Sample records for chain variable region
-
Sequences of heavy and light chain variable regions from four bovine immunoglobulins.
Armour, K L; Tempest, P R; Fawcett, P H; Fernie, M L; King, S I; White, P; Taylor, G; Harris, W J
1994-12-01
Oligodeoxyribonucleotide primers based on the 5' ends of bovine IgG1/2 and lambda constant (C) region genes, together with primers encoding conserved amino acids at the N-terminus of mature variable (V) regions from other species, have been used in cDNA and polymerase chain reactions (PCRs) to amplify heavy and light chain V region cDNA from bovine heterohybridomas. The amino acid sequences of VH and V lambda from four bovine immunoglobulins of different specificities are presented.
-
DISTINCT ANTIBODY SPECIES: STRUCTURAL DIFFERENCES CREATING THERAPEUTIC OPPORTUNITIES
Muyldermans, Serge; Smider, Vaughn V.
2016-01-01
Antibodies have been a remarkably successful class of molecules for binding a large number of antigens in therapeutic, diagnostic, and research applications. Typical antibodies derived from mouse or human sources use the surface formed by complementarity determining regions (CDRs) on the variable regions of the heavy chain/light chain heterodimer, which typically forms a relatively flat binding surface. Alternative species, particularly camelids and bovines, provide a unique paradigm for antigen recognition through novel domains which form the antigen binding paratope. For camelids, heavy chain antibodies bind antigen with only a single heavy chain variable region, in the absence of light chains. In bovines, ultralong CDR-H3 regions form an independently folding minidomain, which protrudes from the surface of the antibody and is diverse in both its sequence and disulfide patterns. The atypical paratopes of camelids and bovines potentially provide the ability to interact with different epitopes, particularly recessed or concave surfaces, compared to traditional antibodies. PMID:26922135
-
Townsend, Catherine L; Laffy, Julie M J; Wu, Yu-Chang Bryan; Silva O'Hare, Joselli; Martin, Victoria; Kipling, David; Fraternali, Franca; Dunn-Walters, Deborah K
2016-01-01
Antibody variable regions are composed of a heavy and a light chain, and in humans, there are two light chain isotypes: kappa and lambda. Despite their importance in receptor editing, the light chain is often overlooked in the antibody literature, with the focus being on the heavy chain complementarity-determining region (CDR)-H3 region. In this paper, we set out to investigate the physicochemical and structural differences between human kappa and lambda light chain CDR regions. We constructed a dataset containing over 29,000 light chain variable region sequences from IgM-transcribing, newly formed B cells isolated from human bone marrow and peripheral blood. We also used a published human naïve dataset to investigate the CDR-H3 properties of heavy chains paired with kappa and lambda light chains and probed the Protein Data Bank to investigate the structural differences between kappa and lambda antibody CDR regions. We found that kappa and lambda light chains have very different CDR physicochemical and structural properties, whereas the heavy chains with which they are paired do not differ significantly. We also observed that the mean CDR3 N nucleotide addition in the kappa, lambda, and heavy chain gene rearrangements are correlated within donors but can differ between donors. This indicates that terminal deoxynucleotidyl transferase may work with differing efficiencies between different people but the same efficiency in the different classes of immunoglobulin chain within one person. We have observed large differences in the physicochemical and structural properties of kappa and lambda light chain CDR regions. This may reflect different roles in the humoral immune response.
-
Townsend, Catherine L.; Laffy, Julie M. J.; Wu, Yu-Chang Bryan; Silva O’Hare, Joselli; Martin, Victoria; Kipling, David; Fraternali, Franca; Dunn-Walters, Deborah K.
2016-01-01
Antibody variable regions are composed of a heavy and a light chain, and in humans, there are two light chain isotypes: kappa and lambda. Despite their importance in receptor editing, the light chain is often overlooked in the antibody literature, with the focus being on the heavy chain complementarity-determining region (CDR)-H3 region. In this paper, we set out to investigate the physicochemical and structural differences between human kappa and lambda light chain CDR regions. We constructed a dataset containing over 29,000 light chain variable region sequences from IgM-transcribing, newly formed B cells isolated from human bone marrow and peripheral blood. We also used a published human naïve dataset to investigate the CDR-H3 properties of heavy chains paired with kappa and lambda light chains and probed the Protein Data Bank to investigate the structural differences between kappa and lambda antibody CDR regions. We found that kappa and lambda light chains have very different CDR physicochemical and structural properties, whereas the heavy chains with which they are paired do not differ significantly. We also observed that the mean CDR3 N nucleotide addition in the kappa, lambda, and heavy chain gene rearrangements are correlated within donors but can differ between donors. This indicates that terminal deoxynucleotidyl transferase may work with differing efficiencies between different people but the same efficiency in the different classes of immunoglobulin chain within one person. We have observed large differences in the physicochemical and structural properties of kappa and lambda light chain CDR regions. This may reflect different roles in the humoral immune response. PMID:27729912
-
Dialynas, D P; Murre, C; Quertermous, T; Boss, J M; Leiden, J M; Seidman, J G; Strominger, J L
1986-01-01
Complementary DNA (cDNA) encoding a human T-cell gamma chain has been cloned and sequenced. At the junction of the variable and joining regions, there is an apparent deletion of two nucleotides in the human cDNA sequence relative to the murine gamma-chain cDNA sequence, resulting simultaneously in the generation of an in-frame stop codon and in a translational frameshift. For this reason, the sequence presented here encodes an aberrantly rearranged human T-cell gamma chain. There are several surprising differences between the deduced human and murine gamma-chain amino acid sequences. These include poor homology in the variable region, poor homology in a discrete segment of the constant region precisely bounded by the expected junctions of exon CII, and the presence in the human sequence of five potential sites for N-linked glycosylation. Images PMID:3458221
-
Change in IgHV Mutational Status of CLL Suggests Origin From Multiple Clones.
Osman, Afaf; Gocke, Christopher D; Gladstone, Douglas E
2017-02-01
Fluorescence in situ hybridization and immunoglobulin (Ig) heavy-chain variable-region (IgHV) mutational status are used to predict outcome in chronic lymphocytic leukemia (CLL). Although DNA aberrations change over time, IgHV sequences and mutational status are considered stable. In a retrospective review, 409 CLL patients, between 2008 and 2015, had IgHV analysis: 56 patients had multiple analyses performed. Seven patients' IgHV results changed: 2 from unmutated to mutated and 5 from mutated to unmutated IgHV sequence. Three concurrently changed their variable heavy-chain sequence. Secondary to allelic exclusion, 2 of the new variable heavy chains produced were biologically nonplausible. The existence of these new nonplausible heavy-chain variable regions suggests either the CLL cancer stem-cell maintains the ability to rearrange a previously silenced IgH allele or more likely that the cancer stem-cell produced at least 2 subclones, suggesting that the CLL cancer stem cell exists before the process of allelic exclusion occurs. Copyright © 2016 Elsevier Inc. All rights reserved.
-
Sotelo, Pablo H.; Collazo, Noberto; Zuñiga, Roberto; Gutiérrez-González, Matías; Catalán, Diego; Ribeiro, Carolina Hager; Aguillón, Juan Carlos; Molina, María Carmen
2012-01-01
Phage display library technology is a common method to produce human antibodies. In this technique, the immunoglobulin variable regions are displayed in a bacteriophage in a way that each filamentous virus displays the product of a single antibody gene on its surface. From the collection of different phages, it is possible to isolate the virus that recognizes specific targets. The most common form in which to display antibody variable regions in the phage is the single chain variable fragment format (scFv), which requires assembly of the heavy and light immunoglobulin variable regions in a single gene. In this work, we describe a simple and efficient method for the assembly of immunoglobulin heavy and light chain variable regions in a scFv format. This procedure involves a two-step reaction: (1) DNA amplification to produce the single strand form of the heavy or light chain gene required for the fusion; and (2) mixture of both single strand products followed by an assembly reaction to construct a complete scFv gene. Using this method, we produced 6-fold more scFv encoding DNA than the commonly used splicing by overlap extension PCR (SOE-PCR) approach. The scFv gene produced by this method also proved to be efficient in generating a diverse scFv phage display library. From this scFv library, we obtained phages that bound several non-related antigens, including recombinant proteins and rotavirus particles. PMID:22692130
-
Picken, M. M.; Gallo, G.; Buxbaum, J.; Frangione, B.
1986-01-01
Amyloid fibrils were extracted from the kidney of a patient (CHE) shown to have tetramers and dimers of a monoclonal lambda light chain in his serum, and whose bone marrow cells in short-term culture synthesized these forms and a smaller lambda fragment of approximately 10,000 to 12,000 daltons. Biochemical and serologic analysis of a fraction of a size (obtained from amyloid fibrils extracted from the kidney) similar to that synthesized by the bone marrow cells revealed a light chain fragment corresponding to the amino terminal end of the variable region of the lambda light chain subgroup II. The presence of similarly sized short fragments of lambda light chain in both the synthesized and deposited protein suggests that aberrant synthesis and/or proteolytic degradation may play a pathogenetic role in the process of amyloidogenesis. Images Figure 1 PMID:3089021
-
Human heavy chain disease protein WIS: implications for the organization of immunoglobulin genes.
Franklin, E C; Prelli, F; Frangione, B
1979-01-01
Protein WIS is a human gamma3 heavy (H) chain disease immunoglobulin variant whose amino acid sequence is most readily interpreted by postulating that three residues of the amino terminus are followed by a deletion of most of the variable (VH) domain, which ends at the variable-constant (VC) joining region. Then there is a stretch of eight residues, three of which are unusual, while the other five have striking homology to the VC junction sequence. This is followed by a second deletion, which ends at the beginning of the quadruplicated hinge region. These findings are consistent with mutations resulting in deletions of most of the gene coding for the V region and CH1 domain followed by splicing at the VC joining region and at the hinge. These structural features fit well the notion of genetic discontinuity between V and C genes and also suggest similar mechanisms of excision and splicing in the interdomain regions of the C gene of the heavy chain. PMID:106391
-
Amino terminal sequence of heavy and light chains from ratfish immunoglobulin.
De Ioannes, A E; Aguila, H L
1989-01-01
The ratfish, Callorhinchus callorhinchus, a representative of the Holocephali, has a natural serum hemagglutinin (Mr 960,000), composed of heavy (Mr 71,000), light (Mr 22,500), and J (Mr 16,000) chains. To approach the mechanisms that generate diversity at this level of evolution, the amino terminal sequence of the heavy and light chains was determined by automated microsequencing. The chains are unblocked and have modest internal sequence heterogeneity. The heavy chains show sequence similarity with the terminal region of the heavy chain from the horned shark, Heterodontus francisci, and other species. In contrast to the heavy chain, the ratfish light chains display low sequence similarity with their shark kappa counterparts. However, their similarity with the variable region of the chicken lambda light chains is about 75%.
-
Method for altering antibody light chain interactions
Stevens, Fred J.; Stevens, Priscilla Wilkins; Raffen, Rosemarie; Schiffer, Marianne
2002-01-01
A method for recombinant antibody subunit dimerization including modifying at least one codon of a nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in the interface segment of the light polypeptide variable region, the charged amino acid having a first polarity; and modifying at least one codon of the nucleic acid sequence to replace an amino acid occurring naturally in the antibody with a charged amino acid at a position in an interface segment of the heavy polypeptide variable region corresponding to a position in the light polypeptide variable region, the charged amino acid having a second polarity opposite the first polarity. Nucleic acid sequences which code for novel light chain proteins, the latter of which are used in conjunction with the inventive method, are also provided.
-
Remote creation of a one-qubit mixed state through a short homogeneous spin-1/2 chain
NASA Astrophysics Data System (ADS)
Zenchuk, A. I.
2014-11-01
We consider a method of remote mixed state creation of a one-qubit subsystem (receiver) in a spin-1/2 chain governed by the nearest-neighbor X Y Hamiltonian. Owing to the evolution of the chain along with the variable local unitary transformation of the one- or two-qubit sender, a large variety of receiver states can be created during a specific time interval starting with a fixed initial state of the whole quantum system. These states form the creatable region of the receiver's state-space. It is remarkable that, with a two-qubit sender, a large creatable region may be covered at a properly fixed time instant t0 using just the variable local unitary transformation of the sender. In this case we have completely local control of remote state creation. In general, for a given initial state, there are some receiver states that may not be created using the above tool. These states form the unavailable region. In turn, this unavailable region might be the creatable region of another sender. Thus, in future, we have a way to share the whole receiver's state-space among the creatable regions of several senders. The effectiveness of remote state creation is characterized by the density function of the creatable region.
-
Giudicelli, Véronique; Duroux, Patrice; Kossida, Sofia; Lefranc, Marie-Paule
2017-06-26
IMGT®, the international ImMunoGeneTics information system® ( http://www.imgt.org ), was created in 1989 in Montpellier, France (CNRS and Montpellier University) to manage the huge and complex diversity of the antigen receptors, and is at the origin of immunoinformatics, a science at the interface between immunogenetics and bioinformatics. Immunoglobulins (IG) or antibodies and T cell receptors (TR) are managed and described in the IMGT® databases and tools at the level of receptor, chain and domain. The analysis of the IG and TR variable (V) domain rearranged nucleotide sequences is performed by IMGT/V-QUEST (online since 1997, 50 sequences per batch) and, for next generation sequencing (NGS), by IMGT/HighV-QUEST, the high throughput version of IMGT/V-QUEST (portal begun in 2010, 500,000 sequences per batch). In vitro combinatorial libraries of engineered antibody single chain Fragment variable (scFv) which mimic the in vivo natural diversity of the immune adaptive responses are extensively screened for the discovery of novel antigen binding specificities. However the analysis of NGS full length scFv (~850 bp) represents a challenge as they contain two V domains connected by a linker and there is no tool for the analysis of two V domains in a single chain. The functionality "Analyis of single chain Fragment variable (scFv)" has been implemented in IMGT/V-QUEST and, for NGS, in IMGT/HighV-QUEST for the analysis of the two V domains of IG and TR scFv. It proceeds in five steps: search for a first closest V-REGION, full characterization of the first V-(D)-J-REGION, then search for a second V-REGION and full characterization of the second V-(D)-J-REGION, and finally linker delimitation. For each sequence or NGS read, positions of the 5'V-DOMAIN, linker and 3'V-DOMAIN in the scFv are provided in the 'V-orientated' sense. Each V-DOMAIN is fully characterized (gene identification, sequence description, junction analysis, characterization of mutations and amino changes). The functionality is generic and can analyse any IG or TR single chain nucleotide sequence containing two V domains, provided that the corresponding species IMGT reference directory is available. The "Analysis of single chain Fragment variable (scFv)" implemented in IMGT/V-QUEST and, for NGS, in IMGT/HighV-QUEST provides the identification and full characterization of the two V domains of full-length scFv (~850 bp) nucleotide sequences from combinatorial libraries. The analysis can also be performed on concatenated paired chains of expressed antigen receptor IG or TR repertoires.
-
J Genes for Heavy Chain Immunoglobulins of Mouse
NASA Astrophysics Data System (ADS)
Newell, Nanette; Richards, Julia E.; Tucker, Philip W.; Blattner, Frederick R.
1980-09-01
A 15.8-kilobase pair fragment of BALB/c mouse liver DNA, cloned in the Charon 4Aλ phage vector system, was shown to contain the μ heavy chain constant region (CHμ ) gene for the mouse immunoglobulin M. In addition, this fragment of DNA contains at least two J genes, used to code for the carboxyl terminal portion of heavy chain variable regions. These genes are located in genomic DNA about eight kilobase pairs to the 5' side of the CHμ gene. The complete nucleotide sequence of a 1120-base pair stretch of DNA that includes the two J genes has been determined.
-
The generation and selection of single-domain, v region libraries from nurse sharks.
Flajnik, Martin F; Dooley, Helen
2009-01-01
The cartilaginous fish (sharks, skates, and rays) are the oldest phylogenetic group in which a human-type adaptive immune system and immunoglobulins (Igs) have been found. In addition to their conventional (heavy-light chain heterodimeric) isotypes, IgM and IgW, sharks produce the novel isotype, IgNAR, a heavy chain homodimer that does not associate with light chains. Instead, its variable (V) regions act as independent, soluble units in order to bind antigen. In this chapter, we detail our immunization protocol in order to raise a humoral IgNAR response in the nurse shark (Ginglymostoma cirratum) and the subsequent cloning of the single-domain V regions from this isotype in order to select antigen-specific binders by phage display.
-
Paired analysis of TCRα and TCRβ chains at the single-cell level in mice
Dash, Pradyot; McClaren, Jennifer L.; Oguin, Thomas H.; Rothwell, William; Todd, Brandon; Morris, Melissa Y.; Becksfort, Jared; Reynolds, Cory; Brown, Scott A.; Doherty, Peter C.; Thomas, Paul G.
2010-01-01
Characterizing the TCRα and TCRβ chains expressed by T cells responding to a given pathogen or underlying autoimmunity helps in the development of vaccines and immunotherapies, respectively. However, our understanding of complementary TCRα and TCRβ chain utilization is very limited for pathogen- and autoantigen-induced immunity. To address this problem, we have developed a multiplex nested RT-PCR method for the simultaneous amplification of transcripts encoding the TCRα and TCRβ chains from single cells. This multiplex method circumvented the lack of antibodies specific for variable regions of mouse TCRα chains and the need for prior knowledge of variable region usage in the TCRβ chain, resulting in a comprehensive, unbiased TCR repertoire analysis with paired coexpression of TCRα and TCRβ chains with single-cell resolution. Using CD8+ CTLs specific for an influenza epitope recovered directly from the pneumonic lungs of mice, this technique determined that 25% of such effectors expressed a dominant, nonproductively rearranged Tcra transcript. T cells with these out-of-frame Tcra mRNAs also expressed an alternate, in-frame Tcra, whereas approximately 10% of T cells had 2 productive Tcra transcripts. The proportion of cells with biallelic transcription increased over the course of a response, a finding that has implications for immune memory and autoimmunity. This technique may have broad applications in mouse models of human disease. PMID:21135507
-
The Complete Nucleotide Sequence of the Human Immunoglobulin Heavy Chain Variable Region Locus
Matsuda, Fumihiko; Ishii, Kazuo; Bourvagnet, Patrice; Kuma, Kei-ichi; Hayashida, Hidenori; Miyata, Takashi; Honjo, Tasuku
1998-01-01
The complete nucleotide sequence of the 957-kb DNA of the human immunoglobulin heavy chain variable (VH) region locus was determined and 43 novel VH segments were identified. The region contains 123 VH segments classifiable into seven different families, of which 79 are pseudogenes. Of the 44 VH segments with an open reading frame, 39 are expressed as heavy chain proteins and 1 as mRNA, while the remaining 4 are not found in immunoglobulin cDNAs. Combinatorial diversity of VH region was calculated to be ∼6,000. Conservation of the promoter and recombination signal sequences was observed to be higher in functional VH segments than in pseudogenes. Phylogenetic analysis of 114 VH segments clearly showed clustering of the VH segments of each family. However, an independent branch in the tree contained a single VH, V4-44.1P, sharing similar levels of homology to human VH families and to those of other vertebrates. Comparison between different copies of homologous units that appear repeatedly across the locus clearly demonstrates that dynamic DNA reorganization of the locus took place at least eight times between 133 and 10 million years ago. One nonimmunoglobulin gene of unknown function was identified in the intergenic region. PMID:9841928
-
Norder, Heléne; Bergström, Åsa; Uhnoo, Ingrid; Aldén, Jöran; Weiss, Lars; Czajkowski, Jan; Magnius, Lars
1998-01-01
Four hepatitis C virus transmission chains at three dialysis units were disclosed by limited sequencing; three of these were disclosed by analysis of the NS5-B region of the genome. Dialysis on the same shift as that during which infected patients were dialyzed was the common factor for seven patients in two chains. Two nurses exposed to needle sticks and their sources of infection constituted two other chains. The strains of three chains belonged to subtype 1a and formed clusters with an intrachain variability of 0 to 6 nucleotides compared to 8 to 37 nucleotides for unrelated strains within this subtype. The clusters were supported by bootstrap values ranging from 89 to 100%. PMID:9738071
-
Grasping the nettle: A bacterial invasin that targets immunoglobulin variable domains.
Barlow, Paul
2018-06-01
In a new paper, the protein InvD from Yersinia pseudotuberculosis , a zoonotic pathogen, is shown to assist late-stage invasion of intestinal epithelia. Remarkably, InvD acts by binding the Fab region of IgG or IgA. It straddles adjacent light-chain and heavy-chain variable domains, but its binding is different from that of antigens in that complementarity-determining regions do not participate. Structure determination revealed that its Fab-interacting domain adopts an immunoglobulin-like fold, fused to the preceding immunoglobulin-like domain and carried on a long stalk anchored to the bacterial outer membrane. Possible roles of this unusual host-pathogen interaction include avoidance of clearance from the intestine by secretory IgA. © 2018 Barlow.
-
Effect of light chain V region duplication on IgG oligomerization and in vivo efficacy.
Shuford, W; Raff, H V; Finley, J W; Esselstyn, J; Harris, L J
1991-05-03
A human immunoglobulin G1 (IgG1) antibody oligomer was isolated from a transfected myeloma cell line that produced a monoclonal antibody to group B streptococci. Compared to the IgG1 monomer, the oligomer was significantly more effective at protecting neonatal rats from infection in vivo. The oligomer was also shown to cross the placenta and to be stable in neonatal rats. Immunochemical analysis and complementary DNA sequencing showed that the transfected cell line produced two distinct kappa light chains: a normal light chain (Ln) with a molecular mass of 25 kilodaltons and a 37-kilodalton species (L37), the domain composition of which was variable-variable-constant (V-V-C). Cotransfection of vectors encoding the heavy chain and L37 resulted in production of oligomeric IgG.
-
Liu, Rui; Liang, Xiao; Xiang, Dandan; Guo, Yirong; Liu, Yihua; Zhu, Guonian
2016-01-01
Triazophos is a widely used organophosphorous insecticide that has potentially adverse effects to organisms. In the present study, a high-affinity single-chain variable fragment (scFv) antibody with specific lambda light chain was developed for residue monitoring. First, the specific variable regions were correctly amplified from a hybridoma cell line 8C10 that secreted monoclonal antibody (mAb) against triazophos. The regions were then assembled as scFv via splicing by overlap extension polymerase chain reaction. Subsequently, the recombinant anti-triazophos scFv-8C10 was successfully expressed in Escherichia coli strain HB2151 in soluble form, purified through immobilized metal ion affinity chromatography, and verified via Western blot and peptide mass fingerprinting analyses. Afterward, an indirect competitive enzyme-linked immunosorbent assay was established based on the purified anti-triazophos scFv-8C10 antibody. The assay exhibited properties similar to those based on the parent mAb, with a high sensitivity (IC50 of 1.73 ng/mL) to triazophos and no cross reaction for other organophosphorus pesticides; it was reliable in detecting triazophos residues in spiked water samples. Moreover, kinetic measurement using a surface plasmon resonance biosensor indicated that the purified scFv-8C10 antibody had a high affinity of 1.8 × 10−10 M and exhibited good binding stability. Results indicated that the recombinant high-affinity scFv-8C10 antibody was an effective detection material that would be promising for monitoring triazophos residues in environment samples. PMID:27338340
-
de Re, Valli; Simula, Maria Paola; Pavan, Alessandro; Garziera, Marica; Marin, Dolores; Dolcetti, Riccardo; de Vita, Salvatore; Sansonno, Domenico; Geremia, Silvano; Toffoli, Giuseppe
2009-09-01
Autoimmune type-II cryoglobulinemia (II-MC) is sustained by hepatitis C virus (HCV) infection and B-cell (oligo)clones. This is the reason why the disease may be considered an "indolent B-cell lymphoma (NHL)." B clones show a restricted use of immunoglobulin variable genes (BCR), in particular in the use of the variable kappa (VK)3-20/15 light chain, and show a homology between their BCR functional regions and those of autoimmune rheumatoid factors. We underlined the BCR unique repertoire with frequent rheumatoid factor activity also observed in other autoimmune disorders associated with NHL. The immunoglobulin idiotype is a clonal B-cell marker and an ideal target for immunotherapy. Five monoclonal antibodies were produced in our laboratory toward the VK3-20 of a subject with HCV infection and a II-MC-associated NHL. Epitope determination was performed using the epitope excision approach. Monoclonal antibody reactivity was tested in vitro in ELISA, Western blot, and cytofluorimetry. Data confirmed that a panel of antibodies, reactive against shared idiotypes, can be produced from patients with HCV-associated B-cell lymphoproliferative diseases, thus obviating the need to produce an anti-idiotype antibody for each patient.
-
Parvari, R; Avivi, A; Lentner, F; Ziv, E; Tel-Or, S; Burstein, Y; Schechter, I
1988-03-01
cDNA clones encoding the variable and constant regions of chicken immunoglobulin (Ig) gamma-chains were obtained from spleen cDNA libraries. Southern blots of kidney DNA show that the variable region sequences of eight cDNA clones reveal the same set of bands corresponding to approximately 30 cross-hybridizing VH genes of one subgroup. Since the VH clones were randomly selected, it is likely that the bulk of chicken H-chains are encoded by a single VH subgroup. Nucleotide sequence determinations of two cDNA clones reveal VH, D, JH and the constant region. The VH segments are closely related to each other (83% homology) as expected for VH or the same subgroup. The JHs are 15 residues long and differ by one amino acid. The Ds differ markedly in sequence (20% homology) and size (10 and 20 residues). These findings strongly indicate multiple (at least two) D genes which by a combinatorial joining mechanism diversify the H-chains, a mechanism which is not operative in the chicken L-chain locus. The most notable among the chicken Igs is the so-called 7S IgG because its H-chain differs in many important aspects from any mammalian IgG. The sequence of the C gamma cDNA reported here resolves this issue. The chicken C gamma is 426 residues long with four CH domains (unlike mammalian C gamma which has three CH domains) and it shows 25% homology to the chicken C mu. The chicken C gamma is most related to the mammalian C epsilon in length, the presence of four CH domains and the distribution of cysteines in the CH1 and CH2 domains. We propose that the unique chicken C gamma is the ancestor of the mammalian C epsilon and C gamma subclasses, and discuss the evolution of the H-chain locus from that of chicken with presumably three genes (mu, gamma, alpha) to the mammalian loci with 8-10 H-chain genes.
-
Array-Based Rational Design of Short Peptide Probe-Derived from an Anti-TNT Monoclonal Antibody.
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.
-
Wiehe, Kevin; Easterhoff, David; Luo, Kan; ...
2014-11-29
In HIV-1, the ability to mount antibody responses to conserved, neutralizing epitopes is critical for protection. Here we have studied the light chain usage of human and rhesus macaque antibodies targeted to a dominant region of the HIV-1 envelope second variable (V2) region involving lysine (K) 169, the site of immune pressure in the RV144 vaccine efficacy trial. We found that humans and rhesus macaques used orthologous lambda variable gene segments encoding a glutamic acid-aspartic acid (ED) motif for K169 recognition. Structure determination of an unmutated ancestor antibody demonstrated that the V2 binding site was preconfigured for ED motif-mediated recognitionmore » prior to maturation. Thus, light chain usage for recognition of the site of immune pressure in the RV144 trial is highly conserved across species. In conclusion, these data indicate that the HIV-1 K169-recognizing ED motif has persisted over the diversification between rhesus macaques and humans, suggesting an evolutionary advantage of this antibody recognition mode.« less
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Wiehe, Kevin; Easterhoff, David; Luo, Kan
In HIV-1, the ability to mount antibody responses to conserved, neutralizing epitopes is critical for protection. Here we have studied the light chain usage of human and rhesus macaque antibodies targeted to a dominant region of the HIV-1 envelope second variable (V2) region involving lysine (K) 169, the site of immune pressure in the RV144 vaccine efficacy trial. We found that humans and rhesus macaques used orthologous lambda variable gene segments encoding a glutamic acid-aspartic acid (ED) motif for K169 recognition. Structure determination of an unmutated ancestor antibody demonstrated that the V2 binding site was preconfigured for ED motif-mediated recognitionmore » prior to maturation. Thus, light chain usage for recognition of the site of immune pressure in the RV144 trial is highly conserved across species. In conclusion, these data indicate that the HIV-1 K169-recognizing ED motif has persisted over the diversification between rhesus macaques and humans, suggesting an evolutionary advantage of this antibody recognition mode.« less
-
Goodhardt, M; Babinet, C; Lutfalla, G; Kallenbach, S; Cavelier, P; Rougeon, F
1989-01-01
We have produced transgenic mice which synthesize chimeric mouse-rabbit immunoglobulin (Ig) kappa light chains following in vivo recombination of an injected unrearranged kappa gene. The exogenous gene construct contained a mouse germ-line kappa variable (V kappa) gene segment, the mouse germ-line joining (J kappa) locus including the enhancer, and the rabbit b9 constant (C kappa) region. A high level of V-J recombination of the kappa transgene was observed in spleen of the transgenic mice. Surprisingly, a particularly high degree of variability in the exact site of recombination and the presence of non germ-line encoded nucleotides (N-regions) were found at the V-J junction of the rearranged kappa transgene. Furthermore, unlike endogenous kappa genes, rearrangement of the exogenous gene occurred in T-cells of the transgenic mice. These results show that additional sequences, other than the heptamer-nonamer signal sequences and the promoter and enhancer elements, are required to obtain stage- and lineage- specific regulation of Ig kappa light chain gene rearrangement in vivo. Images PMID:2508061
-
Mapping of Heavy Chain Genes for Mouse Immunoglobulins M and D
NASA Astrophysics Data System (ADS)
Liu, Chih-Ping; Tucker, Philip W.; Mushinski, J. Frederic; Blattner, Frederick R.
