One of the sites was located adjacent to the 5′ end of the invertase gene, inv. Inversion of the 9-kb segment on pVT745 derivatives has been observed in all A. actinomycetemcomitans strains tested except for the original host, VT745. This would suggest that a host factor that is either inactive or absent in VT745 is required for efficient recombination. Inactivation of the invertase in the donor strain resulted in a 1,000-fold increase in the number of transconjugants upon plasmid transfer. It is proposed that an activated invertase causes the immediate loss of the plasmid in most recipient cells after mating. No biological role has been associated with the invertase as of yet. PMID:12374826

Polycistronic lentiviral vector for "hit and run" reprogramming of adult skin fibroblasts to induced pluripotent stem cells.

PubMed

Chang, Chia-Wei; Lai, Yi-Shin; Pawlik, Kevin M; Liu, Kaimao; Sun, Chiao-Wang; Li, Chao; Schoeb, Trenton R; Townes, Tim M

2009-05-01

We report the derivation of induced pluripotent stem (iPS) cells from adult skin fibroblasts using a single, polycistronic lentiviral vector encoding the reprogramming factors Oct4, Sox2, and Klf4. Porcine teschovirus-1 2A sequences that trigger ribosome skipping were inserted between human cDNAs for these factors, and the polycistron was subcloned downstream of the elongation factor 1 alpha promoter in a self-inactivating (SIN) lentiviral vector containing a loxP site in the truncated 3' long terminal repeat (LTR). Adult skin fibroblasts from a humanized mouse model of sickle cell disease were transduced with this single lentiviral vector, and iPS cell colonies were picked within 30 days. These cells expressed endogenous Oct4, Sox2, Nanog, alkaline phosphatase, stage-specific embryonic antigen-1, and other markers of pluripotency. The iPS cells produced teratomas containing tissue derived from all three germ layers after injection into immunocompromised mice and formed high-level chimeras after injection into murine blastocysts. iPS cell lines with as few as three lentiviral insertions were obtained. Expression of Cre recombinase in these iPS cells resulted in deletion of the lentiviral vector, and sequencing of insertion sites demonstrated that remnant 291-bp SIN LTRs containing a single loxP site did not interrupt coding sequences, promoters, or known regulatory elements. These results suggest that a single, polycistronic "hit and run" vector can safely and effectively reprogram adult dermal fibroblasts into iPS cells.