1980-09-01
A single DNA fragment containing both μ and δ immunoglobulin heavy chain genes has been cloned from normal BALB/c mouse liver DNA with a new λ phage vector Charon 28. The physical distance between the membrane terminal exon of μ and the first domain of δ is 2466 base pairs, with δ on the 3' side of μ . A single transcript could contain a variable region and both μ and δ constant regions. The dual expression of immunoglobulins M and D on spleen B cells may be due to alternate splicing of this transcript.
-
Dooley, Helen; Flajnik, Martin F; Porter, Andrew J
2003-09-01
The novel immunoglobulin isotype novel antigen receptor (IgNAR) is found in cartilaginous fish and is composed of a heavy-chain homodimer that does not associate with light chains. The variable regions of IgNAR function as independent domains similar to those found in the heavy-chain immunoglobulins of Camelids. Here, we describe the successful cloning and generation of a phage-displayed, single-domain library based upon the variable domain of IgNAR. Selection of such a library generated from nurse sharks (Ginglymostoma cirratum) immunized with the model antigen hen egg-white lysozyme (HEL) enabled the successful isolation of intact antigen-specific binders matured in vivo. The selected variable domains were shown to be functionally expressed in Escherichia coli, extremely stable, and bind to antigen specifically with an affinity in the nanomolar range. This approach can therefore be considered as an alternative route for the isolation of minimal antigen-binding fragments with favorable characteristics.
-
Boehm, M K; Corper, A L; Wan, T; Sohi, M K; Sutton, B J; Thornton, J D; Keep, P A; Chester, K A; Begent, R H; Perkins, S J
2000-03-01
MFE-23 is the first single-chain Fv antibody molecule to be used in patients and is used to target colorectal cancer through its high affinity for carcinoembryonic antigen (CEA), a cell-surface member of the immunoglobulin superfamily. MFE-23 contains an N-terminal variable heavy-chain domain joined by a (Gly(4)Ser)(3) linker to a variable light-chain (V(L)) domain (kappa chain) with an 11-residue C-terminal Myc-tag. Its crystal structure was determined at 2.4 A resolution by molecular replacement with an R(cryst) of 19.0%. Five of the six antigen-binding loops, L1, L2, L3, H1 and H2, conformed to known canonical structures. The sixth loop, H3, displayed a unique structure, with a beta-hairpin loop and a bifurcated apex characterized by a buried Thr residue. In the crystal lattice, two MFE-23 molecules were associated back-to-back in a manner not seen before. The antigen-binding site displayed a large acidic region located mainly within the H2 loop and a large hydrophobic region within the H3 loop. Even though this structure is unliganded within the crystal, there is an unusually large region of contact between the H1, H2 and H3 loops and the beta-sheet of the V(L) domain of an adjacent molecule (strands DEBA) as a result of intermolecular packing. These interactions exhibited remarkably high surface and electrostatic complementarity. Of seven MFE-23 residues predicted to make contact with antigen, five participated in these lattice contacts, and this model for antigen binding is consistent with previously reported site-specific mutagenesis of MFE-23 and its effect on CEA binding.
-
Vekony, M A; Holder, J E; Lee, A J; Horrocks, C; Eperon, I C; Camp, R D
1997-07-01
Several groups have investigated the role of T cells in the pathogenesis of psoriasis by determination of T-cell receptor (TCR) B-chain variable (V) region usage, both in chronic plaque (psoriasis vulgaris) and guttate forms, with various results. Because there are no data on TCR expression in early psoriasis vulgaris, when specific cellular immune events may be expected to be most pronounced, we have analyzed early lesions (less than 3 wk old) of ten patients, with highly reproducible results. We have developed a highly controlled anchored polymerase chain reaction (PCR) method in which TCR beta chain species are all amplified with the same primer pair and products are quantified by dot blot hybridization with BV family-specific oligonucleotide probes. Overexpression of certain TCR BV genes was observed in the majority of lesional biopsies, but in samples in which the expanded BV family formed more than 10% of total lesional BV (half of the samples analyzed), BV2 and BV6 predominated. The consistency of overexpression of these BV species between patients was much less than in previous studies of TCRBV usage in established chronic plaque psoriasis lesions. Complementarity-determining region 3 (CDR3) size spectratyping demonstrated evidence for selective clonal T cell accumulation in less than half of the lesional samples showing BV expansion. These results indicate that selective amplification of TCRBV species occurs in early psoriasis vulgaris but is not essential to the pathogenic process and may be more important in the maintenance or expansion of chronic lesions.
-
Walther, Stefanie; Tietze, Manfred; Czerny, Claus-Peter; König, Sven; Diesterbeck, Ulrike S
2016-01-01
We have developed a new bioinformatics framework for the analysis of rearranged bovine heavy chain immunoglobulin (Ig) variable regions by combining and refining widely used alignment algorithms. This bioinformatics framework allowed us to investigate alignments of heavy chain framework regions (FRHs) and the separate alignments of FRHs and heavy chain complementarity determining regions (CDRHs) to determine their germline origin in the four cattle breeds Aubrac, German Black Pied, German Simmental, and Holstein Friesian. Now it is also possible to specifically analyze Ig heavy chains possessing exceptionally long CDR3Hs. In order to gain more insight into breed specific differences in Ig combinatorial diversity, somatic hypermutations and putative gene conversions of IgG, we compared the dominantly transcribed variable (IGHV), diversity (IGHD), and joining (IGHJ) segments and their recombination in the four cattle breeds. The analysis revealed the use of 15 different IGHV segments, 21 IGHD segments, and two IGHJ segments with significant different transcription levels within the breeds. Furthermore, there are preferred rearrangements within the three groups of CDR3H lengths. In the sequences of group 2 (CDR3H lengths (L) of 11-47 amino acid residues (aa)) a higher number of recombination was observed than in sequences of group 1 (L≤10 aa) and 3 (L≥48 aa). The combinatorial diversity of germline IGHV, IGHD, and IGHJ-segments revealed 162 rearrangements that were significantly different. The few preferably rearranged gene segments within group 3 CDR3H regions may indicate specialized antibodies because this length is unique in cattle. The most important finding of this study, which was enabled by using the bioinformatics framework, is the discovery of strong evidence for gene conversion as a rare event using pseudogenes fulfilling all definitions for this particular diversification mechanism.
-
Czerny, Claus-Peter; König, Sven; Diesterbeck, Ulrike S.
2016-01-01
We have developed a new bioinformatics framework for the analysis of rearranged bovine heavy chain immunoglobulin (Ig) variable regions by combining and refining widely used alignment algorithms. This bioinformatics framework allowed us to investigate alignments of heavy chain framework regions (FRHs) and the separate alignments of FRHs and heavy chain complementarity determining regions (CDRHs) to determine their germline origin in the four cattle breeds Aubrac, German Black Pied, German Simmental, and Holstein Friesian. Now it is also possible to specifically analyze Ig heavy chains possessing exceptionally long CDR3Hs. In order to gain more insight into breed specific differences in Ig combinatorial diversity, somatic hypermutations and putative gene conversions of IgG, we compared the dominantly transcribed variable (IGHV), diversity (IGHD), and joining (IGHJ) segments and their recombination in the four cattle breeds. The analysis revealed the use of 15 different IGHV segments, 21 IGHD segments, and two IGHJ segments with significant different transcription levels within the breeds. Furthermore, there are preferred rearrangements within the three groups of CDR3H lengths. In the sequences of group 2 (CDR3H lengths (L) of 11–47 amino acid residues (aa)) a higher number of recombination was observed than in sequences of group 1 (L≤10 aa) and 3 (L≥48 aa). The combinatorial diversity of germline IGHV, IGHD, and IGHJ-segments revealed 162 rearrangements that were significantly different. The few preferably rearranged gene segments within group 3 CDR3H regions may indicate specialized antibodies because this length is unique in cattle. The most important finding of this study, which was enabled by using the bioinformatics framework, is the discovery of strong evidence for gene conversion as a rare event using pseudogenes fulfilling all definitions for this particular diversification mechanism. PMID:27828971
-
Dul, J L; Argon, Y
1990-01-01
Although immunoglobulin light chains are usually secreted in association with heavy chains, free light chains can be secreted by lymphocytes. To identify the structural features of light chains that are essential for their secretion, we mutated a conserved sequence in the variable domain of a lambda I light chain. The effects of the mutations on secretion were assayed by transient expression in COS-1 cells. One mutant (AV60), which replaced Ala-60 with Val, was secreted as efficiently as wild-type lambda I by transfected COS-1 cells. This result was not surprising because secreted lambda II chains contain valine in this position. However, a second lambda I mutant (AV60FS62), which replaced Phe-62 with Ser as well as Ala-60 with Val, was not secreted. This mutant was arrested in the endoplasmic reticulum, as judged by immunofluorescence and by its association with a lumenal endoplasmic reticulum protein, immunoglobulin heavy chain binding protein (BiP). The defect in secretion was not due to gross misfolding of the lambda I chain, since cells cotransfected with AV60FS62 and an immunoglobulin heavy chain gene produced functional antigen-binding antibodies. These assembled IgM molecules were still not secreted. Hence, the replacement of Phe-62 with Ser specifically affects a determinant on the lambda I light chain that is necessary for the intracellular transport of this molecule. Images PMID:2122454
-
Wang, Man; Zhang, Yan; Zhu, Jianguo
2016-03-01
Staphylococcus aureus is a leading causative agent of bovine mastitis, which can result in significant economic losses to the dairy industry. However, available vaccines against bovine mastitis do not confer adequate protection, although passive immunization with antibodies may be useful to prevent disease. Hence, we constructed a bovine single-chain variable region fragment (scFv) phage display library using cDNAs from peripheral blood lymphocytes of cows with S. aureus-induced mastitis. After four rounds of selection, eight scFvs that bound S. aureus antigens with high affinity were obtained. The framework regions of the variable domains (VH and VL) of the eight scFvs were highly conserved, and the complementarity-determining regions (CDRs) displayed significant diversity, especially CDR3 of the VH domain. All eight scFvs inhibited S. aureus growth in culture medium. Lactating mice were challenged by injecting S. aureus into the fourth mammary gland. Histopathological analysis showed that treatment with these scFvs prior to bacterial challenge maintained the structure of the mammary acini, decreased infiltration of polymorphonuclear neutrophils, increased levels of interferon-gamma and interleukin-4, and reduced tumor necrosis factor-alpha levels in mammary tissues, as compared with mice treatment with physiological saline (P < 0.05). These novel bovine scFvs may be suitable candidates for therapeutic agents for the prevention of S. aureus-induced bovine mastitis.
-
Andris-Widhopf, Jennifer; Steinberger, Peter; Fuller, Roberta; Rader, Christoph; Barbas, Carlos F
2011-09-01
The development of therapeutic antibodies for use in the treatment of human diseases has long been a goal for many researchers in the antibody field. One way to obtain these antibodies is through phage-display libraries constructed from human lymphocytes. This protocol describes the construction of human Fab (fragment antigen binding) antibody libraries. In this method, the individual rearranged heavy- and light-chain variable regions are amplified separately and are linked through a series of overlap polymerase chain reaction (PCR) steps to give the final Fab products that are used for cloning.
-
Andris-Widhopf, Jennifer; Steinberger, Peter; Fuller, Roberta; Rader, Christoph; Barbas, Carlos F
2011-09-01
The development of therapeutic antibodies for use in the treatment of human diseases has long been a goal for many researchers in the antibody field. One way to obtain these antibodies is through phage-display libraries constructed from human lymphocytes. This protocol describes the construction of human scFv (single chain antibody fragment) libraries using a short linker (GGSSRSS) or a long linker (GGSSRSSSSGGGGSGGGG). In this method, the individual rearranged heavy- and light-chain variable regions are amplified separately and are linked through a series of overlap polymerase chain reaction (PCR) steps to give the final scFv products that are used for cloning.
-
Genome complexity in the coelacanth is reflected in its adaptive immune system
Saha, Nil Ratan; Ota, Tatsuya; Litman, Gary W.; Hansen, John; Parra, Zuly; Hsu, Ellen; Buonocore, Francesco; Canapa, Adriana; Cheng, Jan-Fang; Amemiya, Chris T.
2014-01-01
We have analyzed the available genome and transcriptome resources from the coelacanth in order to characterize genes involved in adaptive immunity. Two highly distinctive IgW-encoding loci have been identified that exhibit a unique genomic organization, including a multiplicity of tandemly repeated constant region exons. The overall organization of the IgW loci precludes typical heavy chain class switching. A locus encoding IgM could not be identified either computationally or by using several different experimental strategies. Four distinct sets of genes encoding Ig light chains were identified. This includes a variant sigma-type Ig light chain previously identified only in cartilaginous fishes and which is now provisionally denoted sigma-2. Genes encoding α/β and γ/δ T-cell receptors, and CD3, CD4, and CD8 co-receptors also were characterized. Ig heavy chain variable region genes and TCR components are interspersed within the TCR α/δ locus; this organization previously was reported only in tetrapods and raises questions regarding evolution and functional cooption of genes encoding variable regions. The composition, organization and syntenic conservation of the major histocompatibility complex locus have been characterized. We also identified large numbers of genes encoding cytokines and their receptors, and other genes associated with adaptive immunity. In terms of sequence identity and organization, the adaptive immune genes of the coelacanth more closely resemble orthologous genes in tetrapods than those in teleost fishes, consistent with current phylogenomic interpretations. Overall, the work reported described herein highlights the complexity inherent in the coelacanth genome and provides a rich catalog of immune genes for future investigations.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Demaison, C.; Chastagner, P.; Theze, J.
1994-01-18
Monoclonal anti-DNA antibodies bearing a lupus nephritis-associated idiotype were derived from five patients with systemic lupus erythematosus (SLE). Genes encoding their heavy (H)-chain variable (V[sub H]) regions were cloned and sequenced. When compared with their closest V[sub h] germ-line gene relatives, these sequences exhibit a number of silent (S) and replacement (R) substitutions. The ratios of R/S mutations were much higher in the complementarity-determining regions (CDRs) of the antibodies than in the framework regions. Molecular amplification of genomic V[sub H] genes and Southern hybridization with somatic CDR2-specific oligonucleotide probes showed that the configuration of the V[sub H] genes corresponding tomore » V[sub H] sequences in the nephritogenic antibodies is not present in the patient's own germ-line DNA, implying that the B-cell clones underwent somatic mutation in vivo. These findings, together with the characteristics of the diversity and junctional gene elements utilized to form the antibody, indicate that these autoantibodies have been driven through somatic selection processes reminiscent of those that govern antibody responses triggered by exogenous stimuli.« less
-
Berstein, R M; Schluter, S F; Shen, S; Marchalonis, J J
1996-04-16
All immunoglobulins and T-cell receptors throughout phylogeny share regions of highly conserved amino acid sequence. To identify possible primitive immunoglobulins and immunoglobulin-like molecules, we utilized 3' RACE (rapid amplification of cDNA ends) and a highly conserved constant region consensus amino acid sequence to isolate a new immunoglobulin class from the sandbar shark Carcharhinus plumbeus. The immunoglobulin, termed IgW, in its secreted form consists of 782 amino acids and is expressed in both the thymus and the spleen. The molecule overall most closely resembles mu chains of the skate and human and a new putative antigen binding molecule isolated from the nurse shark (NAR). The full-length IgW chain has a variable region resembling human and shark heavy-chain (VH) sequences and a novel joining segment containing the WGXGT motif characteristic of H chains. However, unlike any other H-chain-type molecule, it contains six constant (C) domains. The first C domain contains the cysteine residue characteristic of C mu1 that would allow dimerization with a light (L) chain. The fourth and sixth domains also contain comparable cysteines that would enable dimerization with other H chains or homodimerization. Comparison of the sequences of IgW V and C domains shows homology greater than that found in comparisons among VH and C mu or VL, or CL thereby suggesting that IgW may retain features of the primordial immunoglobulin in evolution.
-
Crystal Structures of T Cell Receptor (Beta) Chains Related to Rheumatoid Arthritis
DOE Office of Scientific and Technical Information (OSTI.GOV)
Li,H.; van Vranken, S.; Zhao, Y.
The crystal structures of the V{beta}17+ {beta} chains of two human T cell receptors (TCRs), originally derived from the synovial fluid (SF4) and tissue (C5-1) of a patient with rheumatoid arthritis (RA), have been determined in native (SF4) and mutant (C5-1{sub F104{yields}Y/C187{yields}S}) forms, respectively. These TCR {beta} chains form homo-dimers in solution and in crystals. Structural comparison reveals that the main-chain conformations in the CDR regions of the C5-1 and SF4 V{beta}17 closely resemble those of a V{beta}17 JM22 in a bound form; however, the CDR3 region shows different conformations among these three V{beta}17 structures. At the side-chain level, conformationalmore » differences were observed at the CDR2 regions between our two ligand-free forms and the bound JM22 form. Other significant differences were observed at the V{beta} regions 8-12, 40-44, and 82-88 between C5-1/SF4 and JM22 V{beta}17, implying that there is considerable variability in the structures of very similar {beta} chains. Structural alignments also reveal a considerable variation in the V{beta}-C{beta} associations, and this may affect ligand recognition. The crystal structures also provide insights into the structure basis of T cell recognition of Mycoplasma arthritidis mitogen (MAM), a superantigen that may be implicated in the development of human RA. Structural comparisons of the V{beta} domains of known TCR structures indicate that there are significant similarities among V{beta} regions that are MAM-reactive, whereas there appear to be significant structural differences among those V{beta} regions that lack MAM-reactivity. It further reveals that CDR2 and framework region (FR) 3 are likely to account for the binding of TCR to MAM.« less
-
NASA Astrophysics Data System (ADS)
Godar, Javier; Suavet, Clément; Gardner, Toby A.; Dawkins, Elena; Meyfroidt, Patrick
2016-03-01
To date, assessments of the sustainability of agricultural commodity supply chains have largely relied on some combination of macro-scale footprint accounts, detailed life-cycle analyses and fine-scale traceability systems. Yet these approaches are limited in their ability to support the sustainability governance of agricultural supply chains, whether because they are intended for coarser-grained analyses, do not identify individual actors, or are too costly to be implemented in a consistent manner for an entire region of production. Here we illustrate some of the advantages of a complementary middle-ground approach that balances detail and scale of supply chain transparency information by combining consistent country-wide data on commodity production at the sub-national (e.g. municipal) level with per shipment customs data to describe trade flows of a given commodity covering all companies and production regions within that country. This approach can support supply chain governance in two key ways. First, enhanced spatial resolution of the production regions that connect to individual supply chains allows for a more accurate consideration of geographic variability in measures of risk and performance that are associated with different production practices. Second, identification of key actors that operate within a specific supply chain, including producers, traders, shippers and consumers can help discriminate coalitions of actors that have shared stake in a particular region, and that together are capable of delivering more cost-effective and coordinated interventions. We illustrate the potential of this approach with examples from Brazil, Indonesia and Colombia. We discuss how transparency information can deepen understanding of the environmental and social impacts of commodity production systems, how benefits are distributed among actors, and some of the trade-offs involved in efforts to improve supply chain sustainability. We then discuss the challenges and opportunities of our approach to strengthen supply chain governance and leverage more effective and fair accountability systems.
-
Sun, Dongbo; Shi, Hongyan; Chen, Jianfei; Shi, Da; Zhu, Qinghe; Zhang, Hong; Liu, Shengwang; Wang, Yunfeng; Qiu, Huaji; Feng, Li
2012-06-01
Porcine aminopeptidase N (pAPN) is a common cellular receptor for swine transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV). To investigate single-chain fragment variable (scFv) repertoire against pAPN, the genes encoding the immunoglobulin light chain variable region (VL) and heavy chain variable region (VH) were amplified by reverse transcript polymerase chain reaction (RT-PCR) using a series of degenerate primers from the spleen of BABL/c mice immunized with native pAPN. The VL and VH amplicons were combined randomly by a 12 amino acid flexible linker by splicing by overlap extension PCR (SOE-PCR), which produced the scFv gene repertoire. After ligation of the scFv gene repertoire into the T7Select10-3b vector, a mouse scFv phage library specific for pAPN was produced through in vitro packaging. The primary scFv library against pAPN contained 2.0×10(7) recombinant phage clones, and the titer of the amplified library was 3.6×10(9)pfu/mL. BstNI restriction analysis and DNA sequencing revealed that 28 phage clones from the primary pAPN scFv library showed excellent diversity. The effectiveness of the scFv library against pAPN was verified further by phage ELISA using the recombinant protein of the pAPN C subunit as coating antigen. The construction and evaluation of a murine scFv library against the common receptor pAPN of porcine coronaviruses TGEV and PEDV using the T7 phage display system are described. Copyright © 2012 Elsevier B.V. All rights reserved.
-
Saeterbakken, Atle H; Andersen, Vidar; van den Tillaar, Roland
2016-04-01
The purpose of the study was to compare kinematic muscle activation when performing 6 repetition maximum (6RM) squats using constant (free weights) or variable resistance (free weights + elastic bands). Twenty recreationally trained women were recruited with 4.6 ± 2.1 years of resistance training experience and a relative strength (6RM/body mass) of 1.1. After a familiarization session identifying the 6RM loads, the participants performed 6RM squats using constant and variable resistance in a randomized order. The total resistance in the variable resistance group was similar to the constant resistance in the presticking region (98%), but greater in the sticking region (105%) and the poststicking region (113%). In addition, the presticking barbell velocity was 21.0% greater using variable than constant resistance, but 22.8% lower in the poststicking region. No significant differences in muscle electromyographic activity, time occurrence, and vertical displacement between the squat modalities were observed, except for higher barbell displacement poststicking using variable resistance. It was concluded that, due to differences in total resistance in the different regions performing variable compared with constant resistance, greater barbell velocity was observed in the presticking region and lower resistance was observed in the poststicking region. However, the extra resistance in the sticking and poststicking regions during the variable resistance modality did not cause increased muscle activity. When performing squats with heavy resistance, the authors recommend using variable resistance, but we suggest increasing the percentage resistance from the elastic bands or using chains.
-
Honey, Denise M.; Best, Annie; Qiu, Huawei
2018-01-01
ABSTRACT Metelimumab (CAT192) is a human IgG4 monoclonal antibody developed as a TGFβ1-specific antagonist. It was tested in clinical trials for the treatment of scleroderma but later terminated due to lack of efficacy. Subsequent characterization of CAT192 indicated that its TGFβ1 binding affinity was reduced by ∼50-fold upon conversion from the parental single-chain variable fragment (scFv) to IgG4. We hypothesized this result was due to decreased conformational flexibility of the IgG that could be altered via engineering. Therefore, we designed insertion mutants in the elbow region and screened for binding and potency. Our results indicated that increasing the elbow region linker length in each chain successfully restored the isoform-specific and high affinity binding of CAT192 to TGFβ1. The crystal structure of the high binding affinity mutant displays large conformational rearrangements of the variable domains compared to the wild-type antigen-binding fragment (Fab) and the low binding affinity mutants. Insertion of two glycines in both the heavy and light chain elbow regions provided sufficient flexibility for the variable domains to extend further apart than the wild-type Fab, and allow the CDR3s to make additional interactions not seen in the wild-type Fab structure. These interactions coupled with the dramatic conformational changes provide a possible explanation of how the scFv and elbow-engineered Fabs bind TGFβ1 with high affinity. This study demonstrates the benefits of re-examining both structure and function when converting scFv to IgG molecules, and highlights the potential of structure-based engineering to produce fully functional antibodies. PMID:29333938
-
NASA Astrophysics Data System (ADS)
Duan, Lingxun; Bagasra, Omar; Laughlin, Mark A.; Oakes, Joseph W.; Pomerantz, Roger J.
1994-05-01
Human immunodeficiency virus type 1 (HIV-1) has a complex life cycle, which has made it a difficult target for conventional therapeutic modalities. A single-chain antibody moiety, directed against the HIV-1 regulatory protein Rev, which rescues unspliced viral RNA from the nucleus of infected cells, has now been developed. This anti-Rev single-chain construct (SFv) consists of both light and heavy chain variable regions of an anti-Rev monoclonal antibody, which, when expressed intracellularly within human cells, potently inhibits HIV-1 replication. This intracellular SFv molecule is demonstrated to specifically antagonize Rev function. Thus, intracellular SFv expression, against a retroviral regulatory protein, may be useful as a gene therapeutic approach to combat HIV-1 infections.
-
Allegrucci, M; Newman, B A; Young-Cooper, G O; Alexander, C B; Meier, D; Kelus, A S; Mage, R G
1990-07-01
Rabbits of the Alicia strain have a mutation (ali) that segregates with the immunoglobulin heavy-chain (lgh) locus and has a cis effect upon the expression of heavy-chain variable-region (VH) genes encoding the a2 allotype. In heterozygous a1/ali or a3/ali rabbits, serum immunoglobulins are almost entirely the products of the normal a1 or a3 allele and only traces of a2 immunoglobulin are detectable. Adult homozygous ali/ali rabbits likewise have normal immunoglobulin levels resulting from increased production of a-negative immunoglobulins and some residual ability to produce the a2 allotype. By contrast, the majority of the immunoglobulins of wild-type a2 rabbits are a2-positive and only a small percentage are a-negative. Genomic DNAs from homozygous mutant and wild-type animals were indistinguishable by Southern analyses using a variety of restriction enzyme digests and lgh probes. However, when digests with infrequently cutting enzymes were analyzed by transverse alternating-field electrophoresis, the ali DNA fragments were 10-15 kilobases smaller than the wild type. These fragments hybridized to probes both for VH and for a region of DNA a few kilobases downstream of the VH genes nearest the joining region. We suggest that this relatively small deletion affects a segment containing 3' VH genes with important regulatory functions, the loss of which leads to the ali phenotype. These results, and the fact that the 3' VH genes rearrange early in B-cell development, indicate that the 3' end of the VH locus probably plays a key role in regulation of VH gene expression.
-
Allegrucci, M; Newman, B A; Young-Cooper, G O; Alexander, C B; Meier, D; Kelus, A S; Mage, R G
1990-01-01
Rabbits of the Alicia strain have a mutation (ali) that segregates with the immunoglobulin heavy-chain (lgh) locus and has a cis effect upon the expression of heavy-chain variable-region (VH) genes encoding the a2 allotype. In heterozygous a1/ali or a3/ali rabbits, serum immunoglobulins are almost entirely the products of the normal a1 or a3 allele and only traces of a2 immunoglobulin are detectable. Adult homozygous ali/ali rabbits likewise have normal immunoglobulin levels resulting from increased production of a-negative immunoglobulins and some residual ability to produce the a2 allotype. By contrast, the majority of the immunoglobulins of wild-type a2 rabbits are a2-positive and only a small percentage are a-negative. Genomic DNAs from homozygous mutant and wild-type animals were indistinguishable by Southern analyses using a variety of restriction enzyme digests and lgh probes. However, when digests with infrequently cutting enzymes were analyzed by transverse alternating-field electrophoresis, the ali DNA fragments were 10-15 kilobases smaller than the wild type. These fragments hybridized to probes both for VH and for a region of DNA a few kilobases downstream of the VH genes nearest the joining region. We suggest that this relatively small deletion affects a segment containing 3' VH genes with important regulatory functions, the loss of which leads to the ali phenotype. These results, and the fact that the 3' VH genes rearrange early in B-cell development, indicate that the 3' end of the VH locus probably plays a key role in regulation of VH gene expression. Images PMID:2115171
-
Naturally Occurring Structural Isomers in Serum IgA1 O-Glycosylation
Takahashi, Kazuo; Smith, Archer D.; Poulsen, Knud; Kilian, Mogens; Julian, Bruce A.; Mestecky, Jiri; Novak, Jan; Renfrow, Matthew B.
2013-01-01
IgA is the most abundantly produced antibody and plays an important role in the mucosal immune system. Human IgA is represented by two isotypes, IgA1 and IgA2. The major structural difference between these two subclasses is the presence of nine potential sites of O-glycosylation in the hinge region between the first and second constant region domains of the heavy chain. Thr225, Thr228, Ser230, Ser232 and Thr236 have been identified as the predominant sites of O-glycan attachment. The range and distribution of O-glycan chains at each site within the context of adjacent sites in this clustered region create a complex heterogeneity of surface epitopes that is incompletely defined. We previously described the analysis of IgA1 O-glycan heterogeneity by use of high resolution LC/MS and electron capture dissociation tandem MS to unambiguously localize all amino acid attachment sites in IgA1 (Ale) myeloma protein. Here, we report the identification and elucidation of IgA1 O-glycopeptide structural isomers that occur based on amino acid position of the attached glycans (positional isomers) and the structure of the O-glycan chains at individual sites (glycan isomers). These isomers are present in a model IgA1 (Mce1) myeloma protein and occur naturally in normal human serum IgA1. Variable O-glycan chains attached to Ser230, Thr233 or Thr236 produce the predominant positional isomers, including O-glycans composed of a single GalNAc residue. These findings represent the first definitive identification of structural isomeric IgA1 O-glycoforms, define the single-site heterogeneity for all O-glycan sites in a single sample, and have implications for defining epitopes based on clustered O-glycan variability. PMID:22067045
-
Hwang, Joyce K.; Wang, Chong; Du, Zhou; Meyers, Robin M.; Kepler, Thomas B.; Neuberg, Donna; Kwong, Peter D.; Mascola, John R.; Joyce, M. Gordon; Bonsignori, Mattia; Haynes, Barton F.; Yeap, Leng-Siew; Alt, Frederick W.
2017-01-01
Variable regions of Ig chains provide the antigen recognition portion of B-cell receptors and derivative antibodies. Ig heavy-chain variable region exons are assembled developmentally from V, D, J gene segments. Each variable region contains three antigen-contacting complementarity-determining regions (CDRs), with CDR1 and CDR2 encoded by the V segment and CDR3 encoded by the V(D)J junction region. Antigen-stimulated germinal center (GC) B cells undergo somatic hypermutation (SHM) of V(D)J exons followed by selection for SHMs that increase antigen-binding affinity. Some HIV-1–infected human subjects develop broadly neutralizing antibodies (bnAbs), such as the potent VRC01-class bnAbs, that neutralize diverse HIV-1 strains. Mature VRC01-class bnAbs, including VRC-PG04, accumulate very high SHM levels, a property that hinders development of vaccine strategies to elicit them. Because many VRC01-class bnAb SHMs are not required for broad neutralization, high overall SHM may be required to achieve certain functional SHMs. To elucidate such requirements, we used a V(D)J passenger allele system to assay, in mouse GC B cells, sequence-intrinsic SHM-targeting rates of nucleotides across substrates representing maturation stages of human VRC-PG04. We identify rate-limiting SHM positions for VRC-PG04 maturation, as well as SHM hotspots and intrinsically frequent deletions associated with SHM. We find that mature VRC-PG04 has low SHM capability due to hotspot saturation but also demonstrate that generation of new SHM hotspots and saturation of existing hotspot regions (e.g., CDR3) does not majorly influence intrinsic SHM in unmutated portions of VRC-PG04 progenitor sequences. We discuss implications of our findings for bnAb affinity maturation mechanisms. PMID:28747530
-
Tiller, Thomas; Schuster, Ingrid; Deppe, Dorothée; Siegers, Katja; Strohner, Ralf; Herrmann, Tanja; Berenguer, Marion; Poujol, Dominique; Stehle, Jennifer; Stark, Yvonne; Heßling, Martin; Daubert, Daniela; Felderer, Karin; Kaden, Stefan; Kölln, Johanna; Enzelberger, Markus; Urlinger, Stefanie
2013-01-01
This report describes the design, generation and testing of Ylanthia, a fully synthetic human Fab antibody library with 1.3E+11 clones. Ylanthia comprises 36 fixed immunoglobulin (Ig) variable heavy (VH)/variable light (VL) chain pairs, which cover a broad range of canonical complementarity-determining region (CDR) structures. The variable Ig heavy and Ig light (VH/VL) chain pairs were selected for biophysical characteristics favorable to manufacturing and development. The selection process included multiple parameters, e.g., assessment of protein expression yield, thermal stability and aggregation propensity in fragment antigen binding (Fab) and IgG1 formats, and relative Fab display rate on phage. The framework regions are fixed and the diversified CDRs were designed based on a systematic analysis of a large set of rearranged human antibody sequences. Care was taken to minimize the occurrence of potential posttranslational modification sites within the CDRs. Phage selection was performed against various antigens and unique antibodies with excellent biophysical properties were isolated. Our results confirm that quality can be built into an antibody library by prudent selection of unmodified, fully human VH/VL pairs as scaffolds. PMID:23571156
-
1985-01-01
Previous studies (21) have shown that two mouse kappa light (L) chain variable (V) region polymorphisms, the IB-peptide and Efla markers, reflect expression of a characteristic group of V kappa regions, called V kappa Ser, by some inbred strains and not others. Expression of V kappa Ser is controlled by a locus on chromosome 6, the chromosome that contains the kappa locus. To further characterize this V kappa group and begin to analyze the basis for its strain-specific expression, full- length complementary DNA (cDNA) copies were produced of L chain mRNA from the M75 myeloma that had been induced in the C.C58 strain of mice, and which produces a V kappa Ser L chain. The C.C58 strain is congenic with BALB/cAn, differing in the region of chromosome 6 that controls expression of the V kappa polymorphisms and the Lyt-2 and Lyt-3 T cell alloantigens. The complete nucleotide sequence of this cloned cDNA was determined and compared with the nucleotide sequences the most closely related BALB/c myeloma L chains known. Results indicated significant differences throughout the variable region, but particularly toward the 5' portion of the sequence. A probe corresponding to 200 bp of the 5' end of the cloned V kappa Ser cDNA was used in Southern hybridizations of restriction digests of liver DNA from a number of inbred, recombinant, and recombinant inbred strains. Under stringent hybridization conditions, one strongly-hybridizing fragment was observed in Bam HI, Hind III, and Eco RI digests, and based on the size of the fragments, strains could be organized into two groups. The presence of strongly hybridizing Bam HI, Hind III, and Eco RI fragments of 3.2, 2.8, and 2.1 kb, respectively, was found to correlate completely with expression by the strain of the IB-peptide and Efla markers. All nonexpressor strains yielded hybridizing fragments of 7.8, 8.4, and 2.8 kb, respectively. Possible explanations for strain- specific expression of V kappa Ser-associated phenotypic markers are discussed. PMID:3926938
-
Shanehbandi, Dariush; Majidi, Jafar; Kazemi, Tohid; Baradaran, Behzad; Aghebati-Maleki, Leili
2017-01-01
CD20-based targeting of B-cells in hematologic malignancies and autoimmune disorders is associated with outstanding clinical outcomes. Isolation and characterization of VH and VL cDNAs encoding the variable regions of the heavy and light chains of monoclonal antibodies (MAb) is necessary to produce next generation MAbs and their derivatives such as bispecific antibodies (bsAb) and single-chain variable fragments (scFv). This study was aimed at cloning and characterization of the VH and VL cDNAs from a hybridoma cell line producing an anti-CD20 MAb. VH and VL fragments were amplified, cloned and characterized. Furthermore, amino acid sequences of VH, VL and corresponding complementarity-determining regions (CDR) were determined and compared with those of four approved MAbs including Rituximab (RTX), Ibritumomab tiuxetan, Ofatumumab and GA101. The cloned VH and VL cDNAs were found to be functional and follow a consensus pattern. Amino acid sequences corresponding to the VH and VL fragments also indicated noticeable homologies to those of RTX and Ibritumomab. Furthermore, amino acid sequences of the relating CDRs had remarkable similarities to their counterparts in RTX and Ibritumomab. Successful recovery of VH and VL fragments encourages the development of novel CD20 targeting bsAbs, scFvs, antibody conjugates and T-cells armed with chimeric antigen receptors.
-
Related Mechanisms of Antibody Somatic Hypermutation and Class Switch Recombination
HWANG, JOYCE K.; ALT, FREDERICK W.; YEAP, LENG-SIEW
2015-01-01
The primary antibody repertoire is generated by mechanisms involving the assembly of the exons that encode the antigen-binding variable regions of immunoglobulin heavy (IgH) and light (IgL) chains during the early development of B lymphocytes. After antigen-dependent activation, mature B lymphocytes can further alter their IgH and IgL variable region exons by the process of somatic hypermutation (SHM), which allows the selection of B cells in which SHMs resulted in the production of antibodies with increased antigen affinity. In addition, during antigen-dependent activation, B cells can also change the constant region of their IgH chain through a DNA double-strand-break (DSB) dependent process referred to as IgH class switch recombination (CSR), which generates B cell progeny that produce antibodies with different IgH constant region effector functions that are best suited for a elimination of a particular pathogen or in a particular setting. Both the mutations that underlie SHM and the DSBs that underlie CSR are initiated in target genes by activation-induced cytidine deaminase (AID). This review describes in depth the processes of SHM and CSR with a focus on mechanisms that direct AID cytidine deamination in activated B cells and mechanisms that promote the differential outcomes of such cytidine deamination. PMID:26104555
-
Rangnoi, Kuntalee; Choowongkomon, Kiattawee; O'Kennedy, Richard; Rüker, Florian; Yamabhai, Montarop
2018-06-06
A human antiaflatoxin B1 (AFB1) scFv antibody (yAFB1-c3), selected from a naı̈ve human phage-displayed scFv library, was used as a template for improving and analysis of antibody-ligand interactions using the chain-shuffling technique. The variable-heavy and variable-light (VH/VL)-shuffled library was constructed from the VH of 25 preselected clones recombined with the VL of yAFB1-c3 and vice versa. Affinity selection from these libraries demonstrated that the VH domain played an important role in the binding of scFv to free AFB1. Therefore, in the next step, VH-shuffled scFv library was constructed from variable-heavy (VH) chain repertoires, amplified from the naı̈ve library, recombined with the variable-light (VL) chain of the clone yAFB1-c3. This library was then used to select a specific scFv antibody against soluble AFB1 by a standard biopanning method. Three clones that showed improved binding properties were isolated. Amino acid sequence analysis indicated that the improved clones have amino acid mutations in framework 1 (FR1) and the complementarity determining region (CDR1) of the VH chain. One clone, designated sAFH-3e3, showed 7.5-fold improvement in sensitivity over the original scFv clone and was selected for molecular binding studies with AFB1. Homology modeling and molecular docking were used to compare the binding of this and the original clones. The results confirmed that VH is more important than VL for AFB1 binding.
-
Vapor-liquid equilibrium and critical asymmetry of square well and short square well chain fluids.
Li, Liyan; Sun, Fangfang; Chen, Zhitong; Wang, Long; Cai, Jun
2014-08-07
The critical behavior of square well fluids with variable interaction ranges and of short square well chain fluids have been investigated by grand canonical ensemble Monte Carlo simulations. The critical temperatures and densities were estimated by a finite-size scaling analysis with the help of histogram reweighting technique. The vapor-liquid coexistence curve in the near-critical region was determined using hyper-parallel tempering Monte Carlo simulations. The simulation results for coexistence diameters show that the contribution of |t|(1-α) to the coexistence diameter dominates the singular behavior in all systems investigated. The contribution of |t|(2β) to the coexistence diameter is larger for the system with a smaller interaction range λ. While for short square well chain fluids, longer the chain length, larger the contribution of |t|(2β). The molecular configuration greatly influences the critical asymmetry: a short soft chain fluid shows weaker critical asymmetry than a stiff chain fluid with same chain length.
-
The mini-exon genes of three Phytomonas isolates that differ in plant tissue tropism.
Sturm, N R; Fernandes, O; Campbell, D A
1995-08-01
The tandem mini-exon gene repeat is an ideal diagnostic target for trypanosomatids because it includes sequences that are conserved absolutely coupled with regions of extreme variability. We have exploited these features and the polymerase chain reaction to differentiate Phytomonas strains isolated from phloem, fruit or latex of various host plants. While the transcribed regions are nearly identical, the intergenic sequences are variable in size and content (130-332 base pairs). The mini-exon genes of these phytomonads can therefore be distinguished from each other and from the corresponding genes in insect trypanosomes, with which they are oft confused.
-
Crystal structure of a shark single-domain antibody V region in complex with lysozyme.
Stanfield, Robyn L; Dooley, Helen; Flajnik, Martin F; Wilson, Ian A
2004-09-17
Cartilaginous fish are the phylogenetically oldest living organisms known to possess components of the vertebrate adaptive immune system. Key to their immune response are heavy-chain, homodimeric immunoglobulins called new antigen receptors (IgNARs), in which the variable (V) domains recognize antigens with only a single immunoglobulin domain, akin to camelid heavy-chain V domains. The 1.45 angstrom resolution crystal structure of the type I IgNAR V domain in complex with hen egg-white lysozyme (HEL) reveals a minimal antigen-binding domain that contains only two of the three conventional complementarity-determining regions but still binds HEL with nanomolar affinity by means of a binding interface comparable in size to conventional antibodies.
-
Antibody repertoire development in camelids.
De Genst, Erwin; Saerens, Dirk; Muyldermans, Serge; Conrath, Katja
2006-01-01
The humoral immune response of the Camelidae is unique as these animals are the only known mammals that seem to possess functional homodimeric heavy-chain antibodies besides the classical heteromeric antibodies composed of heavy (H) and light (L) chains. By definition, the heavy-chain antibodies lack the L-chain, and it was noticed that their H-chain is devoid of the typical first constant domain (CH1) and contains a dedicated variable domain, referred to as VHH. The VHH exon is assembled from separate V-D-J gene segments. The recombined VHH region is subjected to somatic hypermutations; however, the timing and actual mechanism of the class switch from mu to the dedicated gamma-isotype remains elusive. Interestingly, antigen-specific VHHs are easily retrieved after panning of a phage-displayed rearranged V-gene pool cloned from an immunised camelid. These single-domain antigen binding entities possess a number of biophysical properties that offer particular advantages in various medical and biotechnological applications.
-
Catera, Rosa; Hatzi, Katerina; Yan, Xiao-Jie; Zhang, Lu; Wang, Xiao Bo; Fales, Henry M.; Allen, Steven L.; Kolitz, Jonathan E.; Rai, Kanti R.; Chiorazzi, Nicholas
2008-01-01
Leukemic B lymphocytes of a large group of unrelated chronic lymphocytic leukemia (CLL) patients express an unmutated heavy chain immunoglobulin variable (V) region encoded by IGHV1-69, IGHD3-16, and IGHJ3 with nearly identical heavy and light chain complementarity-determining region 3 sequences. The likelihood that these patients developed CLL clones with identical antibody V regions randomly is highly improbable and suggests selection by a common antigen. Monoclonal antibodies (mAbs) from this stereotypic subset strongly bind cytoplasmic structures in HEp-2 cells. Therefore, HEp-2 cell extracts were immunoprecipitated with recombinant stereotypic subset-specific CLL mAbs, revealing a major protein band at approximately 225 kDa that was identified by mass spectrometry as nonmuscle myosin heavy chain IIA (MYHIIA). Reactivity of the stereotypic mAbs with MYHIIA was confirmed by Western blot and immunofluorescence colocalization with anti-MYHIIA antibody. Treatments that alter MYHIIA amounts and cytoplasmic localization resulted in a corresponding change in binding to these mAbs. The appearance of MYHIIA on the surface of cells undergoing stress or apoptosis suggests that CLL mAb may generally bind molecules exposed as a consequence of these events. Binding of CLL mAb to MYHIIA could promote the development, survival, and expansion of these leukemic cells. PMID:18812466
-
Duan, L; Pomerantz, R J
1994-01-01
The pooled degenerate-primer polymerase chain reaction (PCR) technology is now widely used in the amplification and cloning of murine hybridoma-specific immunoglobulin gene cDNAs. The design of primers is mainly based on the highly conserved 5' terminus of immunoglobulin gene variable regions and the constant region in the 3' terminus. Of note, most murine hybridoma cell lines are derived from the Sp2/0 cell line, which is demonstrated to express endogenous aberrant kappa chains (abV kappa). This high-level endogenous abV kappa mixes with specific kappa chains in the hybridomas and interferes with the efficiency of the reverse transcriptase (RT)-PCR cloning strategy. In this report, during the cloning of murine anti-human immunodeficiency virus type I (HIV-1) hybridoma immunoglobulin cDNAs, a specific primer-PCR screening system was developed, based on the abV kappa complementarity-defining region (CDR), to eliminate abV kappa-carrying plasmids. Furthermore, an abV kappa sequence-specific derived ribozyme was developed and packaged in a retroviral expression vector system. This abV kappa ribozyme can be transduced into different murine hybridomas, and expressed intracellularly to potently eliminate endogenous abV kappa RNA. Images PMID:7816635
-
Production of Recombinant Human scFv Against Tetanus Toxin Heavy Chain by Phage Display Technology.
Khalili, Ehsan; Lakzaei, Mostafa; Rasaee, Mohhamad Javad; Aminian, Mahdi
2015-10-01
Tetanus, as a major cause of death in developing countries, is caused by tetanus neurotoxin. Recombinant antibodies against tetanus neurotoxin can be useful in tetanus management. Phage display of antibody fragments from immune human antibody libraries with single chain constructs combining the variable fragments (scFv) has been one of the most prominent technologies in antibody engineering. The aim of this study was the generation of a single chain fragment of variable region (scFv) library and selection of specific antibodies with high affinity against tetanus toxin. Immune human single chain fragment variable (HuscFv) antibody phagemid library was displayed on pIII of filamentous bacteriophage. Selection of scFv clones was performed against tetanus toxin antigens after three rounds of panning. The selected scFv clones were analyzed for inhibition of tetanus toxin binding to ganglioside GT1b. After the third round of panning, over 35 HuscFv phages specific for tetanus toxin were isolated from this library of which 15 clones were found to bind specifically to tetanus toxin. The selected HuscFv phages expressed as a soluble HuscFv peptide and some clones showed positive signals against tetanus toxin. We found that six HuscFv clones inhibit toxin binding to ganglioside GT1b. These selected antibodies can be used in the management of tetanus.
-
A 3,000 year plant leaf wax D/H record of paleohydrology from Southern California
NASA Astrophysics Data System (ADS)
Cheetham, M. I.; Feakins, S. J.
2011-12-01
We report a high resolution record of hydrological variability based on plant leaf wax D/H measurements in a sediment core from Zaca Lake, Southern California spanning 3,000 years. Compound specific analysis of the n-alkanoic acid fraction yields a powerful suite of relative abundance and isotope data. Comparison to modern vegetation and sediments offers insights into the source of sedimentary waxes and their D/H signatures. We identify three potential sources of waxes at Zaca: 1) mid to long chains from emergent aquatic plants (C28 max), 2) mid chains from Pinus coulteri (C20-C24 only) and 3) mid to long chains from Quercus agrifolia (C28 and C30 max). We establish that long chains are dominated by Quercus and other terrestrial vegetation, whereas mid chains could have mixed sources. At Zaca, 80% shared variance between mid chain and long chain records suggests both derive from terrestrial plants influenced by a common driver, presumably the isotopic composition of precipitation or relative humidity influences on leaf water enrichment. Dendrogram analysis of molecular abundance variations allows us to flag where vegetation change cannot be ruled out. We therefore infer that regional atmospheric circulation changes drove the sustained negative isotope excursion of > 20% from about 2,700 to 2,000 years BP and superimposed higher frequency 20% variability throughout the record, including a number of large, rapid excursion (>20%, <20 years). The results of ongoing analyses will be presented at the meeting.
-
Multilocus Association Mapping Using Variable-Length Markov Chains
Browning, Sharon R.
2006-01-01
I propose a new method for association-based gene mapping that makes powerful use of multilocus data, is computationally efficient, and is straightforward to apply over large genomic regions. The approach is based on the fitting of variable-length Markov chain models, which automatically adapt to the degree of linkage disequilibrium (LD) between markers to create a parsimonious model for the LD structure. Edges of the fitted graph are tested for association with trait status. This approach can be thought of as haplotype testing with sophisticated windowing that accounts for extent of LD to reduce degrees of freedom and number of tests while maximizing information. I present analyses of two published data sets that show that this approach can have better power than single-marker tests or sliding-window haplotypic tests. PMID:16685642
-
Multilocus association mapping using variable-length Markov chains.
Browning, Sharon R
2006-06-01
I propose a new method for association-based gene mapping that makes powerful use of multilocus data, is computationally efficient, and is straightforward to apply over large genomic regions. The approach is based on the fitting of variable-length Markov chain models, which automatically adapt to the degree of linkage disequilibrium (LD) between markers to create a parsimonious model for the LD structure. Edges of the fitted graph are tested for association with trait status. This approach can be thought of as haplotype testing with sophisticated windowing that accounts for extent of LD to reduce degrees of freedom and number of tests while maximizing information. I present analyses of two published data sets that show that this approach can have better power than single-marker tests or sliding-window haplotypic tests.
-
Sequence Analysis and Domain Motifs in the Porcine Skin Decorin Glycosaminoglycan Chain*
Zhao, Xue; Yang, Bo; Solakylidirim, Kemal; Joo, Eun Ji; Toida, Toshihiko; Higashi, Kyohei; Linhardt, Robert J.; Li, Lingyun
2013-01-01
Decorin proteoglycan is comprised of a core protein containing a single O-linked dermatan sulfate/chondroitin sulfate glycosaminoglycan (GAG) chain. Although the sequence of the decorin core protein is determined by the gene encoding its structure, the structure of its GAG chain is determined in the Golgi. The recent application of modern MS to bikunin, a far simpler chondroitin sulfate proteoglycans, suggests that it has a single or small number of defined sequences. On this basis, a similar approach to sequence the decorin of porcine skin much larger and more structurally complex dermatan sulfate/chondroitin sulfate GAG chain was undertaken. This approach resulted in information on the consistency/variability of its linkage region at the reducing end of the GAG chain, its iduronic acid-rich domain, glucuronic acid-rich domain, and non-reducing end. A general motif for the porcine skin decorin GAG chain was established. A single small decorin GAG chain was sequenced using MS/MS analysis. The data obtained in the study suggest that the decorin GAG chain has a small or a limited number of sequences. PMID:23423381
-
Kaneko, Mika K; Abe, Shinji; Ogasawara, Satoshi; Fujii, Yuki; Yamada, Shinji; Murata, Takeshi; Uchida, Hiroaki; Tahara, Hideaki; Nishioka, Yasuhiko; Kato, Yukinari
2017-02-01
Podoplanin (PDPN), a type I transmembrane 36-kDa glycoprotein, is expressed not only in normal cells, such as renal epithelial cells (podocytes), lymphatic endothelial cells, and pulmonary type I alveolar cells, but also in cancer cells, including brain tumors and lung squamous cell carcinomas. Podoplanin activates platelet aggregation by binding to C-type lectin-like receptor-2 (CLEC-2) on platelets, and the podoplanin/CLEC-2 interaction facilitates blood/lymphatic vessel separation. We previously produced neutralizing anti-human podoplanin monoclonal antibody (mAb), clone NZ-1 (rat IgG 2a , lambda), which neutralizes the podoplanin/CLEC-2 interaction and inhibits platelet aggregation and cancer metastasis. Human-rat chimeric antibody, NZ-8, was previously developed using variable regions of NZ-1 and human constant regions of heavy chain (IgG 1 ) and light chain (kappa chain). Although NZ-8 showed high antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC) against human podoplanin-expressing cancer cells, the binding affinity of NZ-8 was lower than that of NZ-1. Herein, we produced a novel human-rat chimeric antibody, NZ-12, the constant regions of which consist of IgG 1 heavy chain and lambda light chain. Using flow cytometry, we demonstrated that the binding affinity of NZ-12 was much higher than that of NZ-8. Furthermore, ADCC and CDC activities of NZ-12 were significantly increased against glioblastoma cell lines (LN319 and D397) and lung cancer cell line (PC-10). These results suggested that NZ-12 could become a promising therapeutic antibody against podoplanin-expressing brain tumors and lung cancers.
-
Yuan, Qing; Jordan, Ramon; Brlansky, Ronald H; Istomina, Olga; Hartung, John
2015-10-01
Xylella fastidiosa is a member of the gamma proteobacteria. It is fastidious, insect-vectored and xylem-limited and causes a variety of diseases, some severe, on a wide range of economically important perennial crops, including grape and citrus. Antibody based detection assays are commercially available for X. fastidiosa, and are effective at the species, but not at the subspecies level. We have made a library of scFv antibody fragments directed against X. fastidiosa subsp. pauca strain 9a5c (citrus) by using phage display technology. Antibody gene repertoires were PCR-amplified using 23 primers for the heavy chain variable region (V(H)) and 21 primers for the light chain variable region (V(L)). The V(H) and V(L) were joined by overlap extension PCR, and then the genes of the scFv library were ligated into the phage vector pKM19. The library contained 1.2×10(7) independent clones with full-length scFv inserts. In each of 3cycles of affinity-selection with 9a5c, about 1.0×10(12) phage were used for panning with 4.1×10(6), 7.1×10(6), 2.1×10(7) phage recovered after the first, second and third cycles, respectively. Sixty-six percent of clones from the final library bound X. fastidiosa 9a5c in an ELISA. Some of these scFv antibodies recognized strain 9a5c and did not recognize X. fastidiosa strains that cause Pierce's disease of grapevine. Published by Elsevier B.V.
-
Gadermaier, Elisabeth; Flicker, Sabine; Lupinek, Christian; Steinberger, Peter; Valenta, Rudolf
2013-04-01
Affinity and clonality of allergen-specific IgE antibodies are important determinants for the magnitude of IgE-mediated allergic inflammation. We sought to analyze the contribution of heavy and light chains of human allergen-specific IgE antibodies for allergen specificity and to test whether promiscuous pairing of heavy and light chains with different allergen specificity allows binding and might affect affinity. Ten IgE Fabs specific for 3 non-cross-reactive major timothy grass pollen allergens (Phl p 1, Phl p 2, and Phl p 5) obtained by means of combinatorial cloning from patients with grass pollen allergy were used to construct stable recombinant single chain variable fragments (ScFvs) representing the original Fabs and shuffled ScFvs in which heavy chains were recombined with light chains from IgE Fabs with specificity for other allergens by using the pCANTAB 5 E expression system. Possible ancestor genes for the heavy chain and light chain variable region-encoding genes were determined by using sequence comparison with the ImMunoGeneTics database, and their chromosomal locations were determined. Recombinant ScFvs were tested for allergen specificity and epitope recognition by means of direct and sandwich ELISA, and affinity by using surface plasmon resonance experiments. The shuffling experiments demonstrate that promiscuous pairing of heavy and light chains is possible and maintains allergen specificity, which is mainly determined by the heavy chains. ScFvs consisting of different heavy and light chains exhibited different affinities and even epitope specificity for the corresponding allergen. Our results indicate that allergen specificity of allergen-specific IgE is mainly determined by the heavy chains. Different heavy and light chain pairings in allergen-specific IgE antibodies affect affinity and epitope specificity and thus might influence clinical reactivity to allergens. Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Ben-Avraham, Z.; Nur, A.
The elevation above sea level of circum-Pacific volcanoes situated on continental crust varies greatly, not only between various chains but also within chains. Their edifice heights, however, are essentially constant with each chain. This pattern is reversed for oceanic volcanoes: The elevation circum-Pacific volcanoes situated on oceanic curst is constant within arcs, while edifice heights are greatly variable. In continents the depth to the root zones of volcanoes may be within the elastic part of the lithosphere, whereas in the oceans it may be well below the elastic part of the lithosphere. We suggest that melting, or the onset ofmore » the volcanic uprising, may be controlled in both cases primarily by pressure: in the continental lithosphere by the overburden pressure determined by depth below the local surface and in the oceanic lithosphere by the isostatically compensated pressure zone controlled by depth below sea level. The pattern seems to hold even in complex geological regions and may be used to identify the nature of the crust in such regions.« less
-
Maturation of Shark Single-Domain (IgNAR) Antibodies: Evidence for Induced-Fit Binding
DOE Office of Scientific and Technical Information (OSTI.GOV)
Stanfield, R.L.; Dooley, H.; Verdino, P.
2007-07-13
Sharks express an unusual heavy-chain isotype called IgNAR, whose variable regions bind antigen as independent soluble domains. To further probe affinity maturation of the IgNAR response, we structurally characterized the germline and somatically matured versions of a type II variable (V) region, both in the presence and absence of its antigen, hen egg-white lysozyme. Despite a disulfide bond linking complementarity determining regions (CDRs) 1 and 3, both germline and somatically matured V regions displayed significant structural changes in these CDRs upon complex formation with antigen. Somatic mutations in the IgNAR V region serve to increase the number of contacts withmore » antigen, as reflected by a tenfold increase in affinity, and one of these mutations appears to stabilize the CDR3 region. In addition, a residue in the HV4 loop plays an important role in antibody-antigen interaction, consistent with the high rate of somatic mutations in this non-CDR loop.« less
-
Okazaki, Fumiyoshi; Aoki, Jun-ichi; Tabuchi, Soichiro; Tanaka, Tsutomu; Ogino, Chiaki; Kondo, Akihiko
2012-10-01
We have constructed a filamentous fungus Aspergillus oryzae that secretes a llama variable heavy-chain antibody fragment (V(HH)) that binds specifically to epidermal growth factor receptor (EGFR) in a culture medium. A major improvement in yield was achieved by fusing the V(HH) with a Taka-amylase A signal sequence (sTAA) and a segment of 28 amino acids from the N-terminal region of Rhizopus oryzae lipase (N28). The yields of secreted, immunologically active anti-EGFR V(HH) reached 73.8 mg/1 in a Sakaguchi flask. The V(HH) fragments were released from the sTAA or N28 proteins by an indigenous A. oryzae protease during cultivation. The purified recombinant V(HH) fragment was specifically recognized and could bind to the EGFR with a high affinity.
-
Levi, M; Sällberg, M; Rudén, U; Herlyn, D; Maruyama, H; Wigzell, H; Marks, J; Wahren, B
1993-01-01
A complementarity-determining region (CDR) of the mouse monoclonal antibody (mAb) F58 was constructed with specificity to a neutralization-inducing region of human immunodeficiency virus type 1 (HIV-1). The mAb has its major reactivity to the amino acid sequence I--GPGRA in the V3 viral envelope region. All CDRs including several framework amino acids were synthesized from the sequence deduced by cloning and sequencing mAb F58 heavy- and light-chain variable domains. Peptides derived from the third heavy-chain domain (CDR-H3) alone or in combination with the other CDR sequences competed with F58 mAb for the V3 region. The CDR-H3 peptide was chemically modified by cyclization and then inhibited HIV-1 replication as well as syncytium formation by infected cells. Both the homologous IIIB viral strain to which the F58 mAb was induced and the heterologous SF2 strain were inhibited. This synthetic peptide had unexpectedly potent antiviral activity and may be a potential tool for treatment of HIV-infected persons. PMID:7685100
-
Effects of Variable Resistance Using Chains on Bench Throw Performance in Trained Rugby Players.
Godwin, Mark S; Fernandes, John F T; Twist, Craig
2018-04-01
Godwin, MS, Fernandes, JFT, and Twist, C. Effects of variable resistance using chains on bench throw performance in trained rugby players. J Strength Cond Res 32(4): 950-954, 2018-This study sought to determine the effects of variable resistance using chain resistance on bench throw performance. Eight male rugby union players (19.4 ± 2.3 years, 88.8 ± 6.0 kg, 1RM 105.6 ± 17.0 kg) were recruited from a national league team. In a randomized crossover design, participant's performed 3 bench throws at 45% one repetition maximum (1RM) at a constant load (no chains) or a variable load (30% 1RM constant load and 15% 1RM variable load; chains) with 7 days between conditions. For each repetition, the peak and mean velocity, peak power, peak acceleration, and time to peak velocity were recorded. Differences in peak and mean power were very likely trivial and unclear between the chain and no chain conditions, respectively. Possibly greater peak and likely greater mean bar velocity were accompanied by likely to most likely greater bar velocity between 50 and 400 ms from initiation of bench press in the chain condition compared with the no chain condition. Accordingly, bar acceleration was very likely greater in the chain condition compared with the no chain condition. In conclusion, these results show that the inclusion of chain resistance can acutely enhance several variables in the bench press throw and gives support to this type of training.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Kipps, T.J.; Fong, S.; Tomhave, E.
Malignant B lymphocytes from several patients with chronic lymphocytic leukemia (CLL) were examined for reactivity with murine monoclonal antibody 17.109. This antibody, prepared against the rheumatoid factor (RF) paraprotein Sie, recognizes a cross reactive idiotype on 48% of human IgM RF paraproteins, but does not react with IgM paraproteins without RF activity or substantially with normal pooled immunoglobulin. The 17.109-reactive idiotype is a marker for a kappa III variable-region gene, designated V/sub kappa/RF, that is conserved in outbred human populations. In a limited study of 31 CLL patients, the leukemic cells from 5 of 20 patients with kappa light chain-expressingmore » CLL were recognized by the 17.109 monoclonal antibody. Despite having malignant cells specifically reactive with this antibody, patients with 17.109-positive CLL did not have elevated serum levels of circulating antibody bearing 17.109-reactive determinants. Total RNAs isolated from the CLL B lymphocytes, or from hybridomas produced by fusing the CLL cells with the WI-L2-729-HF/sub 2/ cell line, were fractionated electrophoretically and examined by blot hybridization. Under stringent hybridization conditions capable of discerning a single base-pair mismatch, RNA from the 17.109-idiotype-positive CLL cells hybridized to synthetic oligonucleotide probes corresponding to framework and complementary-determining regions in the V/sub kappa/RF gene. The high frequency of the 17.109-associated idiotype and the V/sub kappa/RF gene in CLL suggests that the disease may arise from B lymphocytes that express a restricted set of inherited immunoglobulin variable-region genes with little or no somatic mutation.« less
-
Cargo selection by specific kinesin light chain 1 isoforms.
Woźniak, Marcin J; Allan, Victoria J
2006-11-29
Kinesin-1 drives the movement of diverse cargoes, and it has been proposed that specific kinesin light chain (KLC) isoforms target kinesin-1 to these different structures. Here, we test this hypothesis using two in vitro motility assays, which reconstitute the movement of rough endoplasmic reticulum (RER) and vesicles present in a Golgi membrane fraction. We generated GST-tagged fusion proteins of KLC1B and KLC1D that included the tetratricopeptide repeat domain and the variable C-terminus. We find that preincubation of RER with KLC1B inhibits RER motility, whereas KLC1D does not. In contrast, Golgi fraction vesicle movement is inhibited by KLC1D but not KLC1B reagents. Both RER and vesicle movement is inhibited by preincubation with the GST-tagged C-terminal domain of ubiquitous kinesin heavy chain (uKHC), which binds to the N-terminal domain of uKHC and alters its interaction with microtubules. We propose that although the TRR domains are required for cargo binding, it is the variable C-terminal region of KLCs that are vital for targeting kinesin-1 to different cellular structures.
-
Cargo selection by specific kinesin light chain 1 isoforms
Woźniak, Marcin J; Allan, Victoria J
2006-01-01
Kinesin-1 drives the movement of diverse cargoes, and it has been proposed that specific kinesin light chain (KLC) isoforms target kinesin-1 to these different structures. Here, we test this hypothesis using two in vitro motility assays, which reconstitute the movement of rough endoplasmic reticulum (RER) and vesicles present in a Golgi membrane fraction. We generated GST-tagged fusion proteins of KLC1B and KLC1D that included the tetratricopeptide repeat domain and the variable C-terminus. We find that preincubation of RER with KLC1B inhibits RER motility, whereas KLC1D does not. In contrast, Golgi fraction vesicle movement is inhibited by KLC1D but not KLC1B reagents. Both RER and vesicle movement is inhibited by preincubation with the GST-tagged C-terminal domain of ubiquitous kinesin heavy chain (uKHC), which binds to the N-terminal domain of uKHC and alters its interaction with microtubules. We propose that although the TRR domains are required for cargo binding, it is the variable C-terminal region of KLCs that are vital for targeting kinesin-1 to different cellular structures. PMID:17093494
-
cDNA sequences and organization of IgM heavy chain genes in two holostean fish.
Wilson, M R; van Ravenstein, E; Miller, N W; Clem, L W; Middleton, D L; Warr, G W
1995-01-01
Immunoglobulin M heavy chain (mu) sequences of two holostean fish, the bowfin, Amia calva, and the longnose gar, Lepisosteus osseus, were amplified from spleen mRNA by RACE-PCR, cloned, and sequenced. Each mu chain showed the conserved four constant domain structure typical of a secreted mu chain. Southern blot analyses with specific heavy chain variable (VH) and constant (CH) region probes suggest that both fish possess an IgH locus that resembles that of the teleosts, amphibians, and mammals in its organization. The overall sequence similarity of gar and bowfin mu chains was 60% and 48% at the nucleotide and amino acid levels, respectively, while similarity to the mu chains of teleosts and elasmobranchs was lower. The bowfin mu chain possesses a distinctive proline-rich sequence at the C mu 1/C mu 2 boundary; a shorter proline-rich sequence is present at this position in the gar mu chain. Both gar and bowfin show, in their C mu 4 sequences, motifs that could serve as cryptic splice donor sites for the production of mRNA encoding the membrane-bound form of the mu chains, and the bowfin also shows a potential cryptic splice donor site in the C mu 3 exon.
-
INDUCTION OF RABBIT ANTIBODY WITH MOLECULAR UNIFORMITY AFTER IMMUNIZATION WITH GROUP C STREPTOCOCCI
Eichmann, Klaus; Lackland, Henry; Hood, Leroy; Krause, Richard M.
1970-01-01
Antibodies with uniform properties may occur in rabbits after immunization with Group C streptococci. These precipitating antibodies possess specificity for the group-specific carbohydrate. Not uncommonly, their concentration is between 20 and 40 mg/ml of antiserum. Evidence for molecular uniformity in the case of one of these antibodies, described in detail here, includes: individual antigenic specificity; monodisperse distribution of the light chains by alkaline urea polyacrylamide disc electrophoresis; and a single amino acid in each of the first three N-terminal positions of the light chains. When the amino acid sequence of rabbit antibody b+ light chains (κ type) are aligned against their human κ counterparts, a definite homology is observed between the N-terminus of the human and the rabbit variable region. PMID:5409946
-
Domier, L L; Latorre, I J; Steinlage, T A; McCoppin, N; Hartman, G L
2003-10-01
The variability of North American and Asian strains and isolates of Soybean mosaic virus was investigated. First, polymerase chain reaction (PCR) products representing the coat protein (CP)-coding regions of 38 SMVs were analyzed for restriction fragment length polymorphisms (RFLP). Second, the nucleotide and predicted amino acid sequence variability of the P1-coding region of 18 SMVs and the helper component/protease (HC/Pro) and CP-coding regions of 25 SMVs were assessed. The CP nucleotide and predicted amino acid sequences were the most similar and predicted phylogenetic relationships similar to those obtained from RFLP analysis. Neither RFLP nor sequence analyses of the CP-coding regions grouped the SMVs by geographical origin. The P1 and HC/Pro sequences were more variable and separated the North American and Asian SMV isolates into two groups similar to previously reported differences in pathogenic diversity of the two sets of SMV isolates. The P1 region was the most informative of the three regions analyzed. To assess the biological relevance of the sequence differences in the HC/Pro and CP coding regions, the transmissibility of 14 SMV isolates by Aphis glycines was tested. All field isolates of SMV were transmitted efficiently by A. glycines, but the laboratory isolates analyzed were transmitted poorly. The amino acid sequences from most, but not all, of the poorly transmitted isolates contained mutations in the aphid transmission-associated DAG and/or KLSC amino acid sequence motifs of CP and HC/Pro, respectively.
-
Skvortsov, Alexander M; Klushin, Leonid I; Polotsky, Alexey A; Binder, Kurt
2012-03-01
The phase transition occurring when a single polymer chain adsorbed at a planar solid surface is mechanically desorbed is analyzed in two statistical ensembles. In the force ensemble, a constant force applied to the nongrafted end of the chain (that is grafted at its other end) is used as a given external control variable. In the z-ensemble, the displacement z of this nongrafted end from the surface is taken as the externally controlled variable. Basic thermodynamic parameters, such as the adsorption energy, exhibit a very different behavior as a function of these control parameters. In the thermodynamic limit of infinite chain length the desorption transition with the force as a control parameter clearly is discontinuous, while in the z-ensemble continuous variations are found. However, one should not be misled by a too-naive application of the Ehrenfest criterion to consider the transition as a continuous transition: rather, one traverses a two-phase coexistence region, where part of the chain is still adsorbed and the other part desorbed and stretched. Similarities with and differences from two-phase coexistence at vapor-liquid transitions are pointed out. The rounding of the singularities due to finite chain length is illustrated by exact calculations for the nonreversal random walk model on the simple cubic lattice. A new concept of local order parameter profiles for the description of the mechanical desorption of adsorbed polymers is suggested. This concept give evidence for both the existence of two-phase coexistence within single polymer chains for this transition and the anomalous character of this two-phase coexistence. Consequences for the proper interpretation of experiments performed in different ensembles are briefly mentioned.
-
Culshaw, Abigail; Ladell, Kristin; Gras, Stephanie; McLaren, James E; Miners, Kelly L; Farenc, Carine; van den Heuvel, Heleen; Gostick, Emma; Dejnirattisai, Wanwisa; Wangteeraprasert, Apirath; Duangchinda, Thaneeya; Chotiyarnwong, Pojchong; Limpitikul, Wannee; Vasanawathana, Sirijitt; Malasit, Prida; Dong, Tao; Rossjohn, Jamie; Mongkolsapaya, Juthathip; Price, David A; Screaton, Gavin R
2017-11-01
Adaptive immune responses protect against infection with dengue virus (DENV), yet cross-reactivity with distinct serotypes can precipitate life-threatening clinical disease. We found that clonotypes expressing the T cell antigen receptor (TCR) β-chain variable region 11 (TRBV11-2) were 'preferentially' activated and mobilized within immunodominant human-leukocyte-antigen-(HLA)-A*11:01-restricted CD8 + T cell populations specific for variants of the nonstructural protein epitope NS3 133 that characterize the serotypes DENV1, DENV3 and DENV4. In contrast, the NS3 133 -DENV2-specific repertoire was largely devoid of such TCRs. Structural analysis of a representative TRBV11-2 + TCR demonstrated that cross-serotype reactivity was governed by unique interplay between the variable antigenic determinant and germline-encoded residues in the second β-chain complementarity-determining region (CDR2β). Extensive mutagenesis studies of three distinct TRBV11-2 + TCRs further confirmed that antigen recognition was dependent on key contacts between the serotype-defined peptide and discrete residues in the CDR2β loop. Collectively, these data reveal an innate-like mode of epitope recognition with potential implications for the outcome of sequential exposure to heterologous DENVs.
-
Sayegh, Camil E.; Demaries, Sandra L.; Iacampo, Sandra; Ratcliffe, Michael J. H.
1999-01-01
Immunoglobulin gene rearrangement in avian B cell precursors generates surface Ig receptors of limited diversity. It has been proposed that specificities encoded by these receptors play a critical role in B lineage development by recognizing endogenous ligands within the bursa of Fabricius. To address this issue directly we have introduced a truncated surface IgM, lacking variable region domains, into developing B precursors by retroviral gene transfer in vivo. Cells expressing this truncated receptor lack endogenous surface IgM, and the low level of endogenous Ig rearrangements that have occurred within this population of cells has not been selected for having a productive reading frame. Such cells proliferate rapidly within bursal epithelial buds of normal morphology. In addition, despite reduced levels of endogenous light chain rearrangement, those light chain rearrangements that have occurred have undergone variable region diversification by gene conversion. Therefore, although surface expression of an Ig receptor is required for bursal colonization and the induction of gene conversion, the specificity encoded by the prediversified receptor is irrelevant and, consequently, there is no obligate ligand for V(D)J-encoded determinants of prediversified avian cell surface IgM receptor. PMID:10485907
-
Brown, Darin J; Stefan, Sarah E; Berden, Giel; Steill, Jeffrey D; Oomens, Jos; Eyler, John R; Bendiak, Brad
2011-11-08
All eight D-aldohexoses and aldohexoses derived from the non-reducing end of disaccharides were investigated by variable-wavelength infrared multiple-photon dissociation (IRMPD) as anions in the negative-ion mode. Spectroscopic evidence supports the existence of a relatively abundant open-chain configuration of the anions in the gas phase, based on the observation of a significant carbonyl absorption band near 1710 cm(-1). The abundance of the open-chain configuration of the aldohexose anions was approximately 1000-fold or greater than that of the neutral sugars in aqueous solution. This provides an explanation as to why it has not been possible to discriminate the anomeric configuration of aldohexose anions in the gas phase when derived from the non-reducing sugar of a disaccharide. Evidence from photodissociation spectra also indicates that the different aldohexoses yield product ions with maximal abundances at different wavelengths, and that the carbonyl stretch region is useful for differentiation of sugar stereochemistries. Quantum-chemical calculations indicate relatively low energy barriers to intramolecular proton transfer between hydroxyl groups and adjacent alkoxy sites located on open-chain sugar anions, suggesting that an ensemble of alkoxy charge locations contributes to their observed photodissociation spectra. Ring opening of monosaccharide anions and interconversion among configurations is an inherent property of the ions themselves and occurs in vacuo independent of solvent participation. Copyright © 2011 Elsevier Ltd. All rights reserved.
-
Bradbury, Andrew R. M.; Trinklein, Nathan D.; Wilkinson, Ian C.; Tandon, Atul K.; Anderson, Stephen; Bladen, Catherine L.; Jones, Brittany; Aldred, Shelley Force; Bestagno, Marco; Burrone, Oscar; Maynard, Jennifer; Ferrara, Fortunato; Görnemann, Janina; Glanville, Jacob; Wolf, Philipp; Frenzel, Andre; Wong, Julin; Koh, Xin Yu; Eng, Hui-Yan; Lane, David; Lefranc, Marie-Paule; Clark, Mike
2018-01-01
ABSTRACT Monoclonal antibodies are commonly assumed to be monospecific, but anecdotal studies have reported genetic diversity in antibody heavy chain and light chain genes found within individual hybridomas. As the prevalence of such diversity has never been explored, we analyzed 185 random hybridomas, in a large multicenter dataset. The hybridomas analyzed were not biased towards those with cloning difficulties or known to have additional chains. Of the hybridomas we evaluated, 126 (68.1%) contained no additional productive chains, while the remaining 59 (31.9%) contained one or more additional productive heavy or light chains. The expression of additional chains degraded properties of the antibodies, including specificity, binding signal and/or signal-to-noise ratio, as determined by enzyme-linked immunosorbent assay and immunohistochemistry. The most abundant mRNA transcripts found in a hybridoma cell line did not necessarily encode the antibody chains providing the correct specificity. Consequently, when cloning antibody genes, functional validation of all possible VH and VL combinations is required to identify those with the highest affinity and lowest cross-reactivity. These findings, reflecting the current state of hybridomas used in research, reiterate the importance of using sequence-defined recombinant antibodies for research or diagnostic use. PMID:29485921
-
Influence of land use and climate on wetland breeding birds in the Prairie Pothole region of Canada
Forcey, G.M.; Linz, G.M.; Thogmartin, W.E.; Bleier, W.J.
2007-01-01
Bird populations are influenced by a variety of factors at both small and large scales that range from the presence of suitable nesting habitat, predators, and food supplies to climate conditions and land-use patterns. We evaluated the influences of regional climate and land-use variables on wetland breeding birds in the Canada section of Bird Conservation Region 11 (CA-BCR11), the Prairie Potholes. We used bird abundance data from the North American Breeding Bird Survey, land-use data from the Prairie Farm Rehabilitation Administration, and weather data from the National Climatic Data and Information Archive to model effects of regional environmental variables on bird abundance. Models were constructed a priori using information from published habitat associations in the literature, and fitting was performed with WinBUGS using Markov chain Monte Carlo techniques. Both land-use and climate variables contributed to predicting bird abundance in CA-BCR11, although climate predictors contributed the most to improving model fit. Examination of regional effects of climate and land use on wetland birds in CA-BCR11 revealed relationships with environmental covariates that are often overlooked by small-scale habitat studies. Results from these studies can be used to improve conservation and management planning for regional populations of avifauna. ?? 2007 NRC.
-
[Research of Human-mouse Chimeric Antibodies Against Ebola Virus Nucleoprotein].
Zhou, Rongping; Sun, Lina; Liu, Yang; Wu, Wei; Li, Chuan; Liang, Mifang; Qiu, Peihong
2016-01-01
The Ebola virus is highly infectious and can result in death in ≤ 90% of infected subjects. Detection of the Ebola virus and diagnosis of infection are extremely important for epidemic control. Presently, Chinese laboratories detect the nucleic acids of the Ebola virus by real-time reverse transcription-polymerase chain reaction (RT-PCR). However, such detection takes a relatively long time and necessitates skilled personnel and expensive equipment. Enzyme-linked immunosorbent assay (ELISA) of serum is simple, easy to operate, and can be used to ascertain if a patient is infected with the Ebola virus as well as the degree of infection. Hence, ELISA can be used in epidemiological investigations and is a strong complement to detection of nucleic acids. Cases of Ebola hemorrhagic fever have not been documented in China, so quality-control material for positive serology is needed. Construction and expression of human-mouse chimeric antibodies against the nucleoprotein of the Ebola virus was carried out. Genes encoding variable heavy (VH) and variable light (VL) chains were extracted and amplified from murine hybridoma cells. Genes encoding the VH and VL chains of monoclonal antibodies were amplified by RT-PCR. According to sequence analyses, a primer was designed to amplify functional sequences relative to VH and VL chain. The eukaryotic expression vector HL51-14 carrying some human antibody heavy chain- and light chain-constant regions was used. IgG antibodies were obtained by transient transfection of 293T cells. Subsequently, immunological detection and immunological identification were identified by ELISA, immunofluorescence assay, and western blotting. These results showed that we constructed and purified two human- mouse chimeric antibodies.
-
Same-Day Imaging Using Small Proteins: Clinical Experience and Translational Prospects in Oncology.
Krasniqi, Ahmet; D'Huyvetter, Matthias; Devoogdt, Nick; Frejd, Fredrik Y; Sörensen, Jens; Orlova, Anna; Keyaerts, Marleen; Tolmachev, Vladimir
2018-06-01
Imaging of expression of therapeutic targets may enable stratification of patients for targeted treatments. The use of small radiolabeled probes based on the heavy-chain variable region of heavy-chain-only immunoglobulins or nonimmunoglobulin scaffolds permits rapid localization of radiotracers in tumors and rapid clearance from normal tissues. This makes high-contrast imaging possible on the day of injection. This mini review focuses on small proteins for radionuclide-based imaging that would allow same-day imaging, with the emphasis on clinical applications and promising preclinical developments within the field of oncology. © 2018 by the Society of Nuclear Medicine and Molecular Imaging.
-
DeKosky, Brandon J; Lungu, Oana I; Park, Daechan; Johnson, Erik L; Charab, Wissam; Chrysostomou, Constantine; Kuroda, Daisuke; Ellington, Andrew D; Ippolito, Gregory C; Gray, Jeffrey J; Georgiou, George
2016-05-10
Elucidating how antigen exposure and selection shape the human antibody repertoire is fundamental to our understanding of B-cell immunity. We sequenced the paired heavy- and light-chain variable regions (VH and VL, respectively) from large populations of single B cells combined with computational modeling of antibody structures to evaluate sequence and structural features of human antibody repertoires at unprecedented depth. Analysis of a dataset comprising 55,000 antibody clusters from CD19(+)CD20(+)CD27(-) IgM-naive B cells, >120,000 antibody clusters from CD19(+)CD20(+)CD27(+) antigen-experienced B cells, and >2,000 RosettaAntibody-predicted structural models across three healthy donors led to a number of key findings: (i) VH and VL gene sequences pair in a combinatorial fashion without detectable pairing restrictions at the population level; (ii) certain VH:VL gene pairs were significantly enriched or depleted in the antigen-experienced repertoire relative to the naive repertoire; (iii) antigen selection increased antibody paratope net charge and solvent-accessible surface area; and (iv) public heavy-chain third complementarity-determining region (CDR-H3) antibodies in the antigen-experienced repertoire showed signs of convergent paired light-chain genetic signatures, including shared light-chain third complementarity-determining region (CDR-L3) amino acid sequences and/or Vκ,λ-Jκ,λ genes. The data reported here address several longstanding questions regarding antibody repertoire selection and development and provide a benchmark for future repertoire-scale analyses of antibody responses to vaccination and disease.
-
Zhang, B; Evans, J S
2001-01-01
Molecular elasticity is associated with a select number of polypeptides and proteins, such as titin, Lustrin A, silk fibroin, and spider silk dragline protein. In the case of titin, the globular (Ig) and non-globular (PEVK) regions act as extensible springs under stretch; however, their unfolding behavior and force extension characteristics are different. Using our time-dependent macroscopic method for simulating AFM-induced titin Ig domain unfolding and refolding, we simulate the extension and relaxation of hypothetical titin chains containing Ig domains and a PEVK region. Two different models are explored: 1) a series-linked WLC expression that treats the PEVK region as a distinct entropic spring, and 2) a summation of N single WLC expressions that simulates the extension and release of a discrete number of parallel titin chains containing constant or variable amounts of PEVK. In addition to these simulations, we also modeled the extension of a hypothetical PEVK domain using a linear Hooke's spring model to account for "enthalpic" contributions to PEVK elasticity. We find that the modified WLC simulations feature chain length compensation, Ig domain unfolding/refolding, and force-extension behavior that more closely approximate AFM, laser tweezer, and immunolocalization experimental data. In addition, our simulations reveal the following: 1) PEVK extension overlaps with the onset of Ig domain unfolding, and 2) variations in PEVK content within a titin chain ensemble lead to elastic diversity within that ensemble. PMID:11159428
-
Peptide docking of HIV-1 p24 with single chain fragment variable (scFv) by CDOCKER algorithm
NASA Astrophysics Data System (ADS)
Karim, Hana Atiqah Abdul; Tayapiwatana, Chatchai; Nimmanpipug, Piyarat; Zain, Sharifuddin M.; Rahman, Noorsaadah Abdul; Lee, Vannajan Sanghiran
2014-10-01
In search for the important residues that might have involve in the binding interaction between the p24 caspid protein of HIV-1 fragment (MET68 - PRO90) with the single chain fragment variable (scFv) of FAB23.5, modern computational chemistry approach has been conducted and applied. The p24 fragment was initially taken out from the 1AFV protein molecule consisting of both light (VL) and heavy (VH) chains of FAB23.5 as well as the HIV-1 caspid protein. From there, the p24 (antigen) fragment was made to dock back into the protein pocket receptor (antibody) by using the CDOCKER algorithm to conduct the molecular docking process. The score calculated from the CDOCKER gave 15 possible docked poses with various docked ligand's positions, the interaction energy as well as the binding energy. The best docked pose that imitates the original antigen's position was determined and further processed to the In Situ minimization to obtain the residues interaction energy as well as to observe the hydrogen bonds interaction in the protein-peptide complex. Based on the results demonstrated, the specific residues in the complex that have shown immense lower interaction energies in the 5Å vicinity region from the peptide are from the heavy chain (VH:TYR105) and light chain (VL: ASN31, TYR32, and GLU97). Those residues play vital roles in the binding mechanism of Antibody-Antigen (Ab-Ag) complex of p24 with FAB23.5.
-
Injectable Absorbable Ocular Inserts for Controlled Drug Delivery
1997-07-01
conjunctiva for prolonged delivery of drugs to the anterior region of the eye (Gwon & Meadows, 1992). The dosage system was an elliptically shaped unit...1979) have reviewed many other gel formers which are available for preparing pharmaceutical gels. A.3.4.1. Hydrogels -- Hydrogels are materials which...denoted as hydrogels (or aquagels). Hydrogels based on crosslinked polymeric chains of methoxy poly(ethylene glycol) monomethacrylate having variable
-
Tertiary structure of human {Lambda}6 light chains.
DOE Office of Scientific and Technical Information (OSTI.GOV)
Pokkuluri, P. R.; Solomon, A.; Weiss, D. T.
1999-01-01
AL amyloidosis is a disease process characterized by the pathologic deposition of monoclonal light chains in tissue. To date, only limited information has been obtained on the molecular features that render such light chains amyloidogenic. Although protein products of the major human V kappa and V lambda gene families have been identified in AL deposits, one particular subgroup--lambda 6--has been found to be preferentially associated with this disease. Notably, the variable region of lambda 6 proteins (V lambda 6) has distinctive primary structural features including the presence in the third framework region (FR3) of two additional amino acid residues thatmore » distinguish members of this subgroup from other types of light chains. However, the structural consequences of these alterations have not been elucidated. To determine if lambda 6 proteins possess unique tertiary structural features, as compared to light chains of other V lambda subgroups, we have obtained x-ray diffraction data on crystals prepared from two recombinant V lambda 6 molecules. These components, isolated from a bacterial expression system, were generated from lambda 6-related cDNAs cloned from bone marrow-derived plasma cells from a patient (Wil) who had documented AL amyloidosis and another (Jto) with multiple myeloma and tubular cast nephropathy, but no evident fibrillar deposits. The x-ray crystallographic analyses revealed that the two-residue insertion located between positions 68 and 69 (not between 66 and 67 as previously surmised) extended an existing loop region that effectively increased the surface area adjacent to the first complementarity determining region (CDR1). Further, an unusual interaction between the Arg 25 and Phe 2 residues commonly found in lambda 6 molecules was noted. However, the structures of V lambda 6 Wil and Jto also differed from each other, as evidenced by the presence in the latter of certain ionic and hydrophobic interactions that we posit increased protein stability and thus prevented amyloid formation.« less
-
Estimating variability in grain legume yields across Europe and the Americas
NASA Astrophysics Data System (ADS)
Cernay, Charles; Ben-Ari, Tamara; Pelzer, Elise; Meynard, Jean-Marc; Makowski, David
2015-06-01
Grain legume production in Europe has recently come under scrutiny. Although legume crops are often promoted to provide environmental services, European farmers tend to turn to non-legume crops. It is assumed that high variability in legume yields explains this aversion, but so far this hypothesis has not been tested. Here, we estimate the variability of major grain legume and non-legume yields in Europe and the Americas from yield time series over 1961-2013. Results show that grain legume yields are significantly more variable than non-legume yields in Europe. These differences are smaller in the Americas. Our results are robust at the level of the statistical methods. In all regions, crops with high yield variability are allocated to less than 1% of cultivated areas. Although the expansion of grain legumes in Europe may be hindered by high yield variability, some species display risk levels compatible with the development of specialized supply chains.
-
Somatic diversification of chicken immunoglobulin light chains by point mutations.
Parvari, R; Ziv, E; Lantner, F; Heller, D; Schechter, I
1990-04-01
The light-chain locus of chicken has 1 functional V lambda 1 gene, 1 J gene, and 25 pseudo-V lambda-genes (where V = variable and J = joining). A major problem is which somatic mechanisms expand this extremely limited germ-line information to generate many different antibodies. Weill's group [Reynaud, C. A., Anquez, V., Grimal, H. & Weill, J. C. (1987) Cell 48, 379-388] has shown that the pseudo-V lambda-genes diversify the rearranged V lambda 1 by gene conversion. Here we demonstrate that chicken light chains are further diversified by somatic point mutations and by V lambda 1-J flexible joining. Somatic point mutations were identified in the J and 3' noncoding DNA of rearranged light-chain genes of chicken. These regions were analyzed because point mutations in V lambda 1 are obscured by gene conversion; the J and 3' noncoding DNA are presented in one copy per haploid genome and are not subject to gene conversion. In rodents point mutations occur as frequently in the V-J coding regions as in the adjacent flanking DNA. Therefore, we conclude that somatic point mutations diversify the V lambda 1 of chicken. The frequency (0-1%) and distribution of the mutations (decreasing in number with increased distance from the V lambda 1 segment) in chicken were as observed in rodents. Sequence variability at the V lambda 1-J junctions could be attributed to imprecise joining of the V lambda 1 and J genes. The modification by gene conversion of rearranged V lambda 1 genes in the bursa was similar in chicken aged 3 months (9.5%) or 3 weeks (9.1%)--i.e., gene conversion that generates the preimmune repertoire in the bursa seems to level off around 3 weeks of age. This preimmune repertoire can be further diversified by somatic point mutations that presumably lead to the formation of antibodies with increased affinity. A segment with structural features of a matrix association region [(A + T)-rich and four topoisomerase II binding sites] was identified in the middle of the J-C lambda intron (where C = constant).
-
Aliesky, Holly; Banuelos, Bianca; Magana, Jessica; Williams, Robert W.; Rapoport, Basil
2014-01-01
Graves' hyperthyroidism is caused by antibodies to the TSH receptor (TSHR) that mimic thyroid stimulation by TSH. Stimulating TSHR antibodies and hyperthyroidism can be induced by immunizing mice with adenovirus expressing the human TSHR A-subunit. Prior analysis of induced Graves' disease in small families of recombinant inbred (RI) female mice demonstrated strong genetic control but did not resolve trait loci for TSHR antibodies or elevated serum T4. We investigated the genetic basis for induced Graves' disease in female mice of two large RI families and combined data with earlier findings to provide phenotypes for 178 genotypes. TSHR antibodies measured by inhibition of TSH binding to its receptor were highly significantly linked in the BXD set to the major histocompatibility region (chromosome 17), consistent with observations in 3 other RI families. In the LXS family, we detected linkage between T4 levels after TSHR-adenovirus immunization and the Ig heavy chain variable region (Igvh, chromosome 12). This observation is a key finding because components of the antigen binding region of Igs determine antibody specificity and have been previously linked to induced thyroid-stimulating antibodies. Data from the LXS family provide the first evidence in mice of a direct link between induced hyperthyroidism and Igvh genes. A role for major histocompatibility genes has now been established for genetic susceptibility to Graves' disease in both humans and mice. Future studies using arrays incorporating variation in the complex human Ig gene locus will be necessary to determine whether Igvh genes are also linked to Graves' disease in humans. PMID:25051451
-
Roux, K H; Greenberg, A S; Greene, L; Strelets, L; Avila, D; McKinney, E C; Flajnik, M F
1998-09-29
We recently have identified an antigen receptor in sharks called NAR (new or nurse shark antigen receptor) that is secreted by splenocytes but does not associate with Ig light (L) chains. The NAR variable (V) region undergoes high levels of somatic mutation and is equally divergent from both Ig and T cell receptors (TCR). Here we show by electron microscopy that NAR V regions, unlike those of conventional Ig and TCR, do not form dimers but rather are independent, flexible domains. This unusual feature is analogous to bona fide camelid IgG in which modifications of Ig heavy chain V (VH) sequences prevent dimer formation with L chains. NAR also displays a uniquely flexible constant (C) region. Sequence analysis and modeling show that there are only two types of expressed NAR genes, each having different combinations of noncanonical cysteine (Cys) residues in the V domains that likely form disulfide bonds to stabilize the single antigen-recognition unit. In one NAR class, rearrangement events result in mature genes encoding an even number of Cys (two or four) in complementarity-determining region 3 (CDR3), which is analogous to Cys codon expression in an unusual human diversity (D) segment family. The NAR CDR3 Cys generally are encoded by preferred reading frames of rearranging D segments, providing a clear design for use of preferred reading frame in antigen receptor D regions. These unusual characteristics shared by NAR and unconventional mammalian Ig are most likely the result of convergent evolution at the molecular level.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Cafferty, Kara G.; Searcy, Erin M.; Nguyen, Long
To meet Energy Independence and Security Act (EISA) cellulosic biofuel mandates, the United States will require an annual domestic supply of about 242 million Mg of biomass by 2022. To improve the feedstock logistics of lignocellulosic biofuels and access available biomass resources from areas with varying yields, commodity systems have been proposed and designed to deliver on-spec biomass feedstocks at preprocessing “depots”, which densify and stabilize the biomass prior to long-distance transport and delivery to centralized biorefineries. The harvesting, preprocessing, and logistics (HPL) of biomass commodity supply chains thus could introduce spatially variable environmental impacts into the biofuel life cyclemore » due to needing to harvest, move, and preprocess biomass from multiple distances that have variable spatial density. This study examines the uncertainty in greenhouse gas (GHG) emissions of corn stover logisticsHPL within a bio-ethanol supply chain in the state of Kansas, where sustainable biomass supply varies spatially. Two scenarios were evaluated each having a different number of depots of varying capacity and location within Kansas relative to a central commodity-receiving biorefinery to test GHG emissions uncertainty. Monte Carlo simulation was used to estimate the spatial uncertainty in the HPL gate-to-gate sequence. The results show that the transport of densified biomass introduces the highest variability and contribution to the carbon footprint of the logistics HPL supply chain (0.2-13 g CO 2e/MJ). Moreover, depending upon the biomass availability and its spatial density and surrounding transportation infrastructure (road and rail), logistics HPL processes can increase the variability in life cycle environmental impacts for lignocellulosic biofuels. Within Kansas, life cycle GHG emissions could range from 24 to 41 g CO 2e/MJ depending upon the location, size and number of preprocessing depots constructed. However, this range can be minimized through optimizing the siting of preprocessing depots where ample rail infrastructure exists to supply biomass commodity to a regional biorefinery supply system« less
-
Cafferty, Kara G.; Searcy, Erin M.; Nguyen, Long; ...
2014-11-04
To meet Energy Independence and Security Act (EISA) cellulosic biofuel mandates, the United States will require an annual domestic supply of about 242 million Mg of biomass by 2022. To improve the feedstock logistics of lignocellulosic biofuels and access available biomass resources from areas with varying yields, commodity systems have been proposed and designed to deliver on-spec biomass feedstocks at preprocessing “depots”, which densify and stabilize the biomass prior to long-distance transport and delivery to centralized biorefineries. The harvesting, preprocessing, and logistics (HPL) of biomass commodity supply chains thus could introduce spatially variable environmental impacts into the biofuel life cyclemore » due to needing to harvest, move, and preprocess biomass from multiple distances that have variable spatial density. This study examines the uncertainty in greenhouse gas (GHG) emissions of corn stover logisticsHPL within a bio-ethanol supply chain in the state of Kansas, where sustainable biomass supply varies spatially. Two scenarios were evaluated each having a different number of depots of varying capacity and location within Kansas relative to a central commodity-receiving biorefinery to test GHG emissions uncertainty. Monte Carlo simulation was used to estimate the spatial uncertainty in the HPL gate-to-gate sequence. The results show that the transport of densified biomass introduces the highest variability and contribution to the carbon footprint of the logistics HPL supply chain (0.2-13 g CO 2e/MJ). Moreover, depending upon the biomass availability and its spatial density and surrounding transportation infrastructure (road and rail), logistics HPL processes can increase the variability in life cycle environmental impacts for lignocellulosic biofuels. Within Kansas, life cycle GHG emissions could range from 24 to 41 g CO 2e/MJ depending upon the location, size and number of preprocessing depots constructed. However, this range can be minimized through optimizing the siting of preprocessing depots where ample rail infrastructure exists to supply biomass commodity to a regional biorefinery supply system« less
-
Marini, Joan C.; Forlino, Antonella; Cabral, Wayne A.; Barnes, Aileen M.; San Antonio, James D.; Milgrom, Sarah; Hyland, James C.; Körkkö, Jarmo; Prockop, Darwin J.; De Paepe, Anne; Coucke, Paul; Symoens, Sofie; Glorieux, Francis H.; Roughley, Peter J.; Lund, Alan M.; Kuurila-Svahn, Kaija; Hartikka, Heini; Cohn, Daniel H.; Krakow, Deborah; Mottes, Monica; Schwarze, Ulrike; Chen, Diana; Yang, Kathleen; Kuslich, Christine; Troendle, James; Dalgleish, Raymond; Byers, Peter H.
2014-01-01
Osteogenesis imperfecta (OI) is a generalized disorder of connective tissue characterized by fragile bones and easy susceptibility to fracture. Most cases of OI are caused by mutations in type I collagen. We have identified and assembled structural mutations in type I collagen genes (COL1A1 and COL1A2, encoding the proα1(I) and proα2(I) chains, respectively) that result in OI. Quantitative defects causing type I OI were not included. Of these 832 independent mutations, 682 result in substitution for glycine residues in the triple helical domain of the encoded protein and 150 alter splice sites. Distinct genotype–phenotype relationships emerge for each chain. One-third of the mutations that result in glycine substitutions in α1(I) are lethal, especially when the substituting residues are charged or have a branched side chain. Substitutions in the first 200 residues are nonlethal and have variable outcome thereafter, unrelated to folding or helix stability domains. Two exclusively lethal regions (helix positions 691–823 and 910–964) align with major ligand binding regions (MLBRs), suggesting crucial interactions of collagen monomers or fibrils with integrins, matrix metalloproteinases (MMPs), fibronectin, and cartilage oligomeric matrix protein (COMP). Mutations in COL1A2 are predominantly nonlethal (80%). Lethal substitutions are located in eight regularly spaced clusters along the chain, supporting a regional model. The lethal regions align with proteoglycan binding sites along the fibril, suggesting a role in fibril–matrix interactions. Recurrences at the same site in α2(I) are generally concordant for outcome, unlike α1(I). Splice site mutations comprise 20% of helical mutations identified in OI patients, and may lead to exon skipping, intron inclusion, or the activation of cryptic splice sites. Splice site mutations in COL1A1 are rarely lethal; they often lead to frameshifts and the mild type I phenotype. In α2(I), lethal exon skipping events are located in the carboxyl half of the chain. Our data on genotype–phenotype relationships indicate that the two collagen chains play very different roles in matrix integrity and that phenotype depends on intracellular and extracellular events. PMID:17078022
-
Remily-Wood, Elizabeth R.; Benson, Kaaron; Baz, Rachid C.; Chen, Y. Ann; Hussein, Mohamad; Hartley-Brown, Monique A.; Sprung, Robert W.; Perez, Brianna; Liu, Richard Z.; Yoder, Sean; Teer, Jamie; Eschrich, Steven A.; Koomen, John M.
2014-01-01
Purpose Quantitative mass spectrometry assays for immunoglobulins (Igs) are compared with existing clinical methods in samples from patients with plasma cell dyscrasias, e.g. multiple myeloma. Experimental design Using LC-MS/MS data, Ig constant region peptides and transitions were selected for liquid chromatography-multiple reaction monitoring mass spectrometry (LC-MRM). Quantitative assays were used to assess Igs in serum from 83 patients. Results LC-MRM assays quantify serum levels of Igs and their isoforms (IgG1–4, IgA1–2, IgM, IgD, and IgE, as well as kappa(κ) and lambda(λ) light chains). LC-MRM quantification has been applied to single samples from a patient cohort and a longitudinal study of an IgE patient undergoing treatment, to enable comparison with existing clinical methods. Proof-of-concept data for defining and monitoring variable region peptides are provided using the H929 multiple myeloma cell line and two MM patients. Conclusions and Clinical Relevance LC-MRM assays targeting constant region peptides determine the type and isoform of the involved immunoglobulin and quantify its expression; the LC-MRM approach has improved sensitivity compared with the current clinical method, but slightly higher interassay variability. Detection of variable region peptides is a promising way to improve Ig quantification, which could produce a dramatic increase in sensitivity over existing methods, and could further complement current clinical techniques. PMID:24723328
-
Remily-Wood, Elizabeth R; Benson, Kaaron; Baz, Rachid C; Chen, Y Ann; Hussein, Mohamad; Hartley-Brown, Monique A; Sprung, Robert W; Perez, Brianna; Liu, Richard Z; Yoder, Sean J; Teer, Jamie K; Eschrich, Steven A; Koomen, John M
2014-10-01
Quantitative MS assays for Igs are compared with existing clinical methods in samples from patients with plasma cell dyscrasias, for example, multiple myeloma (MM). Using LC-MS/MS data, Ig constant region peptides, and transitions were selected for LC-MRM MS. Quantitative assays were used to assess Igs in serum from 83 patients. RNA sequencing and peptide-based LC-MRM are used to define peptides for quantification of the disease-specific Ig. LC-MRM assays quantify serum levels of Igs and their isoforms (IgG1-4, IgA1-2, IgM, IgD, and IgE, as well as kappa (κ) and lambda (λ) light chains). LC-MRM quantification has been applied to single samples from a patient cohort and a longitudinal study of an IgE patient undergoing treatment, to enable comparison with existing clinical methods. Proof-of-concept data for defining and monitoring variable region peptides are provided using the H929 MM cell line and two MM patients. LC-MRM assays targeting constant region peptides determine the type and isoform of the involved Ig and quantify its expression; the LC-MRM approach has improved sensitivity compared with the current clinical method, but slightly higher inter-assay variability. Detection of variable region peptides is a promising way to improve Ig quantification, which could produce a dramatic increase in sensitivity over existing methods, and could further complement current clinical techniques. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
-
DeKosky, Brandon J.; Lungu, Oana I.; Park, Daechan; Johnson, Erik L.; Charab, Wissam; Chrysostomou, Constantine; Kuroda, Daisuke; Ellington, Andrew D.; Ippolito, Gregory C.; Gray, Jeffrey J.; Georgiou, George
2016-01-01
Elucidating how antigen exposure and selection shape the human antibody repertoire is fundamental to our understanding of B-cell immunity. We sequenced the paired heavy- and light-chain variable regions (VH and VL, respectively) from large populations of single B cells combined with computational modeling of antibody structures to evaluate sequence and structural features of human antibody repertoires at unprecedented depth. Analysis of a dataset comprising 55,000 antibody clusters from CD19+CD20+CD27− IgM-naive B cells, >120,000 antibody clusters from CD19+CD20+CD27+ antigen–experienced B cells, and >2,000 RosettaAntibody-predicted structural models across three healthy donors led to a number of key findings: (i) VH and VL gene sequences pair in a combinatorial fashion without detectable pairing restrictions at the population level; (ii) certain VH:VL gene pairs were significantly enriched or depleted in the antigen-experienced repertoire relative to the naive repertoire; (iii) antigen selection increased antibody paratope net charge and solvent-accessible surface area; and (iv) public heavy-chain third complementarity-determining region (CDR-H3) antibodies in the antigen-experienced repertoire showed signs of convergent paired light-chain genetic signatures, including shared light-chain third complementarity-determining region (CDR-L3) amino acid sequences and/or Vκ,λ–Jκ,λ genes. The data reported here address several longstanding questions regarding antibody repertoire selection and development and provide a benchmark for future repertoire-scale analyses of antibody responses to vaccination and disease. PMID:27114511
-
Crystal structure of a 3B3 variant - A broadly neutralizing HIV-1 scFv antibody
DOE Office of Scientific and Technical Information (OSTI.GOV)
Clark, K. Reed; Walsh, Scott T.R.; NCH)
2009-12-10
We present the crystal structure determination of an anti-HIV-1 gp120 single-chain variable fragment antibody variant, 3B3, at 2.5 {angstrom} resolution. This 3B3 variant was derived from the b12 antibody, using phage display and site-directed mutagenesis of the variable heavy chain (V{sub H}) complementary-determining regions (CDRs). 3B3 exhibits enhanced binding affinity and neutralization activity against several cross-clade primary isolates of HIV-1 by interaction with the recessed CD4-binding site on the gp120 envelope protein. Comparison with the structures of the unbound and bound forms of b12, the 3B3 structure closely resembles these structures with minimal differences with two notable exceptions. First, theremore » is a reorientation of the CDR-H3 of the V{sub H} domain where the primary sequences evolved from b12 to 3B3. The structural changes in CDR-H3 of 3B3, in light of the b12-gp120 complex structure, allow for positioning an additional Trp side chain in the binding interface with gp120. Finally, the second region of structural change involves two peptide bond flips in CDR-L3 of the variable light (VL) domain triggered by a point mutation in CDR-H3 of Q100eY resulting in changes in the intramolecular hydrogen bonding patterning between the VL and VH domains. Thus, the enhanced binding affinities and neutralization capabilities of 3B3 relative to b12 probably result from higher hydrophobic driving potential by burying more aromatic residues at the 3B3-gp120 interface and by indirect stabilization of intramolecular contacts of the core framework residues between the VL and VH domains possibly through more favorable entropic effect through the expulsion of water.« less
-
Mutation affecting the expression of immunoglobulin variable regions in the rabbit.
Kelus, A S; Weiss, S
1986-07-01
We have found a variant of the allotype allele a2 in the rabbit, which presumably arose by mutation, that segregates as expected for an allele at the a locus. This allele is called "ali" and the corresponding rabbit strain is called "Alicia." In heterozygous animals (ali/a1 and ali/a3) the concentration of a2 molecules is lower by a factor of 1000 than in standard a2/a2 homozygotes. In homozygous ali/ali individuals the a2 concentration varies with age--i.e., very low in young rabbits and higher in older ones--but it never reaches normal levels. The low level of a2 is compensated by increased amounts of a-negative molecules. Southern blot analysis did not reveal any gross changes in the intron between JH and C mu (joining region of immunoglobulin heavy chain and constant region of immunoglobulin mu chain) or in the number of VH gene segments encoding a locus specificities. We suggest that the ali phenotype is due to a mutation in a control element.
-
Mutation affecting the expression of immunoglobulin variable regions in the rabbit.
Kelus, A S; Weiss, S
1986-01-01
We have found a variant of the allotype allele a2 in the rabbit, which presumably arose by mutation, that segregates as expected for an allele at the a locus. This allele is called "ali" and the corresponding rabbit strain is called "Alicia." In heterozygous animals (ali/a1 and ali/a3) the concentration of a2 molecules is lower by a factor of 1000 than in standard a2/a2 homozygotes. In homozygous ali/ali individuals the a2 concentration varies with age--i.e., very low in young rabbits and higher in older ones--but it never reaches normal levels. The low level of a2 is compensated by increased amounts of a-negative molecules. Southern blot analysis did not reveal any gross changes in the intron between JH and C mu (joining region of immunoglobulin heavy chain and constant region of immunoglobulin mu chain) or in the number of VH gene segments encoding a locus specificities. We suggest that the ali phenotype is due to a mutation in a control element. Images PMID:3014517
-
Breaux, Breanna; Deiss, Thaddeus C.; Chen, Patricia L.; Cruz-Schneider, Maria Paula; Sena, Leonardo; Hunter, Margaret E.; Bonde, Robert K.; Criscitiello, Michael F.
2017-01-01
Manatees are a vulnerable, charismatic sentinel species from the evolutionarily divergent Afrotheria. Manatee health and resistance to infectious disease is of great concern to conservation groups, but little is known about their immune system. To develop manatee-specific tools for monitoring health, we first must have a general knowledge of how the immunoglobulin heavy (IgH) chain locus is organized and transcriptionally expressed. Using the genomic scaffolds of the Florida manatee (Trichechus manatus latirostris), we characterized the potential IgH segmental diversity and constant region isotypic diversity and performed the first Afrotherian repertoire analysis. The Florida manatee has low V(D)J combinatorial diversity (3744 potential combinations) and few constant region isotypes. They also lack clan III V segments, which may have caused reduced VH segment numbers. However, we found productive somatic hypermutation concentrated in the complementarity determining regions. In conclusion, manatees have limited IGHV clan and combinatorial diversity. This suggests that clan III V segments are essential for maintaining IgH locus diversity.
-
Linkage mapping of a mouse gene, iv, that controls left-right asymmetry of the heart and viscera.
Brueckner, M; D'Eustachio, P; Horwich, A L
1989-01-01
Inherited single gene defects have been identified in both humans and mice that lead to loss of developmental control over the left-right asymmetry of the heart and viscera. In mice the recessively inherited mutation iv leads to such apparent loss of control over situs: 50% of iv/iv mice exhibit situs inversus and 50% exhibit normal situs. The affected gene product has not been identified in these animals. To study the normal function of iv, we have taken an approach directed to the gene itself. As a first step, we have mapped iv genetically, by examining its segregation in backcrosses with respect to markers defined by restriction fragment length polymorphisms. The iv locus lies 3 centimorgans (cM) from the immunoglobulin heavy-chain constant-region gene complex (Igh-C) on chromosome 12. A multilocus map of the region suggests the gene order centromere-Aat (alpha 1-antitrypsin gene complex)-(11 cM)-iv-(3 cM)-Igh-C-(1 cM)-Igh-V (immunoglobulin heavy-chain variable-region gene complex). Images PMID:2740340
-
The complementarity-determining region sequences in IgY antivenom hypervariable regions.
da Rocha, David Gitirana; Fernandez, Jorge Hernandez; de Almeida, Claudia Maria Costa; da Silva, Claudia Letícia; Magnoli, Fabio Carlos; da Silva, Osmair Élder; da Silva, Wilmar Dias
2017-08-01
The data presented in this article are related to the research article entitled "Development of IgY antibodies against anti-snake toxins endowed with highly lethal neutralizing activity" (da Rocha et al., 2017) [1]. Complementarity-determining region (CDR) sequences are variable antibody (Ab) sequences that respond with specificity, duration and strength to identify and bind to antigen (Ag) epitopes. B lymphocytes isolated from hens immunized with Bitis arietans (Ba) and anti- Crotalus durissus terrificus (Cdt) venoms and expressing high specificity, affinity and toxicity neutralizing antibody titers were used as DNA sources. The VLF1, CDR1, CDR2, VLR1 and CDR3 sequences were validated by BLASTp, and values corresponding to IgY V L and V H anti-Ba or anti-Cdt venoms were identified, registered [ Gallus gallus IgY Fv Light chain (GU815099)/ Gallus gallus IgY Fv Heavy chain (GU815098)] and used for molecular modeling of IgY scFv anti-Ba. The resulting CDR1, CDR2 and CDR3 sequences were combined to construct the three - dimensional structure of the Ab paratope.
-
Breaux, Breanna; Deiss, Thaddeus C; Chen, Patricia L; Cruz-Schneider, Maria Paula; Sena, Leonardo; Hunter, Margaret E; Bonde, Robert K; Criscitiello, Michael F
2017-07-01
Manatees are a vulnerable, charismatic sentinel species from the evolutionarily divergent Afrotheria. Manatee health and resistance to infectious disease is of great concern to conservation groups, but little is known about their immune system. To develop manatee-specific tools for monitoring health, we first must have a general knowledge of how the immunoglobulin heavy (IgH) chain locus is organized and transcriptionally expressed. Using the genomic scaffolds of the Florida manatee (Trichechus manatus latirostris), we characterized the potential IgH segmental diversity and constant region isotypic diversity and performed the first Afrotherian repertoire analysis. The Florida manatee has low V(D)J combinatorial diversity (3744 potential combinations) and few constant region isotypes. They also lack clan III V segments, which may have caused reduced VH segment numbers. However, we found productive somatic hypermutation concentrated in the complementarity determining regions. In conclusion, manatees have limited IGHV clan and combinatorial diversity. This suggests that clan III V segments are essential for maintaining IgH locus diversity. Copyright © 2017 Elsevier Ltd. All rights reserved.
-
Tidal and residual currents across the northern Ryukyu Island chain observed by ferryboat ADCP
NASA Astrophysics Data System (ADS)
Liu, Zhao-Jun; Nakamura, Hirohiko; Zhu, Xiao-Hua; Nishina, Ayako; Dong, Menghong
2017-09-01
Ferryboat Acoustic Doppler Current Profiler (ADCP) data from 2003 to 2012 are used to estimate the tidal and residual currents across the northern Ryukyu Island chain (RIC) between the islands of Okinawa and Amamioshima. In this region, the M2 tide current is the strongest tidal component, and the K1 tide current is the strongest diurnal tidal component. The corresponding maximum amplitudes are 40 and 34 cm s-1, respectively. After removal of the tidal currents, the mean volume transport, 1.5 ± 2.7 Sv, flows into the East China Sea (ECS) from the western North Pacific through four channels in this area. In an empirical orthogonal function (EOF) analysis performed to clarify the temporal and spatial variability of currents through the four channels, the first two EOF modes account for 71% and 18% of the total variance, respectively. The EOF1 mode shows a clear bottom-intensified mode through the deep channel, which is likely to be formed by the propagation of bottom-trapped long topographic Rossby wave caused by the impingement of westward-propagating mesoscale eddies upon the eastern slope of the northern RIC. The EOF2 mode has significant seasonal variability and may be driven by the wind stress prevailing over the Kuroshio flow region around the northern RIC in October-November. This study provides observational evidence of the water exchanges across the northern RIC, which is essential for constructing a circulation scheme in the North Pacific subtropical western boundary region.
-
Ghiotto, Fabio; Marcatili, Paolo; Tenca, Claudya; Calevo, Maria Grazia; Yan, Xiao-Jie; Albesiano, Emilia; Bagnara, Davide; Colombo, Monica; Cutrona, Giovanna; Chu, Charles C; Morabito, Fortunato; Bruno, Silvia; Ferrarini, Manlio; Tramontano, Anna; Fais, Franco; Chiorazzi, Nicholas
2011-01-01
B-cell chronic lymphocytic leukemia (CLL) patients display leukemic clones bearing either germline or somatically mutated immunoglobulin heavy variable (IGHV ) genes. Most information on CLL immunoglobulins (Igs), such as the definition of stereotyped B-cell receptors (BCRs), was derived from germline unmutated Igs. In particular, detailed studies on the distribution and nature of mutations in paired heavy- and light-chain domains of CLL clones bearing mutated Igs are lacking. To address the somatic hyper-mutation dynamics of CLL Igs, we analyzed the mutation pattern of paired IGHV–diversity-joining (IGHV-D-J ) and immunoglobulin kappa/lambda variable-joining (IGK/LV-J ) rearrangements of 193 leukemic clones that displayed ≥2% mutations in at least one of the two immunoglobulin variable (IGV ) genes (IGHV and/or IGK/LV ). The relationship between the mutation frequency in IGHV and IGK/LV complementarity determining regions (CDRs) and framework regions (FRs) was evaluated by correlation analysis. Replacement (R) mutation frequency within IGK/LV chain CDRs correlated significantly with mutation frequency of paired IGHV CDRs in λ but not κ isotype CLL clones. CDRs of IGKV-J rearrangements displayed a lower percentage of R mutations than IGHVs. The frequency/pattern of mutations in kappa CLL Igs differed also from that in κ-expressing normal B cells described in the literature. Instead, the mutation frequency within the FRs of IGHV and either IGKV or IGLV was correlated. Notably, the amount of diversity introduced by replaced amino acids was comparable between IGHVs and IGKVs. The data indicate a different mutation pattern between κ and λ isotype CLL clones and suggest an antigenic selection that, in κ samples, operates against CDR variation. PMID:21785810
-
The Ultrasensitivity of Living Polymers
NASA Astrophysics Data System (ADS)
O'Shaughnessy, Ben; Vavylonis, Dimitrios
2003-03-01
Synthetic and biological living polymers are self-assembling chains whose chain length distributions (CLDs) are dynamic. We show these dynamics are ultrasensitive: Even a small perturbation (e.g., temperature jump) nonlinearly distorts the CLD, eliminating or massively augmenting short chains. The origin is fast relaxation of mass variables (mean chain length, monomer concentration) which perturbs CLD shape variables before these can relax via slow chain growth rate fluctuations. Viscosity relaxation predictions agree with experiments on the best-studied synthetic system, α-methylstyrene.
-
Kahr, Walter H A; Savoia, Anna; Pluthero, Fred G; Li, Ling; Christensen, Hilary; De Rocco, Daniela; Traivaree, Chanchai; Butchart, Sheila E; Curtin, Julie; Stollar, Elliott J; Forman-Kay, Julie D; Blanchette, Victor S
2009-12-01
Heterozygous mutations in MYH9, which encodes non-muscle myosin heavy chain IIA (MHC-IIA), result in autosomal dominant inherited MYH9-related disorders characterised by macro-thrombocytopenia, granulocyte inclusions, variable sensorineural deafness, cataracts and nephritis. MHC-IIA is assembled into a complex consisting of two pairs of light chains and two heavy chains, where the latter contain a neck region, SH3-like, motor and rod domains. We describe a patient with a Trp33Cys missense mutation in the SH3-like domain of MHC-IIA. Abnormal platelet function was observed using platelet aggregometry with the agonists epinephrine and adenosine diphosphate (ADP). Patient granulocytes and megakaryocytes, but not platelets, contained abnormal MHC-IIA inclusions visualised by confocal immunofluorescence or electron microscopy. Megakaryocytes grown in culture were smaller and contained hypolobulated nuclei compared to controls. Bone marrow-derived megakaryocytes revealed a preponderance of immature forms, the presence of structurally diverse inclusion bodies, and frequent emperipolesis as assessed by electron microscopy. Platelets and leukocytes contained indistinguishable amounts of total MHC-IIA determined by immunoblotting. Molecular modelling studies indicated that mutation of Trp33 destabilises the interface between the SH3-like and motor domain of MHC-IIA, which is close to previously described motor domain mutations, implying an important structural and/or functional role for this region in MHC-IIA.
-
Attallah, Carolina; Aguilar, María Fernanda; Garay, A Sergio; Herrera, Fernando E; Etcheverrigaray, Marina; Oggero, Marcos; Rodrigues, Daniel E
2017-10-01
The Cys residues are almost perfectly conserved in all antibodies. They contribute significantly to the antibody fragment stability. The relevance of two natural contiguous Cys residues of an anti-recombinant human-follicle stimulation hormone (rhFSH) in a format of single-chain variable fragment (scFv) was studied. This scFv contains 5 Cys residues: V H 22 and V H 92 in the variable heavy chain (V H ) and V L 23, V L 87 and V L 88 in the variable light chain (V L ). The influence of two unusual contiguous Cys at positions V L 87 and V L 88 was studied by considering the wild type fragment and mutant variants: V L -C88S, V L -C87S, V L -C87Y. The analysis was carried out using antigen-binding ability measurement by indirect specific ELISA and a detailed molecular modeling that comprises homology methods, long molecular dynamics simulations and docking. We found that V L -C87 affected the antibody fragment stability without interfering with the disulfide bond formation. The effect of mutating the V L -C87 by a usual residue at this position like Tyr caused distant structural changes at the V H region that confers a higher mobility to the V H -CDR2 and V H -CDR3 loops improving the scFv binding to the antigen. Copyright © 2017 Elsevier Ltd. All rights reserved.
-
NASA Astrophysics Data System (ADS)
Chocholáč, Jan; Průša, Petr
2016-12-01
The bullwhip effect generally refers to the phenomenon where order variability increases as the orders move upstream in the supply chain. It is serious problem for every member of the supply chain. This effect begins at customers and passes through the chain to producers, which are at the end of the logistic chain. Especially food supply chains are affected by this issue. These chains are unique for problems of expiration of goods (particularly perishable goods), variable demand, orders with quantity discounts and effort to maximize the customer satisfaction. This paper will present the problem of the bullwhip effect in the real supply chain in the food industry. This supply chain consists of approximately 350 stores, four central warehouses and more than 1000 suppliers, but the case study will examine 87 stores, one central warehouse and one supplier in 2015. The aim of this paper is the analysis of the order variability between the various links in this chain and confirmation of the bullwhip effect in this chain. The subject of the analysis will be perishable goods.
-
Yang, Rong; Jain, Tushar; Lynaugh, Heather; Nobrega, R Paul; Lu, Xiaojun; Boland, Todd; Burnina, Irina; Sun, Tingwan; Caffry, Isabelle; Brown, Michael; Zhi, Xiaoyong; Lilov, Asparouh; Xu, Yingda
Susceptibility of methionine to oxidation is an important concern for chemical stability during the development of a monoclonal antibody (mAb) therapeutic. To minimize downstream risks, leading candidates are usually screened under forced oxidation conditions to identify oxidation-labile molecules. Here we report results of forced oxidation on a large set of in-house expressed and purified mAbs with variable region sequences corresponding to 121 clinical stage mAbs. These mAb samples were treated with 0.1% H 2 O 2 for 24 hours before enzymatic cleavage below the hinge, followed by reduction of inter-chain disulfide bonds for the detection of the light chain, Fab portion of heavy chain (Fd) and Fc by liquid chromatography-mass spectrometry. This high-throughput, middle-down approach allows detection of oxidation site(s) at the resolution of 3 distinct segments. The experimental oxidation data correlates well with theoretical predictions based on the solvent-accessible surface area of the methionine side-chains within these segments. These results validate the use of upstream computational modeling to predict mAb oxidation susceptibility at the sequence level.
-
Li, Jiang-Wei; Xia, Lijie; Su, Youhong; Liu, Hongchun; Xia, Xueqing; Lu, Qinxia; Yang, Chunjin; Reheman, Kalbinur
2012-04-20
Screening of inhibitory Ab1 antibodies is a critical step for producing catalytic antibodies in the anti-idiotypic approach. However, the incompatible surface of the active site of the enzyme and the antigen-binding site of heterotetrameric conventional antibodies become the limiting step. Because camelid-derived nanobodies possess the potential to preferentially bind to the active site of enzymes due to their small size and long CDR3, we have developed a novel approach to produce antibodies with alliinase activities by exploiting the molecular mimicry of camel nanobodies. By screening the camelid-derived variable region of the heavy chain cDNA phage display library with alliinase, we obtained an inhibitory nanobody VHHA4 that recognizes the active site. Further screening with VHHA4 from the same variable domain of the heavy chain of a heavy-chain antibody library led to a higher incidence of anti-idiotypic Ab2 abzymes with alliinase activities. One of the abzymes, VHHC10, showed the highest activity that can be inhibited by Ab1 VHHA4 and alliinase competitive inhibitor penicillamine and significantly suppressed the B16 tumor cell growth in the presence of alliin in vitro. The results highlight the feasibility of producing abzymes via anti-idiotypic nanobody approach.
-
NASA Astrophysics Data System (ADS)
Vidal, V.; Adam, C.; Escartin, J.
2007-12-01
Walvis and St.~Helena are the only long-lived hotspot chains in the South Atlantic. Therefore, their characterization is important to constrain the processes associated with mantle plume formation, their temporal evolution, and the interaction with plate and mantle dynamics in the region. We study the temporal evolution of plume buoyancy and magma production rate along both hotspot chains, which are constrained from the swell and volume of volcanic materials emplaced along the chain. The regional depth anomaly is calculated by correcting the 2' bathymetry grid of Smith & Sandwell (1997) for thermal subsidence and sediment loading. We separate the topography associated with volcanism and the swell surrounding the hotspot chains using the MiFil filtering method (Adam et al., 2005). We then estimate the temporal variations associated with both parameters by computing volumes along the hotspot tracks. Neither Walvis nor St.~Helena show a 'classical' hotspot behavior. We find that two plumes are at the origin of the St.~Helena chain. This study also shows a swell associated with the Circe seamount, supporting the existence of a hotspot NW of the St.~Helena trail. The variation in swell and volcanic fluxes suggests temporal variability in the plume behavior at time scales of 10-20~m.y. and 5~m.y., which may be related to oscillations and instabilities of the plume conduit, respectively. Cumulative fluxes in the area are largest for Walvis and weakest for Circe, and all are significantly lower than that reported for the Hawai'i hotspot.
-
Liu, Fang; Li, Li; Zhang, Wei; Wang, Qi
2013-04-01
This research was to construct the lentiviral expression vector for anti- p185(erbB2) mouse/human chimeric antibody and to determine the expression of the chimeric antibody gene in 293T cells transfected with this vector. The genes (vL and vH) coding light and heavy chain of variable regions of anti-p185(erbB2) mAb and the constant regions of human IgG1 (kappa and gamma1) were cloned with PCR method. The target genes were assembled by three-primers PCR method to obtain the chimeric light chain (L) and the chimeric heavy chain (H). Both chains inserted into the down stream and upper stream of IRES gene of the plasmid pVAX1/IRES respectively. We digested the plasmid pVAX1/ H-IRES-L with endoenzyme and subcloned H-IRES-L into the lentiviral vector pWPI. The enzyme digestion and sequence analysis showed that the lentiviral expression vector pWPI/H-IRES-L was constructed correctly. Then, it was transfected into 293T cells and after 48h, GFP protein expression in 293T cells were detected by fluorescent microscope and the chimeric antibody expression was detected by RT-PCR and direct ELISA. The results showed that after 293T cells were transfected with recombination plasmid, both light and heavy chains of the chimeric antibody genes could express together. The chimeric antibody expressed could bind to p185(erbB2) specifically. This research may lay a sound foundation for further study of anti-p185(erbB2) engineered antibody.
-
Stoops, Janelle; Byrd, Samantha; Hasegawa, Haruki
2012-10-01
Russell bodies are intracellular aggregates of immunoglobulins. Although the mechanism of Russell body biogenesis has been extensively studied by using truncated mutant heavy chains, the importance of the variable domain sequences in this process and in immunoglobulin biosynthesis remains largely unknown. Using a panel of structurally and functionally normal human immunoglobulin Gs, we show that individual immunoglobulin G clones possess distinctive Russell body inducing propensities that can surface differently under normal and abnormal cellular conditions. Russell body inducing predisposition unique to each immunoglobulin G clone was corroborated by the intrinsic physicochemical properties encoded in the heavy chain variable domain/light chain variable domain sequence combinations that define each immunoglobulin G clone. While the sequence based intrinsic factors predispose certain immunoglobulin G clones to be more prone to induce Russell bodies, extrinsic factors such as stressful cell culture conditions also play roles in unmasking Russell body propensity from immunoglobulin G clones that are normally refractory to developing Russell bodies. By taking advantage of heterologous expression systems, we dissected the roles of individual subunit chains in Russell body formation and examined the effect of non-cognate subunit chain pair co-expression on Russell body forming propensity. The results suggest that the properties embedded in the variable domain of individual light chain clones and their compatibility with the partnering heavy chain variable domain sequences underscore the efficiency of immunoglobulin G biosynthesis, the threshold for Russell body induction, and the level of immunoglobulin G secretion. We propose that an interplay between the unique properties encoded in variable domain sequences and the state of protein homeostasis determines whether an immunoglobulin G expressing cell will develop the Russell body phenotype in a dynamic cellular setting. Copyright © 2012 Elsevier B.V. All rights reserved.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Wilbur, Jeremy D., E-mail: jwilbur@msg.ucsf.edu; Hwang, Peter K.; Brodsky, Frances M.
2010-03-01
Variable packing interaction related to the conformational flexibility within the huntingtin-interacting protein 1 coiled coil domain. Huntingtin-interacting protein 1 (HIP1) is an important link between the actin cytoskeleton and clathrin-mediated endocytosis machinery. HIP1 has also been implicated in the pathogenesis of Huntington’s disease. The binding of HIP1 to actin is regulated through an interaction with clathrin light chain. Clathrin light chain binds to a flexible coiled-coil domain in HIP1 and induces a compact state that is refractory to actin binding. To understand the mechanism of this conformational regulation, a high-resolution crystal structure of a stable fragment from the HIP1 coiled-coilmore » domain was determined. The flexibility of the HIP1 coiled-coil region was evident from its variation from a previously determined structure of a similar region. A hydrogen-bond network and changes in coiled-coil monomer interaction suggest that the HIP1 coiled-coil domain is uniquely suited to allow conformational flexibility.« less
-
Jhunjhunwala, Suchit; van Zelm, Menno C.; Peak, Mandy M.; Cutchin, Steve; Riblet, Roy; van Dongen, Jacques J.M.; Grosveld, Frank G.; Knoch, Tobias A.; Murre, Cornelis
2009-01-01
SUMMARY The immunoglobulin heavy-chain (Igh) locus is organized into distinct regions that contain multiple variable (VH), diversity (DH), joining (JH) and constant (CH) coding elements. How the Igh locus is structured in 3D space is unknown. To probe the topography of the Igh locus, spatial distance distributions were determined between 12 genomic markers that span the entire Igh locus. Comparison of the distance distributions to computer simulations of alternative chromatin arrangements predicted that the Igh locus is organized into compartments containing clusters of loops separated by linkers. Trilateration and triple-point angle measurements indicated the mean relative 3D positions of the VH, DH, JH, and CH elements, showed compartmentalization and striking conformational changes involving VH and DH-JH elements during early B cell development. In pro-B cells, the entire repertoire of VH regions (2 Mbp) appeared to have merged and juxtaposed to the DH elements, mechanistically permitting long-range genomic interactions to occur with relatively high frequency. PMID:18423198
-
Bianchi, Sergio; Moreno, Pilar; Landoni, Ana Inés; Naya, Hugo; Oppezzo, Pablo; Dighiero, Guillermo; Gabús, Raúl; Pritsch, Otto
2010-11-01
B-cell chronic lymphocytic leukemia (CLL) is characterized by the accumulation of long-lived circulating clonal leukemic B-cells, although the etiopathogenesis remains unclear. The incidence of CLL is variable in different regions around the world. While it is the most frequent chronic leukemia in Western countries, it has a low incidence in Asia. In this work we have investigated the immunoglobulin heavy chain gene rearrangements and mutational status in 80 Uruguayan patients with CLL, and compared these results with those obtained in other geographic regions. Our results demonstrate that Uruguayan patients with CLL display an IGHV gene usage which resembles that observed in Mediterranean countries and exhibits certain differences compared with Brazilian and Asian series, as expected, considering the ethnic basis of the Uruguayan population. This suggests that genetic influences could be important in the development and etiopathogenesis of CLL, but larger studies are necessary to substantiate this possibility.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Mullinax, R.L.; Gross, E.A.; Amberg, J.R.
1990-10-01
The authors have applied a molecular biology approach to the identification of human monoclonal antibodies. Human peripheral blood lymphocyte mRNA was converted to cDNA and a select subset was amplified by the polymerase chain reaction. These products, containing coding sequences for numerous immunoglobulin heavy- and {kappa} light-chain variable and constant region domains, were inserted into modified bacteriophase {lambda} expression vectors and introduced into Escherichia coli by infection to yield a combinatorial immunoexpression library. Clones with binding activity to tetanus toxoid were identified by filter hybridization with radiolabeled antigen and appeared at a frequency of 0.2{percent} in the library. These humanmore » antigen binding fragments, consisting of a heavy-chain fragment covalently linked to a light chain, displayed high affinity of binding to tetanus toxoid with equilibrium constants in the nanomolar range but did not cross-react with other proteins tested. They estimate that this human immunoexpression library contains 20,000 clones with high affinity and specificity to our chosen antigen.« less
-
Using Kalman Filter Chemical Data Assimilation to Study Ozone Catalytic Loss Cycles in January 1992
NASA Technical Reports Server (NTRS)
Lary, David J.
2002-01-01
This paper presents for the first time a global study of the ozone catalytic destruction cycles operating in the stratosphere using a stratospheric analyses for January 1992. The chemical analyses were produced using a Kalman filter data assimilation system. Because a major component of the variability of trace gases is due to the atmospheric motions the analyses have been cast in a flow-tracking coordinate system that moves with the large scale flow pattern. Particular attention is paid to the kinetic aspects of these cycles such as the rate limiting step and chain length. Although it is an important kinetic parameter, the chain length of the various cycles is seldom considered when the various catalytic cycles are discussed. This survey highlights that in the low stratosphere the cycles involving HO2 and halogens (notably bromine) are particularly important. In approximate order of effectiveness the most important ozone loss cycles in the polar lower stratosphere are the BrO/ClO, HO2/BrO, and OH/HO2 cycles. The ClO/ClO cycle clearly delineates the regions of chlorine activation. The chain length of the HO2/ClO, OH/HO2, Br/BrO, and ClO/NO2, clearly delineate the vortex edge region. The chain length of the BrO/NO2 and Cl/NO2 cycles highlight the regions of chemical processing outside the vortex where streamers of chemically processed air are stripped-off and transported away from the vortex. This is also true in the very low stratosphere for the Cl/ClO and BrO/ClO cycles.
-
NASA Astrophysics Data System (ADS)
Pineda-Martinez, Luis F.; Carbajal, Noel
2009-08-01
A series of numerical experiments were carried out to study the effect of meteorological events such as warm and cold air masses on climatic features and variability of a understudied region with strong topographic gradients in the northeastern part of Mexico. We applied the mesoscale model MM5. We investigated the influence of soil moisture availability in the performance of the model under two representative events for winter and summer. The results showed that a better resolution in land use cover improved the agreement among observed and calculated data. The topography induces atmospheric circulation patterns that determine the spatial distribution of climate and seasonal behavior. The numerical experiments reveal regions favorable to forced convection on the eastern side of the mountain chains Eastern Sierra Madre and Sierra de Alvarez. These processes affect the vertical and horizontal structure of the meteorological variables along the topographic gradient.
-
Sweeney, Lisa M.; Parker, Ann; Haber, Lynne T.; Tran, C. Lang; Kuempel, Eileen D.
2015-01-01
A biomathematical model was previously developed to describe the long-term clearance and retention of particles in the lungs of coal miners. The model structure was evaluated and parameters were estimated in two data sets, one from the United States and one from the United Kingdom. The three-compartment model structure consists of deposition of inhaled particles in the alveolar region, competing processes of either clearance from the alveolar region or translocation to the lung interstitial region, and very slow, irreversible sequestration of interstitialized material in the lung-associated lymph nodes. Point estimates of model parameter values were estimated separately for the two data sets. In the current effort, Bayesian population analysis using Markov chain Monte Carlo simulation was used to recalibrate the model while improving assessments of parameter variability and uncertainty. When model parameters were calibrated simultaneously to the two data sets, agreement between the derived parameters for the two groups was very good, and the central tendency values were similar to those derived from the deterministic approach. These findings are relevant to the proposed update of the ICRP human respiratory tract model with revisions to the alveolar-interstitial region based on this long-term particle clearance and retention model. PMID:23454101
-
Spatially explicit modeling of blackbird abundance in the Prairie Pothole Region
Forcey, Greg M.; Thogmartin, Wayne E.; Linz, George M.; McKann, Patrick C.; Crimmins, Shawn M.
2015-01-01
Knowledge of factors influencing animal abundance is important to wildlife biologists developing management plans. This is especially true for economically important species such as blackbirds (Icteridae), which cause more than $100 million in crop damages annually in the United States. Using data from the North American Breeding Bird Survey, the National Land Cover Dataset, and the National Climatic Data Center, we modeled effects of regional environmental variables on relative abundance of 3 blackbird species (red-winged blackbird,Agelaius phoeniceus; yellow-headed blackbird, Xanthocephalus xanthocephalus; common grackle, Quiscalus quiscula) in the Prairie Pothole Region of the central United States. We evaluated landscape covariates at 3 logarithmically related spatial scales (1,000 ha, 10,000 ha, and 100,000 ha) and modeled weather variables at the 100,000-ha scale. We constructed models a priori using information from published habitat associations. We fit models with WinBUGS using Markov chain Monte Carlo techniques. Both landscape and weather variables contributed strongly to predicting blackbird relative abundance (95% credibility interval did not overlap 0). Variables with the strongest associations with blackbird relative abundance were the percentage of wetland area and precipitation amount from the year before bird surveys were conducted. The influence of spatial scale appeared small—models with the same variables expressed at different scales were often in the best model subset. This large-scale study elucidated regional effects of weather and landscape variables, suggesting that management strategies aimed at reducing damages caused by these species should consider the broader landscape, including weather effects, because such factors may outweigh the influence of localized conditions or site-specific management actions. The regional species distributional models we developed for blackbirds provide a tool for understanding these broader landscape effects and guiding wildlife management practices to areas that are optimally beneficial. Published 2015. This article is a U.S. Government work and is in the public domain in the USA.
-
Bagheri, Salman; Yousefi, Mehdi; Safaie Qamsari, Elmira; Riazi-Rad, Farhad; Abolhassani, Mohsen; Younesi, Vahid; Dorostkar, Ruhollah; Movassaghpour, Ali Akbar; Sharifzadeh, Zahra
2017-03-01
The 4-1BB is a surface glycoprotein that pertains to the tumor necrosis factor-receptor family. There is compelling evidence suggesting important roles for 4-1BB in the immune response, including cell activation and proliferation and also cytokine induction. Because of encouraging results of different agonistic monoclonal antibodies against 4-1BB in the treatment of cancer, infectious, and autoimmune diseases, 4-1BB has been suggested as an attractive target for immunotherapy. In this study, single chain variable fragment phage display libraries, Tomlinson I+J, were screened against specific synthetic oligopeptides (peptides I and II) designed from 4-1BB extracellular domain. Five rounds of panning led to selection of four 4-1BB specific single chain variable fragments (PI.12, PI.42, PII.16, and PII.29) which showed specific reaction to relevant peptides in phage enzyme-linked immunosorbent assay. The selected clones were successfully expressed in Escherichia coli Rosetta-gami 2, and their expression was confirmed by western blot analysis. Enzyme-linked immunosorbent assay experiments indicated that these antibodies were able to specifically recognize 4-1BB without any cross-reactivity with other antigens. Flow cytometry analysis demonstrated an acceptable specific binding of the single chain variable fragments to 4-1BB expressed on CCRF-CEM cells, while no binding was observed with an irrelevant antibody. Anti-4-1BB single chain variable fragments enhanced surface CD69 expression and interleukin-2 production in stimulated CCRF-CEM cells which confirmed the agonistic effect of the selected single chain variable fragments. The data from this study have provided a rationale for further experiments involving the biological functions of anti-4-1BB single chain variable fragments in future studies.
-
Gladiolus plants transformed with single-chain variable fragment antibodies to Cucumber mosaic virus
USDA-ARS?s Scientific Manuscript database
Transgenic plants of Gladiolus ‘Peter Pears’ or ‘Jenny Lee’ were developed that contain single-chain variable fragments (scFv) to Cucumber mosaic virus (CMV) subgroup I or II. The CMV subgroup I heavy and light chain scFv fragments were placed under control of either the duplicated CaMV 35S or suga...
-
Falling Chains as Variable-Mass Systems: Theoretical Model and Experimental Analysis
ERIC Educational Resources Information Center
de Sousa, Celia A.; Gordo, Paulo M.; Costa, Pedro
2012-01-01
In this paper, we revisit, theoretically and experimentally, the fall of a folded U-chain and of a pile-chain. The model calculation implies the division of the whole system into two subsystems of variable mass, allowing us to explore the role of tensional contact forces at the boundary of the subsystems. This justifies, for instance, that the…
-
Interannual variability of ring formations in the Gulf Stream region
NASA Astrophysics Data System (ADS)
Sasaki, Y. N.
2016-02-01
An oceanic ring in the Gulf Stream (GS) region plays important roles in across-jet transport of heat, salt, momentum, and nutrients. This study examines interannual variability of rings shed from the GS jet and their properties using satellite altimeter observations from 1993 to 2013. An objective method is used to capture a ring shedding from the GS jet and track its movement. A spatial distribution of the ring formations in the GS region showed that both cyclonic (cold-core) and anticyclonic (warm-core) rings were most frequently formed around the New England Seamount chain between 62°-65°W, suggesting the importance of the bottom topography on the pinch-off process. These rings moved westward, although about two-third of these rings was reabsorbed by the GS jet. The number of ring formations, especially cyclonic ring formations, indicated prominent fluctuations on interannual to decadal timescales. The annual maximum number of the pinched-off rings is four times larger than the annual minimum number of the rings. These fluctuations of the ring formations were negatively correlated with the strength of the GS. This situation is similar that in the Kuroshio Extension region. The interannual variability of the number of ring formations is also negatively correlated with the North Atlantic Oscillation (NAO) index with one-year lag (NAO leads). Interannual variations of the propagation tendency and shape of rings are also discussed.
-
Neuberger, M S; Rajewsky, K
1981-01-01
From a hybrid mouse cell line (B1-8) that secreted an IgM, lambda 1 anti-(4-hydroxy-3-nitrophenyl)acetyl antibody but that had no detectable surface IgM, selection for a variant with lambda 1 chains on the surface resulted in the isolation of a line that had switched from mu to delta expression. The surface and secreted Igs of this line were typed as IgD with two monoclonal antibodies, and the parental IgM and variant IgD molecules carried the same variable regions as judged by hapten-binding and idiotypic analysis. The surface and secreted delta chains of the IgD variant have apparent molecular weights of 64,000 and 61,000, respectively. However, the unglycosylated secreted delta polypeptide chain has a molecular weight of only 44,000. The secreted IgD exists predominantly in the delta 2 lambda A2 form, does not contain J protein, is relatively stable in serum, and does not fix complement. Images PMID:6940132
-
Contextual determinants of US nursing home racial/ethnic diversity.
Davis, Jullet A; Weech-Maldonado, Robert; Lapane, Kate L; Laberge, Alex
2014-03-01
We hypothesized that for-profit/chain affiliated nursing homes, those in states with higher Medicaid reimbursement, and those in more competitive markets would have greater resident racial/ethnic diversity than nursing homes not meeting these criteria. Using 2004 Online Survey, Certification and Reporting data, Minimum Data Set, Lewis Mumford Center for Comparative Urban and Regional Research data, and the Area Resource File, we included U.S. Medicare/Medicaid certified nursing homes (N = 8950) located in 310 Metropolitan Statistical Areas. The dependent variable quantified facility-level multiracial diversity. Ordinary least squares regression showed support for the hypothesized relationships: for-profit/chain affiliated nursing homes were more diverse than nursing homes in all other ownership/chain member categories, while higher Medicaid per-diem rates, greater residential diversity, and stronger market competition were also positively associated with nursing home racial/ethnic composition. Results suggest there is room for policy changes to achieve equitable access to all levels of nursing home services for minority elders. Copyright © 2013 Elsevier Ltd. All rights reserved.
-
40 CFR 264.276 - Food-chain crops.
Code of Federal Regulations, 2012 CFR
2012-07-01
... 40 Protection of Environment 27 2012-07-01 2012-07-01 false Food-chain crops. 264.276 Section 264... Treatment § 264.276 Food-chain crops. The Regional Administrator may allow the growth of food-chain crops in... Regional Administrator will specify in the facility permit the specific food-chain crops which may be grown...
-
40 CFR 264.276 - Food-chain crops.
Code of Federal Regulations, 2011 CFR
2011-07-01
... 40 Protection of Environment 26 2011-07-01 2011-07-01 false Food-chain crops. 264.276 Section 264... Treatment § 264.276 Food-chain crops. The Regional Administrator may allow the growth of food-chain crops in... Regional Administrator will specify in the facility permit the specific food-chain crops which may be grown...
-
40 CFR 264.276 - Food-chain crops.
Code of Federal Regulations, 2010 CFR
2010-07-01
... 40 Protection of Environment 25 2010-07-01 2010-07-01 false Food-chain crops. 264.276 Section 264... Treatment § 264.276 Food-chain crops. The Regional Administrator may allow the growth of food-chain crops in... Regional Administrator will specify in the facility permit the specific food-chain crops which may be grown...
-
40 CFR 264.276 - Food-chain crops.
Code of Federal Regulations, 2013 CFR
2013-07-01
... 40 Protection of Environment 27 2013-07-01 2013-07-01 false Food-chain crops. 264.276 Section 264... Treatment § 264.276 Food-chain crops. The Regional Administrator may allow the growth of food-chain crops in... Regional Administrator will specify in the facility permit the specific food-chain crops which may be grown...
-
40 CFR 264.276 - Food-chain crops.
Code of Federal Regulations, 2014 CFR
2014-07-01
... 40 Protection of Environment 26 2014-07-01 2014-07-01 false Food-chain crops. 264.276 Section 264... Treatment § 264.276 Food-chain crops. The Regional Administrator may allow the growth of food-chain crops in... Regional Administrator will specify in the facility permit the specific food-chain crops which may be grown...
-
Effects of Variable Resistance Training on Maximal Strength: A Meta-Analysis.
Soria-Gila, Miguel A; Chirosa, Ignacio J; Bautista, Iker J; Baena, Salvador; Chirosa, Luis J
2015-11-01
Variable resistance training (VRT) methods improve the rate of force development, coordination between antagonist and synergist muscles, the recruitment of motor units, and reduce the drop in force produced in the sticking region. However, the beneficial effects of long-term VRT on maximal strength both in athletes and untrained individuals have been much disputed. The purpose of this study was to compare in a meta-analysis the effects of a long-term (≥7 weeks) VRT program using chains or elastic bands and a similar constant resistance program in both trained adults practicing different sports and untrained individuals. Intervention effect sizes were compared among investigations meeting our selection and inclusion criteria using a random-effects model. The published studies considered were those addressing VRT effects on the 1 repetition maximum. Seven studies involving 235 subjects fulfilled the selection and inclusion criteria. Variable resistance training led to a significantly greater mean strength gain (weighted mean difference: 5.03 kg; 95% confidence interval: 2.26-7.80 kg; Z = 3.55; p < 0.001) than the gain recorded in response to conventional weight training. Long-term VRT training using chains or elastic bands attached to the barbell emerged as an effective evidence-based method of improving maximal strength both in athletes with different sports backgrounds and untrained subjects.
-
Closed loop supply chain network design with fuzzy tactical decisions
NASA Astrophysics Data System (ADS)
Sherafati, Mahtab; Bashiri, Mahdi
2016-09-01
One of the most strategic and the most significant decisions in supply chain management is reconfiguration of the structure and design of the supply chain network. In this paper, a closed loop supply chain network design model is presented to select the best tactical and strategic decision levels simultaneously considering the appropriate transportation mode in activated links. The strategic decisions are made for a long term; thus, it is more satisfactory and more appropriate when the decision variables are considered uncertain and fuzzy, because it is more flexible and near to the real world. This paper is the first research which considers fuzzy decision variables in the supply chain network design model. Moreover, in this study a new fuzzy optimization approach is proposed to solve a supply chain network design problem with fuzzy tactical decision variables. Finally, the proposed approach and model are verified using several numerical examples. The comparison of the results with other existing approaches confirms efficiency of the proposed approach. Moreover the results confirms that by considering the vagueness of tactical decisions some properties of the supply chain network will be improved.
-
Measuring the influence of industry sector membership on supply chain disruption reporting.
Alcantara, Patrick
2015-01-01
The global Supply Chain Resilience Survey by the Business Continuity Institute and Zurich Insurance is a comprehensive study on the state of supply chains in different organisations worldwide. As a benchmarking tool, it also contains data about business continuity arrangements in place to ensure supply chain resilience. Given this study's historically qualitative approach to reporting, this paper aims to introduce quantitative analysis. In this paper, responses that report membership in Standard Industrial Classification 2007 industry sectors from the 2013 Supply Chain Resilience Survey were disaggregated and related to supply chain disruption reporting. A chi-square test of independence reveals that membership in a particular industry sector influences reporting of supply chain disruption. Nonetheless, the relationship between these variables is weak. This study demonstrates interesting differences between industry sectors in terms of supply chain resilience. Further research is required in terms of other variables in order to provide granularity and relevant findings to supply chain planners.
-
Dolimbek, Behzod Z; Steward, Lance E; Aoki, K Roger; Atassi, M Zouhair
2011-06-01
The continuous regions on botulinum neurotoxin A (BoNT/A) light (L) chain recognized by anti-toxin antibodies (Abs) from mouse, horse and chicken have been mapped. We synthesized a panel of thirty-two 19-residue peptides that overlapped consecutively by 5 residues and encompassed the entire L chain (residues 1-453). Mouse Abs recognized 5 major antigenic regions on the L chain, horse Abs recognized 9 while chicken Abs recognized 8 major antigenic regions. Overall, however, the three host species recognized, to some extent, similar, but not identical, peptides and the levels of Abs directed against a given region varied with the immunized host. Differences in the MHC of the host caused variation in levels of Ab recognition and some epitopes showed right or left frame-shifts among the species. Selected region(s) were also uniquely recognized by one species (e.g., peptide L1 by horse Abs). Mapping of the L chain antigenic regions and the previous localization of the regions on the H chain with the same antisera, has permitted description of the complete antigenic structure of BoNT/A. The locations in the 3-dimensional structure of the antigenic regions of the entire toxin are shown for mouse Abs. In the 3-D structure, the antigenic regions are on the surface of the toxin and when antibodies are bound the enzymatic activity of the light chain is obstructed. Copyright © 2010 Elsevier GmbH. All rights reserved.
-
Comparison of llama VH sequences from conventional and heavy chain antibodies.
Vu, K B; Ghahroudi, M A; Wyns, L; Muyldermans, S
1997-01-01
Forty different PCR clones encoding a llama variable heavy chain domain were analysed. The majority of these clones are derived from heavy-chain antibody cDNA in which the entire CH1 exon is absent. It appears from the amino acid within the VHH framework 1 and 3 that all the llama clones belong to the VH III family. However, the individual llama VHH sequences differ more substantially from each other than expected for members of the same family. Several remarkable amino acid substitutions in the framework 2 hinder the proper association of the VL. However, they lay the foundation for the secretion from the endoplasmic reticulum and good solubility behaviour of llama H2 antibodies. The repertoire of the llama VHHs may be extensive due to the presence of a long CDR3-loop, often constrained by a disulfide bridge and the occurrence of H1 and H2 loop conformations not yet encountered in mice or human VHs. The variability plot of the amino acids in the VHH shows that the first hypervariable region coincides with the structural H1 loop in contrast to the situation found in mice and man where the CDR1 and H1 are slightly offset. We propose that the amino acids of the llama H1 loop participate actively in the antigen binding. All these observations are characteristic for the llama VHHs of the homodimeric heavy-chain H2 antibodies, but are not maintained in the llama clones from conventional heterotetrameric H2L2 immunoglobulins.
-
Kozik, A V; Matvienko, M A; Men', A E; Zalenskiĭ, A O; Tikhonovich, I A
1992-01-01
We have determined the length of early noduline gene ENOD12 in various varieties and lines of pea (Pisum sativum) using the polymerase chain reaction (PCR). It was demonstrated that promoter regions of ENOD12A and ENOD12B genes in line 2150 (Afghanistan) are longer than these in variety "Feltham first". The disparity is 14 bp. When studying these genes in 7 different lines and varieties of pea it was found that ENOD12A gene is more variable in size than the ENOD12B gene. We showed the possibility to analyze the heritage of ENOD12 gene's alleles by using the PCR method.
-
Lee, Nam-Kyung; Bidlingmaier, Scott; Su, Yang; Liu, Bin
2018-01-01
Monoclonal antibodies and antibody-derived therapeutics have emerged as a rapidly growing class of biological drugs for the treatment of cancer, autoimmunity, infection, and neurological diseases. To support the development of human antibodies, various display techniques based on antibody gene repertoires have been constructed over the last two decades. In particular, scFv-antibody phage display has been extensively utilized to select lead antibodies against a variety of target antigens. To construct a scFv phage display that enables efficient antibody discovery, and optimization, it is desirable to develop a system that allows modular assembly of highly diverse variable heavy chain and light chain (Vκ and Vλ) repertoires. Here, we describe modular construction of large non-immune human antibody phage-display libraries built on variable gene cassettes from heavy chain and light chain repertoires (Vκ- and Vλ-light can be made into independent cassettes). We describe utility of such libraries in antibody discovery and optimization through chain shuffling.
-
Rumfelt, L L; Avila, D; Diaz, M; Bartl, S; McKinney, E C; Flajnik, M F
2001-02-13
In most vertebrate embryos and neonates studied to date unique antigen receptors (antibodies and T cell receptors) are expressed that possess a limited immune repertoire. We have isolated a subclass of IgM, IgM(1gj), from the nurse shark Ginglymostoma cirratum that is preferentially expressed in neonates. The variable (V) region gene encoding the heavy (H) chain underwent V-D-J rearrangement in germ cells ("germline-joined"). Such H chain V genes were discovered over 10 years ago in sharks but until now were not shown to be expressed at appreciable levels; we find expression of H(1gj) in primary and secondary lymphoid tissues early in life, but in adults only in primary lymphoid tissue, which is identified in this work as the epigonal organ. H(1gj) chain associates covalently with light (L) chains and is most similar in sequence to IgM H chains, but like mammalian IgG has three rather than the four IgM constant domains; deletion of the ancestral IgM C2 domain thus defines both IgG and IgM(1gj). Because sharks are the members of the oldest vertebrate class known to possess antibodies, unique or specialized antibodies expressed early in ontogeny in sharks and other vertebrates were likely present at the inception of the adaptive immune system.
-
Studies conducted as a search for physical mechanisms relating solar variability and the troposphere
NASA Technical Reports Server (NTRS)
Wu, S. T. (Editor)
1981-01-01
Chains of causative mechanisms that are hypothesized to relate solar variability to the behavior of the Earth's lower atmosphere were assessed. Solar variations believed most likely to constitute the forcing functions in hypothesized solar terrestrial atmosphere chain, changes in the Earth's atmospheric electrical characteristics due to solar variability, and the observed variations in atmospheric behavior that are influenced by solar variability were also examined.
-
Lucas, Alexander H.; Reason, Donald C.
1998-01-01
Anti-Haemophilus influenzae b polysaccharide (Hib PS) antibodies elicited in elderly subjects following conjugate vaccination expressed a light-chain variable-region (VL)-associated idiotype and had functional activities similar to those previously observed in children and younger adults. These findings indicate that advanced age is not accompanied by shifts in the major VL component of the Hib PS-specific repertoire or by diminution of the protective function of antibodies. PMID:9529108
-
Joint modelling rationale for chained equations
2014-01-01
Background Chained equations imputation is widely used in medical research. It uses a set of conditional models, so is more flexible than joint modelling imputation for the imputation of different types of variables (e.g. binary, ordinal or unordered categorical). However, chained equations imputation does not correspond to drawing from a joint distribution when the conditional models are incompatible. Concurrently with our work, other authors have shown the equivalence of the two imputation methods in finite samples. Methods Taking a different approach, we prove, in finite samples, sufficient conditions for chained equations and joint modelling to yield imputations from the same predictive distribution. Further, we apply this proof in four specific cases and conduct a simulation study which explores the consequences when the conditional models are compatible but the conditions otherwise are not satisfied. Results We provide an additional “non-informative margins” condition which, together with compatibility, is sufficient. We show that the non-informative margins condition is not satisfied, despite compatible conditional models, in a situation as simple as two continuous variables and one binary variable. Our simulation study demonstrates that as a consequence of this violation order effects can occur; that is, systematic differences depending upon the ordering of the variables in the chained equations algorithm. However, the order effects appear to be small, especially when associations between variables are weak. Conclusions Since chained equations is typically used in medical research for datasets with different types of variables, researchers must be aware that order effects are likely to be ubiquitous, but our results suggest they may be small enough to be negligible. PMID:24559129
-
Prosser, Christine E; Waters, Lorna C; Muskett, Frederick W; Veverka, Vaclav; Addis, Philip W; Griffin, Laura M; Baker, Terry S; Lawson, Alastair D G; Wernery, Ulrich; Kinne, Jorg; Henry, Alistair J; Taylor, Richard J; Carr, Mark D
2014-04-01
Heavy chain antibodies differ in structure to conventional antibodies lacking both the light chain and the first heavy chain constant domain (CH1). Characteristics of the antigen-binding variable heavy domain of the heavy chain antibody (VHH) including the smaller size, high solubility and stability make them an attractive alternative to more traditional antibody fragments for detailed NMR-based structural analysis. Here we report essentially complete backbone and side chain (15)N, (13)C and (1)H assignments for a free VHH. Analysis of the backbone chemical shift data obtained indicates that the VHH is comprised predominantly of β-sheets corresponding to nearly 60% of the protein backbone.
-
Lape, Michael; Paula, Stefan; Ball, William J
2010-06-01
Immunotherapy by cocaine-binding monoclonal antibodies (mAbs) has emerged as a promising strategy for the treatment of cocaine addiction. The human (gamma1 heavy chain)/murine (lambda light chain) chimeric mAb 2E2 has excellent affinity and specificity for cocaine and recent animal studies have demonstrated 2E2's ability in vivo to reduce cocaine levels in the brain as well as alter cocaine self-administration behavior in rats. In this study, we used mAb 2E2 amino acid sequence information to create a homology model for the 3-D structure of its Fv fragment. Subsequent computational docking studies revealed the intermolecular interactions potentially responsible for mAb 2E2's cocaine binding properties. The driving force of cocaine binding was identified as a combination of hydrophobic interactions and a single hydrogen bond between a light chain tyrosine residue and a carbonyl oxygen atom of cocaine. The model also allowed for an in silico evaluation of single/double residue mutations in the heavy and light chain variable regions that might further enhance mAb 2E2's cocaine binding properties. Copyright (c) 2010 Elsevier Masson SAS. All rights reserved.
-
Lape, Michael; Paula, Stefan; Ball, William J.
2010-01-01
Immunotherapy by cocaine-binding monoclonal antibodies (mAbs) has emerged as a promising strategy for the treatment of cocaine addiction. The human (γ1 heavy chain)/murine (λ light chain) chimeric mAb 2E2 has excellent affinity and specificity for cocaine and recent animal studies have demonstrated 2E2’s ability in vivo to reduce cocaine levels in the brain as well as alter cocaine self-administration behavior in rats. In this study, we used mAb 2E2 amino acid sequence information to create a homology model for the 3-D structure of its Fv fragment. Subsequent computational docking studies revealed the intermolecular interactions potentially responsible for mAb 2E2’s cocaine binding properties. The driving force of cocaine binding was identified as a combination of hydrophobic interactions and a single hydrogen bond between a light chain tyrosine residue and a carbonyl oxygen atom of cocaine. The model also allowed for an in silico evaluation of single/double residue mutations in the heavy and light chain variable regions that might further enhance mAb 2E2’s cocaine binding properties. PMID:20185210
-
Oxidative Post-Translational Modifications of an Amyloidogenic Immunoglobulin Light Chain Protein.
Lu, Yanyan; Jiang, Yan; Prokaeva, Tatiana; Connors, Lawreen H; Costello, Catherine E
2017-05-01
Immunoglobulin light chain amyloidosis (AL) is a plasma cell disorder characterized by overproduction and deposition of monoclonal immunoglobulin (Ig) light chains (LC) or variable region fragments as amyloid fibrils in various organs and tissues. Much clinical evidence indicates that patients with AL amyloidosis sustain cardiomyocyte impairment and suffer from oxidative stress. We seek to understand the underlying biochemical pathways whose disruption or amplification during sporadic or sustained disease states leads to harmful physiological consequences and to determine the detailed structures of intermediates and products that serve as signposts for the biochemical changes and represent potential biomarkers. In this study, matrix-assisted laser desorption/ionization mass spectrometry provided extensive evidence for oxidative post-translational modifications (PTMs) of an amyloidogenic Ig LC protein from a patient with AL amyloidosis. Some of the tyrosine residues were heavily mono- or di-chlorinated. In addition, a novel oxidative conversion to a nitrile moiety was observed for many of the terminal aminomethyl groups on lysine side chains. In vitro experiments using model peptides, in-solution oxidation, and click chemistry demonstrated that hypochlorous acid produced by the myeloperoxidase - hydrogen peroxide - chloride system could be responsible for these and other, more commonly observed modifications.
-
NASA Astrophysics Data System (ADS)
Aalbers, Emma E.; Lenderink, Geert; van Meijgaard, Erik; van den Hurk, Bart J. J. M.
2018-06-01
High-resolution climate information provided by e.g. regional climate models (RCMs) is valuable for exploring the changing weather under global warming, and assessing the local impact of climate change. While there is generally more confidence in the representativeness of simulated processes at higher resolutions, internal variability of the climate system—`noise', intrinsic to the chaotic nature of atmospheric and oceanic processes—is larger at smaller spatial scales as well, limiting the predictability of the climate signal. To quantify the internal variability and robustly estimate the climate signal, large initial-condition ensembles of climate simulations conducted with a single model provide essential information. We analyze a regional downscaling of a 16-member initial-condition ensemble over western Europe and the Alps at 0.11° resolution, similar to the highest resolution EURO-CORDEX simulations. We examine the strength of the forced climate response (signal) in mean and extreme daily precipitation with respect to noise due to internal variability, and find robust small-scale geographical features in the forced response, indicating regional differences in changes in the probability of events. However, individual ensemble members provide only limited information on the forced climate response, even for high levels of global warming. Although the results are based on a single RCM-GCM chain, we believe that they have general value in providing insight in the fraction of the uncertainty in high-resolution climate information that is irreducible, and can assist in the correct interpretation of fine-scale information in multi-model ensembles in terms of a forced response and noise due to internal variability.
-
NASA Astrophysics Data System (ADS)
Aalbers, Emma E.; Lenderink, Geert; van Meijgaard, Erik; van den Hurk, Bart J. J. M.
2017-09-01
High-resolution climate information provided by e.g. regional climate models (RCMs) is valuable for exploring the changing weather under global warming, and assessing the local impact of climate change. While there is generally more confidence in the representativeness of simulated processes at higher resolutions, internal variability of the climate system—`noise', intrinsic to the chaotic nature of atmospheric and oceanic processes—is larger at smaller spatial scales as well, limiting the predictability of the climate signal. To quantify the internal variability and robustly estimate the climate signal, large initial-condition ensembles of climate simulations conducted with a single model provide essential information. We analyze a regional downscaling of a 16-member initial-condition ensemble over western Europe and the Alps at 0.11° resolution, similar to the highest resolution EURO-CORDEX simulations. We examine the strength of the forced climate response (signal) in mean and extreme daily precipitation with respect to noise due to internal variability, and find robust small-scale geographical features in the forced response, indicating regional differences in changes in the probability of events. However, individual ensemble members provide only limited information on the forced climate response, even for high levels of global warming. Although the results are based on a single RCM-GCM chain, we believe that they have general value in providing insight in the fraction of the uncertainty in high-resolution climate information that is irreducible, and can assist in the correct interpretation of fine-scale information in multi-model ensembles in terms of a forced response and noise due to internal variability.
-
NASA Astrophysics Data System (ADS)
Brochier, Timothée; Auger, Pierre-Amaël; Pecquerie, Laure; Machu, Eric; Capet, Xavier; Thiaw, Modou; Mbaye, Baye Cheikh; Braham, Cheikh-Baye; Ettahiri, Omar; Charouki, Najib; Sène, Ousseynou Ndaw; Werner, Francisco; Brehmer, Patrice
2018-05-01
Small pelagic fish (SPF) species are heavily exploited in eastern boundary upwelling systems (EBUS) as their transformation products are increasingly used in the world's food chain. Management relies on regular monitoring, but there is a lack of robust theories for the emergence of the populations' traits and their evolution in highly variable environments. This work aims to address existing knowledge gaps by combining physical and biogeochemical modelling with an individual life-cycle based model applied to round sardinella (Sardinella aurita) off northwest Africa, a key species for regional food security. Our approach focused on the processes responsible for seasonal migrations, spatio-temporal size-structure, and interannual biomass fluctuations. Emergence of preferred habitat resulted from interactions between natal homing behavior and environmental variability that impacts early life stages. Exploration of the environment by the fishes was determined by swimming capabilities, mesoscale to regional habitat structure, and horizontal currents. Fish spatio-temporal abundance variability emerged from a complex combination of distinct life-history traits. An alongshore gradient in fish size distributions is reported and validated by in situ measurements. New insights into population structure are provided, within an area where the species is abundant year-round (Mauritania) and with latitudinal migrations of variable (300-1200 km) amplitude. Interannual biomass fluctuations were linked to modulations of fish recruitment over the Sahara Bank driven by variability in alongshore current intensity. The identified processes constitute an analytical framework that can be implemented in other EBUS and used to explore impacts of regional climate change on SPF.
-
Recovery and fine structure variability of RGII sub-domains in wine (Vitis vinifera Merlot)
Buffetto, F.; Ropartz, D.; Zhang, X. J.; Gilbert, H. J.; Guillon, F.; Ralet, M.-C.
2014-01-01
Background and Aims Rhamnogalacturonan II (RGII) is a structurally complex pectic sub-domain composed of more than 12 different sugars and 20 different linkages distributed in five side chains along a homogalacturonan backbone. Although RGII has long been described as highly conserved over plant evolution, recent studies have revealed variations in the structure of the polysaccharide. This study examines the fine structure variability of RGII in wine, focusing on the side chains A and B obtained after sequential mild acid hydrolysis. Specifically, this study aims to differentiate intrinsic structural variations in these RGII side chains from structural variations due to acid hydrolysis. Methods RGII from wine (Vitis vinifera Merlot) was sequentially hydrolysed with trifluoroacetic acid (TFA) and the hydrolysis products were separated by anion-exchange chromatography (AEC). AEC fractions or total hydrolysates were analysed by MALDI-TOF mass spectrometry. Key Results The optimal conditions to recover non-degraded side chain B, side chain A and RGII backbone were 0·1 m TFA at 40 °C for 16 h, 0·48 m TFA at 40 °C for 16 h (or 0·1 m TFA at 60 °C for 8 h) and 0·1 m TFA at 60 °C for 16 h, respectively. Side chain B was particularly prone to acid degradation. Side chain A and the RGII GalA backbone were partly degraded by 0·1 m TFA at 80 °C for 1–4 h. AEC allowed separation of side chain B, methyl-esterified side chain A and non-methyl-esterified side chain A. The structure of side chain A and the GalA backbone were highly variable. Conclusions Several modifications to the RGII structure of wine were identified. The observed dearabinosylation and deacetylation were primarily the consequence of acidic treatment, while variation in methyl-esterification, methyl-ether linkages and oxidation reflect natural diversity. The physiological significance of this variability, however, remains to be determined. PMID:24908680
-
Highly sensitive and unbiased approach for elucidating antibody repertoires
Lin, Sherry G.; Ba, Zhaoqing; Du, Zhou; Zhang, Yu; Hu, Jiazhi; Alt, Frederick W.
2016-01-01
Developing B lymphocytes undergo V(D)J recombination to assemble germ-line V, D, and J gene segments into exons that encode the antigen-binding variable region of Ig heavy (H) and light (L) chains. IgH and IgL chains associate to form the B-cell receptor (BCR), which, upon antigen binding, activates B cells to secrete BCR as an antibody. Each of the huge number of clonally independent B cells expresses a unique set of IgH and IgL variable regions. The ability of V(D)J recombination to generate vast primary B-cell repertoires results from a combinatorial assortment of large numbers of different V, D, and J segments, coupled with diversification of the junctions between them to generate the complementary determining region 3 (CDR3) for antigen contact. Approaches to evaluate in depth the content of primary antibody repertoires and, ultimately, to study how they are further molded by secondary mutation and affinity maturation processes are of great importance to the B-cell development, vaccine, and antibody fields. We now describe an unbiased, sensitive, and readily accessible assay, referred to as high-throughput genome-wide translocation sequencing-adapted repertoire sequencing (HTGTS-Rep-seq), to quantify antibody repertoires. HTGTS-Rep-seq quantitatively identifies the vast majority of IgH and IgL V(D)J exons, including their unique CDR3 sequences, from progenitor and mature mouse B lineage cells via the use of specific J primers. HTGTS-Rep-seq also accurately quantifies DJH intermediates and V(D)J exons in either productive or nonproductive configurations. HTGTS-Rep-seq should be useful for studies of human samples, including clonal B-cell expansions, and also for following antibody affinity maturation processes. PMID:27354528
-
Borek, Aleksandra; Sokolowska-Wedzina, Aleksandra; Chodaczek, Grzegorz; Otlewski, Jacek
2018-01-01
Fibroblast growth factor receptors (FGFRs) are promising targets for antibody-based cancer therapies, as their substantial overexpression has been found in various tumor cells. Aberrant activation of FGF receptor 2 (FGFR2) signaling through overexpression of FGFR2 and/or its ligands, mutations, or receptor amplification has been reported in multiple cancer types, including gastric, colorectal, endometrial, ovarian, breast and lung cancer. In this paper, we describe application of the phage display technology to produce a panel of high affinity single chain variable antibody fragments (scFvs) against the extracellular ligand-binding domain of FGFR2 (ECD_FGFR2). The binders were selected from the human single chain variable fragment scFv phage display libraries Tomlinson I + J and showed high specificity and binding affinity towards human FGFR2 with nanomolar KD values. To improve the affinity of the best binder selected, scFvF7, we reformatted it to a bivalent diabody format, or fused it with the Fc region (scFvF7-Fc). The scFvF7-Fc antibody construct presented the highest affinity for FGFR2, with a KD of 0.76 nM, and was selectively internalized into cancer cells overexpressing FGFR2, Snu-16 and NCI-H716. Finally, we prepared a conjugate of scFvF7-Fc with the cytotoxic drug monomethyl-auristatin E (MMAE) and evaluated its cytotoxicity. The conjugate delivered MMAE selectively to FGFR2-positive tumor cells. These results indicate that scFvF7-Fc-vcMMAE is a highly potent molecule for the treatment of cancers with FGFR2 overexpression.
-
Integration of environmental aspects in modelling and optimisation of water supply chains.
Koleva, Mariya N; Calderón, Andrés J; Zhang, Di; Styan, Craig A; Papageorgiou, Lazaros G
2018-04-26
Climate change becomes increasingly more relevant in the context of water systems planning. Tools are necessary to provide the most economic investment option considering the reliability of the infrastructure from technical and environmental perspectives. Accordingly, in this work, an optimisation approach, formulated as a spatially-explicit multi-period Mixed Integer Linear Programming (MILP) model, is proposed for the design of water supply chains at regional and national scales. The optimisation framework encompasses decisions such as installation of new purification plants, capacity expansion, and raw water trading schemes. The objective is to minimise the total cost incurring from capital and operating expenditures. Assessment of available resources for withdrawal is performed based on hydrological balances, governmental rules and sustainable limits. In the light of the increasing importance of reliability of water supply, a second objective, seeking to maximise the reliability of the supply chains, is introduced. The epsilon-constraint method is used as a solution procedure for the multi-objective formulation. Nash bargaining approach is applied to investigate the fair trade-offs between the two objectives and find the Pareto optimality. The models' capability is addressed through a case study based on Australia. The impact of variability in key input parameters is tackled through the implementation of a rigorous global sensitivity analysis (GSA). The findings suggest that variations in water demand can be more disruptive for the water supply chain than scenarios in which rainfalls are reduced. The frameworks can facilitate governmental multi-aspect decision making processes for the adequate and strategic investments of regional water supply infrastructure. Copyright © 2018. Published by Elsevier B.V.
-
Seal, S N; Hoet, R M; Raats, J M; Radic, M Z
2000-09-01
To examine anti-double-stranded DNA (anti-dsDNA) IgG autoantibodies from the bone marrow of individuals with systemic lupus erythematosus (SLE). A library of single-chain variable fragments (scFv) was constructed from SLE bone marrow complementary DNA of gamma, kappa, and lambda isotype by cloning into the pHENIX phagemid vector. The library was screened with dsDNA in solution, and 2 anti-DNA phage, DNA1 and DNA4, were isolated and their Ig V genes sequenced. Soluble scFv corresponding to DNA1 and DNA4, and their heavy (H)- and light (L)-chain recombinants, were prepared, purified, and analyzed for binding to DNA by enzyme-linked immunosorbent assay. DNA1 and DNA4 used different Ig H-chain (3-30 and 5-51, respectively) and L-chain (DPK15 and DPK22, respectively) V genes. The ratios of replacement mutations to silent mutations in DNA1 and DNA4 suggest that their V genes were selected for improved antigen binding in vivo. The recombinant between DNA4VH and DNA1VL showed the highest relative affinity for both single-stranded DNA and dsDNA. These 2 Ig subunits contained third complementarity-determining region arginines and had acquired the majority of replacement mutations. Anti-dsDNA IgG autoantibodies from the bone marrow of SLE patients exploit diverse V genes and cationic V-D-J and V-J junctions for DNA binding, and accumulate replacement mutations that enhance binding.
-
Production of novel recombinant single-domain antibodies against tandem repeat region of MUC1 mucin.
Rahbarizadeh, F; Rasaee, M J; Forouzandeh Moghadam, M; Allameh, A A; Sadroddiny, E
2004-06-01
Recently, the existence of "heavy-chain" antibody in Camelidae has been described. However, as yet there is no data on the binding of this type of antibody to peptides. In addition, there was not any report of production of single-domain antibodies in two-humped camels (Camelus bactrianus). In the present study, these questions are addressed. We showed the feasibility of immunizing old world camels, cloning the repertoire of the variable domain of their heavy-chain antibodies, panning and selection, leading to the successful identification of minimum-sized antigen binders. Antigen-specific fragments of the heavy-chain IgGs (V(HH)) are of great interest in biotechnology because they are very stable, highly soluble, and react specifically and with high affinity to the antigens. In this study, we immunized two camels (Camelus dromedarius and Camelus bactrianus) with homogenized cancerous tissues, synthetic peptide, and human milk fat globule membrane (HMFG), and generated two V(HH) libraries displayed on phage particles. Some single-domain antibody fragments have been isolated that specifically recognize the tandem repeat region of MUC1. The camels' single-domain V(HH) harbor the original, intact antigen binding site and reacted specifically and with high affinity to the tandem repeat region of MUC1. Indeed soluble, specific antigen binders and good affinities (in the range of 0.2 x 10(9) M(-1) to 0.6 x 10(9) M(-1)) were identified from these libraries. This is the first example of the isolation of camel anti-peptide V(HH) domains.
-
Yin, Ling-Ling; Ruan, Su-Hong; Tian, Yu; Zhao, Kai; Xu, Kai Lin
2015-10-01
To clone the variable region genes of human anti-IL1RAP (IL-1 receptor accessory protein) monoclonal antibodies (McAb) and to construct IL1RAP chimeric antigen receptors (CARs). The VH and VL DNA of IL1RAP single chain antibodies were amplified by RACE and overlap extension PCR from total RNA extracted from 3H6E10 and 10D8A7 hybridoma and ligated into specific IL1RAP single-chain variable fragments (scFv). CD8α transmembrane domain, CD137 intracellular domain, TCR ζ chain, human CD8α signal peptide and scFv-anti-IL1RAP were cloned into plasmid LV-lac. Recombinant lentiviruses were generated by co-transfection of recombinant plasmid LV-lac, pMD2. G, and psPAX2 helper vectors into 293FT packing cells. The VH and VL genes of 2 human anti-IL1RAP McAb were acquired. The 3H6E10 VH and VL genes consisted of 402 bp and 393 bp encoding 134 and 131 aminoacid residues, respectively; 10D8A7 VH and VL genes consisted of 423 bp and 381 bp encoding 141 and 127 amine acid residues, respectively. Recombinant expression vertors LV-3H6E10 scFv-ICD and LV-10D8A7 scFv-ICD (ICD: CD8α transmembrane domain-CD137 intracellular domain-TCR ζ chain) were constructed. The target fragments were demonstrated by sequencing analysis. Recombinant plasmids were transfected into 293FT cells and lentiviral particles were acquired. Human anti-IL1RAP recombinant receptors are constructed successfully and lay a good foundation for the construction of IL1RAP-CAR killer T cell vaccine.
-
Continuously Variable Transmission
NASA Technical Reports Server (NTRS)
Grana, D. C.
1985-01-01
Chain slides along two cones, in novel transmission concept. Transmission includes chain drive between two splined shafts. Chain sprockets follow surfaces of two cones. As one chain sprocket moves toward smaller diameter other chain sprocket moves toward larger diameter, thereby changing "gear" ratio. Movement initiated by tension applied to chain by planetary gear mechanism. Device positive, simple, and efficient over wide range of speed ratios.
-
Abe, Ryoji; Jeong, Hee-Jin; Arakawa, Dai; Dong, Jinhua; Ohashi, Hiroyuki; Kaigome, Rena; Saiki, Fujio; Yamane, Kyosuke; Takagi, Hiroaki; Ueda, Hiroshi
2014-04-11
Recently, we described a novel reagentless fluorescent biosensor strategy named Quenchbody, which functions via the antigen-dependent removal of the quenching effect on a fluorophore that is attached to a single-chain antibody variable region. To explore the practical utility of Quenchbodies, we prepared antibody Fab fragments that were fluorolabeled at either one or two of the N-terminal regions, using a cell-free translation-mediated position-specific protein labeling system. Unexpectedly, the Fab fragment labeled at the heavy chain N-terminal region demonstrated a deeper quenching and antigen-dependent release compared to that observed using scFv. Moreover, when the Fab was fluorolabeled at the two N-termini with either the same dye or with two different dyes, an improved response due to enhanced quenching via dye-dye interactions was observed. On the basis of this approach, several targets, including peptides, proteins, and haptens, as well as narcotics, were quantified with a higher response up to 50-fold. In addition, differentiation of osteosarcoma to osteoblasts was successfully imaged using a similarly fluorolabeled recombinant Fab protein prepared from E. coli. Due to its versatility, this "Ultra-Quenchbody" is expected to exhibit a range of applications from in vitro diagnostics to the live imaging of various targets in situ.
-
Rumfelt, Lynn L.; Avila, David; Diaz, Marilyn; Bartl, Simona; McKinney, E. Churchill; Flajnik, Martin F.
2001-01-01
In most vertebrate embryos and neonates studied to date unique antigen receptors (antibodies and T cell receptors) are expressed that possess a limited immune repertoire. We have isolated a subclass of IgM, IgM1gj, from the nurse shark Ginglymostoma cirratum that is preferentially expressed in neonates. The variable (V) region gene encoding the heavy (H) chain underwent V-D-J rearrangement in germ cells (“germline-joined”). Such H chain V genes were discovered over 10 years ago in sharks but until now were not shown to be expressed at appreciable levels; we find expression of H1gj in primary and secondary lymphoid tissues early in life, but in adults only in primary lymphoid tissue, which is identified in this work as the epigonal organ. H1gj chain associates covalently with light (L) chains and is most similar in sequence to IgM H chains, but like mammalian IgG has three rather than the four IgM constant domains; deletion of the ancestral IgM C2 domain thus defines both IgG and IgM1gj. Because sharks are the members of the oldest vertebrate class known to possess antibodies, unique or specialized antibodies expressed early in ontogeny in sharks and other vertebrates were likely present at the inception of the adaptive immune system. PMID:11172027
-
Landeira, José M.; Ferron, Bruno; Lunven, Michel; Morin, Pascal; Marié, Louis; Sourisseau, Marc
2014-01-01
Phytoplankton blooms are usually dominated by chain-forming diatom species that can alter food pathways from primary producers to predators by reducing the interactions between intermediate trophic levels. The food-web modifications are determined by the length of the chains; however, the estimation is biased because traditional sampling strategies damage the chains and, therefore, change the phytoplankton size structure. Sedimentological studies around oceanic fronts have shown high concentrations of giant diatom mats (>1 cm in length), suggesting that the size of diatom chains is underestimated in the pelagic realm. Here, we investigate the variability in size and abundance of phytoplankton chains at the Ushant tidal front (NW France) using the Video Fluorescence Analyzer (VFA), a novel and non-invasive system. CTD and Scanfish profiling characterized a strong temperature and chlorophyll front, separating mixed coastal waters from the oceanic-stratified domain. In order to elucidate spring-neap variations in the front, vertical microstructure profiler was used to estimate the turbulence and vertical nitrate flux. Key findings were: (1) the VFA system recorded large diatom chains up to 10.7 mm in length; (2) chains were mainly distributed in the frontal region, with maximum values above the pycnocline in coincidence with the maximum chlorophyll; (3) the diapycnal fluxes of nitrate enabled the maintenance of the bloom in the frontal area throughout the spring-neap tidal cycle; (4) from spring to neap tide the chains length was significantly reduced; (5) during neap tide, the less intense vertical diffusion of nutrients, as well as the lower turbulence around the chains, intensified nutrient-depleted conditions and, thus, very large chains became disadvantageous. To explain this pattern, we suggest that size plasticity is an important ecological trait driving phytoplankton species competition. Although this plasticity behavior is well known from experiments in the laboratory, it has never been reported from observations in the field. PMID:24587384
-
NASA Astrophysics Data System (ADS)
Hijuzaman, O.; Rahayu, A.; Kusnendi
2018-05-01
The purpose of this study was to determine and analyze the effect of direct and indirect implementation of Green Supply-Chain Management (GSCM) on the Performance Marketing and Competitiveness Mediation Paper Company in West Java province. Object This research is the paper company that existed in West Java Province, which amounts to 30 companies as well as a sample of research, while variables studied are Variable Green Supply- Chain Management, Company Competitiveness Variable and Variable of Paper Marketing Industry Performance in West Java Province. The method used SEM with Variance Based or Component Based with Software used is SmartPLS 3.0. On this research are identified that no effect of Competitiveness on Marketing Performance in Paper Companies in West Java Province.
-
Kühne, Annett; Kaiser, Rolf; Schirmer, Markus; Heider, Ulrike; Muhlke, Sabine; Niere, Wiebke; Overbeck, Tobias; Hohloch, Karin; Trümper, Lorenz; Sezer, Orhan; Brockmöller, Jürgen
2007-07-01
Melphalan is widely used in the treatment of multiple myeloma. Pharmacokinetics of this alkylating drug shows high inter-individual variability. As melphalan is a phenylalanine derivative, the pharmacokinetic variability may be determined by genetic polymorphisms in the L-type amino acid transporters LAT1 (SLC7A5) and LAT2 (SLC7A8). Pharmacokinetics were analysed in 64 patients after first administration of intravenous melphalan. Severity of side effects was documented according to WHO criteria. Genomic DNA was analysed for polymorphisms in LAT1 and LAT2 by sequencing of the entire coding region, intron-exon boundaries and 2 kb upstream promoter region. Selected polymorphisms in the common heavy chain of both transporters, the protein 4F2hc (SLC3A2), were analysed by single nucleotide primer extension. Melphalan pharmacokinetics was highly variable with up to 6.2-fold differences in total clearance. A total of 44 polymorphisms were identified in LAT1 and 21 polymorphisms in LAT2. From all variants, only five were in the coding region and only one heterozygous non-synonymous polymorphism (Ala94Thr) was found in LAT2. Numerous polymorphisms were found in the LAT1 and LAT2 5'-flanking regions but did not correlate with expression of the respective genes. No significant correlations could be observed between the polymorphisms in 4F2hc, LAT1, and LAT2 with melphalan pharmacokinetics or with melphalan side effects. The study confirmed that these transporter genes are highly conserved, particularly in the coding sequences. Genetic variation in 4F2hc, LAT1, and LAT2 does not appear to be a major cause of inter-individual variability in pharmacokinetics and of adverse reactions to melphalan.
-
Ghadge, Ghanashyam D; Pavlovic, John D; Koduvayur, Sujatha P; Kay, Brian K; Roos, Raymond P
2013-08-01
Approximately 10% of amyotrophic lateral sclerosis (ALS) cases are familial (known as FALS) with an autosomal dominant inheritance pattern, and ~25% of FALS cases are caused by mutations in Cu/Zn superoxide dismutase (SOD1). There is convincing evidence that mutant SOD1 (mtSOD1) kills motor neurons (MNs) because of a gain-of-function toxicity, most likely related to aggregation of mtSOD1. A number of recent reports have suggested that antibodies can be used to treat mtSOD1-induced FALS. To follow up on the use of antibodies as potential therapeutics, we generated single chain fragments of variable region antibodies (scFvs) against SOD1, and then expressed them as 'intrabodies' within a motor neuron cell line. In the present study, we describe isolation of human scFvs that interfere with mtSOD1 in vitro aggregation and toxicity. These scFvs may have therapeutic potential in sporadic ALS, as well as FALS, given that sporadic ALS may also involve abnormalities in the SOD1 protein or activity. Copyright © 2013 Elsevier Inc. All rights reserved.
-
Perera, Piyumali K; Gasser, Robin B; Jabbar, Abdul
2015-03-01
Oriental theileriosis is a tick-borne, protozoan disease of cattle caused by one or more genotypes of Theileria orientalis complex. In this study, we assessed sequence variability in a region of the 23kDa piroplasm membrane protein (p23) gene within and among three T. orientalis genotypes (designated buffeli, chitose and ikeda) in south-eastern Australia. Genomic DNA (n=100) was extracted from blood of infected cattle from various locations endemic for oriental theileriosis and tested by polymerase chain reaction (PCR)-coupled mutation scanning (single-strand conformation polymorphism (SSCP)) and targeted sequencing analysis. Eight distinct sequences represented all DNA samples, and three genotypes were found: buffeli (n=3), chitose (3) and ikeda (2). Nucleotide pairwise comparisons among these eight sequences revealed considerably higher variability among the genotypes (6.6-11.7%) than within them (0-1.9%), indicating that the p23 gene region allows the accurate identification of T. orientalis genotypes. In the future, we will combine this gene with other molecular markers to study the genetic structure of T. orientalis populations in Australasia, which will pave the way to establish a highly sensitive and specific PCR-based assay for genotypic diagnosis of infection and for assessing levels of parasitaemia in cattle. Copyright © 2014 Elsevier GmbH. All rights reserved.
-
SAbDab: the structural antibody database
Dunbar, James; Krawczyk, Konrad; Leem, Jinwoo; Baker, Terry; Fuchs, Angelika; Georges, Guy; Shi, Jiye; Deane, Charlotte M.
2014-01-01
Structural antibody database (SAbDab; http://opig.stats.ox.ac.uk/webapps/sabdab) is an online resource containing all the publicly available antibody structures annotated and presented in a consistent fashion. The data are annotated with several properties including experimental information, gene details, correct heavy and light chain pairings, antigen details and, where available, antibody–antigen binding affinity. The user can select structures, according to these attributes as well as structural properties such as complementarity determining region loop conformation and variable domain orientation. Individual structures, datasets and the complete database can be downloaded. PMID:24214988
-
NASA Astrophysics Data System (ADS)
Curtin, L.; D'Andrea, W. J.; de Wet, G.; Balascio, N.; Bradley, R. S.
2017-12-01
The climate of the North Atlantic region is extremely sensitive to changes in ocean and atmospheric circulation, and understanding past natural variability in North Atlantic climate provides important context for modern climate change. Here, we present Holocene and Eemian hydrogen isotope (δD) records from leaf waxes preserved in lacustrine sediments from the North Atlantic Faroe Islands and interpret them as a proxy for temperature and hydroclimate variability. In addition to helping to constrain the timing and amplitude of climate evolution during each of these interglacial periods, the data can be used to directly compare Eemian and Holocene climate using the same proxy from the same terrestrial location. Of the leaf waxes measured, the δD values of long-chain and mid-chain n-alkanes showed two different signals, which we interpret to represent leaf water δD values and lake water δD values, respectively. The δD values for long-chain and mid-chain fatty acids were most similar to the mid-chain n-alkanes, and likely represent a mixture of terrestrial and aquatic sources. Leaf wax-inferred δD values of precipitation during the early Holocene (10,000 to 8,000 cal yr BP) are 13‰ enriched compared to the remainder of the Holocene (after 8,000 cal yr BP), which show relatively stable values over time. Inferred lake water δD values decreased slowly over the late Holocene, suggesting a gradual transition to a wetter climate after 4,000 cal yr BP. At 2,000 cal yr BP there was a significant change in the distribution of leaf waxes that suggests a transition from shrubland to grassland, but which pre-dates the pollen evidence for this transition. The last interglacial period has been suggested as an analog for future climate conditions. We found that long-chain alkane δD values from the Eemian are most similar to the earliest Holocene, which corroborate previous pollen studies suggesting a warmer climate at the Faroe Islands during this period.
-
A System-Oriented Approach for the Optimal Control of Process Chains under Stochastic Influences
NASA Astrophysics Data System (ADS)
Senn, Melanie; Schäfer, Julian; Pollak, Jürgen; Link, Norbert
2011-09-01
Process chains in manufacturing consist of multiple connected processes in terms of dynamic systems. The properties of a product passing through such a process chain are influenced by the transformation of each single process. There exist various methods for the control of individual processes, such as classical state controllers from cybernetics or function mapping approaches realized by statistical learning. These controllers ensure that a desired state is obtained at process end despite of variations in the input and disturbances. The interactions between the single processes are thereby neglected, but play an important role in the optimization of the entire process chain. We divide the overall optimization into two phases: (1) the solution of the optimization problem by Dynamic Programming to find the optimal control variable values for each process for any encountered end state of its predecessor and (2) the application of the optimal control variables at runtime for the detected initial process state. The optimization problem is solved by selecting adequate control variables for each process in the chain backwards based on predefined quality requirements for the final product. For the demonstration of the proposed concept, we have chosen a process chain from sheet metal manufacturing with simplified transformation functions.
-
Variable context Markov chains for HIV protease cleavage site prediction.
Oğul, Hasan
2009-06-01
Deciphering the knowledge of HIV protease specificity and developing computational tools for detecting its cleavage sites in protein polypeptide chain are very desirable for designing efficient and specific chemical inhibitors to prevent acquired immunodeficiency syndrome. In this study, we developed a generative model based on a generalization of variable order Markov chains (VOMC) for peptide sequences and adapted the model for prediction of their cleavability by certain proteases. The new method, called variable context Markov chains (VCMC), attempts to identify the context equivalence based on the evolutionary similarities between individual amino acids. It was applied for HIV-1 protease cleavage site prediction problem and shown to outperform existing methods in terms of prediction accuracy on a common dataset. In general, the method is a promising tool for prediction of cleavage sites of all proteases and encouraged to be used for any kind of peptide classification problem as well.
-
Kalynych, Sergei; Ruan, Xiang; Valvano, Miguel A; Cygler, Miroslaw
2011-08-01
The O-antigen component of the lipopolysaccharide (LPS) represents a population of polysaccharide molecules with nonrandom (modal) chain length distribution. The number of the repeat O units in each individual O-antigen polymer depends on the Wzz chain length regulator, an inner membrane protein belonging to the polysaccharide copolymerase (PCP) family. Different Wzz proteins confer vastly different ranges of modal lengths (4 to >100 repeat units), despite having remarkably conserved structural folds. The molecular mechanism responsible for the selective preference for a certain number of O units is unknown. Guided by the three-dimensional structures of PCPs, we constructed a panel of chimeric molecules containing parts of two closely related Wzz proteins from Salmonella enterica and Shigella flexneri which confer different O-antigen chain length distributions. Analysis of the O-antigen length distribution imparted by each chimera revealed the region spanning amino acids 67 to 95 (region 67 to 95), region 200 to 255, and region 269 to 274 as primarily affecting the length distribution. We also showed that there is no synergy between these regions. In particular, region 269 to 274 also influenced chain length distribution mediated by two distantly related PCPs, WzzB and FepE. Furthermore, from the 3 regions uncovered in this study, region 269 to 274 appeared to be critical for the stability of the oligomeric form of Wzz, as determined by cross-linking experiments. Together, our data suggest that chain length determination depends on regions that likely contribute to stabilize a supramolecular complex.
-
Wen, Yangming; Lan, Kaijian; Wang, Junjie; Yu, Jingyi; Qu, Yarong; Zhao, Wei; Zhang, Fuchun; Tan, Wanlong; Cao, Hong; Zhou, Chen
2013-06-01
To construct dengue virus-specific full-length fully human antibody libraries using mammalian cell surface display technique. Total RNA was extracted from peripheral blood mononuclear cells (PBMCs) from convalescent patients with dengue fever. The reservoirs of the light chain and heavy chain variable regions (LCκ and VH) of the antibody genes were amplified by RT-PCR and inserted into the vector pDGB-HC-TM separately to construct the light chain and heavy chain libraries. The library DNAs were transfected into CHO cells and the expression of full-length fully human antibodies on the surface of CHO cells was analyzed by flow cytometry. Using 1.2 µg of the total RNA isolated from the PBMCs as the template, the LCκ and VH were amplified and the full-length fully human antibody mammalian display libraries were constructed. The kappa light chain gene library had a size of 1.45×10(4) and the heavy chain gene library had a size of 1.8×10(5). Sequence analysis showed that 8 out of the 10 light chain clones and 7 out of the 10 heavy chain clones randomly picked up from the constructed libraries contained correct open reading frames. FACS analysis demonstrated that all the 15 clones with correct open reading frames expressed full-length antibodies, which could be detected on CHO cell surfaces. After co-transfection of the heavy chain and light chain gene libraries into CHO cells, the expression of full-length antibodies on CHO cell surfaces could be detected by FACS analysis with an expressible diversity of the antibody library reaching 1.46×10(9) [(1.45×10(4)×80%)×(1.8×10(5)×70%)]. Using 1.2 µg of total RNA as template, the LCκ and VH full-length fully human antibody libraries against dengue virus have been successfully constructed with an expressible diversity of 10(9).
-
Ladenson, Ruth C; Crimmins, Dan L; Landt, Yvonne; Ladenson, Jack H
2006-07-01
We have isolated and characterized a caffeine-specific, heavy-chain-only antibody fragment (V(HH)) from llama that is capable of being utilized to analyze caffeine in hot and cold beverages. Camelid species (llama and camel) were selected for immunization because of their potential to make heat-stable, heavy-chain-only antibodies. Llamas and camels were immunized with caffeine covalently linked to keyhole limpet hemocyanin, and recombinant antibody techniques were used to create phage displayed libraries of variable region fragments of the heavy-chain antibodies. Caffeine-specific V(HH) fragments were selected by their ability to bind to caffeine/bovine serum albumin (BSA) and confirmed by a positive reaction in a caffeine enzyme-linked immunosorbent assay (caffeine ELISA). One of these V(HH) fragments (VSA2) was expressed as a soluble protein and shown to recover its reactivity after exposure to temperatures up to 90 degrees C. In addition, VSA2 was able to bind caffeine at 70 degrees C. A competition caffeine ELISA was developed for the measurement of caffeine in beverages, and concentrations of caffeine obtained for coffee, Coca-Cola Classic, and Diet Coke agreed well with high performance liquid chromatography (HPLC) determination and literature values. VSA2 showed minimal cross reactivity with structurally related methylxanthines.
-
Measles re-emergence in Northern Italy: Pathways of measles virus genotype D8, 2013-2014.
Amendola, Antonella; Bianchi, Silvia; Lai, Alessia; Canuti, Marta; Piralla, Antonio; Baggieri, Melissa; Ranghiero, Alberto; Piatti, Alessandra; Tanzi, Elisabetta; Zehender, Gianguglielmo; Magurano, Fabio; Baldanti, Fausto
2017-03-01
Molecular surveillance and advanced phylogenetic methods are important tools to track the pathways of Measles virus (MV) genotypes, provide evidence for the interruption of endemic transmission and verify the elimination of the disease. The aims of this study were to describe the genetic profile of MV genotype D8 (D8-MV) strains circulating in Northern Italy (Lombardy Region) during the 2013-2014 period and to analyze the transmission chains and estimate the introduction time points using a phylogenetic approach. Forty-four strains of D8-MV identified from 12 outbreaks and 28 cases reported as sporadic were analyzed. Molecular analysis was performed by sequencing the highly variable 450nt region of the N gene of MV genome (N-450), as recommended by the WHO. Phylogenetic analyses and tree time-scaled reconstruction were performed with BEAST software. We could trace back the transmission pathways that resulted in three chains of transmission, two introductions with limited spread (two familiar outbreaks), and two single introductions (true sporadic cases). The D8-Taunton transmission chain, which was involved in 7 outbreaks and 13 sporadic cases, was endemic during the studied period. Furthermore, two novel local variants emerged independently in March 2014 and caused two transmission chains linked to at least 3 outbreaks. Overall, viral diversity was high and strains belonging to 5 different variants were identified. The results of this study clearly demonstrate that multiple lineages of D8-MV co-circulated in Northern Italy. Measles can be considered a re-emerging disease in Italy and additional efforts are necessary to achieve measles elimination goal. Copyright © 2016 Elsevier B.V. All rights reserved.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Hernández-Santoyo, A.; Del Pozo Yauner, L; Fuentes-Silva, D
Systemic amyloid light-chain (LC) amyloidosis is a disease process characterized by the pathological deposition of monoclonal LCs in tissue. All LC subtypes are capable of fibril formation although {lambda} chains, particularly those belonging to the {lambda}6 type, are overrepresented. Here, we report the thermodynamic and in vitro fibrillogenic properties of several mutants of the {lambda}6 protein 6aJL2 in which Pro7 and/or His8 was substituted by Ser or Pro. The H8P and H8S mutants were almost as stable as the wild-type protein and were poorly fibrillogenic. In contrast, the P7S mutation decreased the thermodynamic stability of 6aJL2 and greatly enhanced itsmore » capacity to form amyloid-like fibrils in vitro. The crystal structure of the P7S mutant showed that the substitution induced both local and long-distance effects, such as the rearrangement of the VL (variable region of the light chain)-VL interface. This mutant crystallized in two orthorhombic polymorphs, P2{sub 1}2{sub 1}2{sub 1} and C222{sub 1}. In the latter, a monomer that was not arranged in the typical Bence-Jones dimer was observed for the first time. Crystal-packing analysis of the C222{sub 1} lattice showed the establishment of intermolecular {beta}-{beta} interactions that involved the N-terminus and {beta}-strand B and that these could be relevant in the mechanism of LC fibril formation. Our results strongly suggest that Pro7 is a key residue in the conformation of the N-terminal sheet switch motif and, through long-distance interactions, is also critically involved in the contacts that stabilized the VL interface in {lambda}6 LCs.« less
-
Velázquez-López, Isabel; Valdés-García, Gilberto; Romero Romero, Sergio; Maya Martínez, Roberto; Leal-Cervantes, Ana I; Costas, Miguel; Sánchez-López, Rosana; Amero, Carlos; Pastor, Nina; Fernández Velasco, D Alejandro
2018-07-01
Solvent conditions modulate the expression of the amyloidogenic potential of proteins. In this work the effect of pH on the fibrillogenic behavior and the conformational properties of 6aJL2, a model protein of the highly amyloidogenic variable light chain λ6a gene segment, was examined. Ordered aggregates showing the ultrastructural and spectroscopic properties observed in amyloid fibrils were formed in the 2.0-8.0 pH range. At pH <3.0 a drastic decrease in lag time and an increase in fibril formation rate were found. In the 4.0-8.0 pH range there was no spectroscopic evidence for significant conformational changes in the native state. Likewise, heat capacity measurements showed no evidence for residual structure in the unfolded state. However, at pH <3.0 stability is severely decreased and the protein suffers conformational changes as detected by circular dichroism, tryptophan and ANS fluorescence, as well as by NMR spectroscopy. Molecular dynamics simulations indicate that acid-induced conformational changes involve the exposure of the loop connecting strands E and F. These results are compatible with pH-induced changes in the NMR spectra. Overall, the results indicate that the mechanism involved in the acid-induced increase in the fibrillogenic potential of 6aJL2 is profoundly different to that observed in κ light chains, and is promoted by localized conformational changes in a region of the protein that was previously not known to be involved in acid-induced light chain fibril formation. The identification of this region opens the potential for the design of specific inhibitors. Copyright © 2018 Elsevier B.V. All rights reserved.
-
USDA-ARS?s Scientific Manuscript database
Deoxynivalenol (DON)is a mycotoxin produced by certain fungi that infest cereal grains worldwide. A hybridoma cell line producing a monoclonal antibody (Mab) recognizing DON was used as the starting point in the development of a recombinant single chain variable fragment (scFv) antibody. The scFv wa...
-
Bacterial Polysaccharide Co-Polymerases Share a Common Framework for Control of Polymer Length
DOE Office of Scientific and Technical Information (OSTI.GOV)
Tocilj,A.; Munger, C.; Proteau, A.
2008-01-01
The chain length distribution of complex polysaccharides present on the bacterial surface is determined by polysaccharide co-polymerases (PCPs) anchored in the inner membrane. We report crystal structures of the periplasmic domains of three PCPs that impart substantially different chain length distributions to surface polysaccharides. Despite very low sequence similarities, they have a common protomer structure with a long central alpha-helix extending 100 Angstroms into the periplasm. The protomers self-assemble into bell-shaped oligomers of variable sizes, with a large internal cavity. Electron microscopy shows that one of the full-length PCPs has a similar organization as that observed in the crystal formore » its periplasmic domain alone. Functional studies suggest that the top of the PCP oligomers is an important region for determining polysaccharide modal length. These structures provide a detailed view of components of the bacterial polysaccharide assembly machinery.« less
-
Dynamics of Extreme Floods in Southeast and South Brazil
NASA Astrophysics Data System (ADS)
Ribeiro Lima, C. H.; Lall, U.
2015-12-01
Many extreme floods result from a causal chain, where exceptional rain and floods in water basins from different sizes are related to large scale, anomalous and persistent patterns in atmospheric and oceanic circulation. Organized moisture plumes from oceanic sources are often implicated. One could use an Eulerian-Lagrangian climate model to test a causal chain hypothesis, but the parameterization and testing of such a model covering convection and transport continues to be a challenge. Consequently, empirical data based studies can be useful to establish the need to formally model such events using this approach. Here we consider two flood-prone regions in Southeast and South Brazil as case studies. A hypothesis of the causal chain of extreme floods in these regions is investigated by means of observed streamflow and reanalysis data and some machine learning tools. The signatures of the organization of the large scale atmospheric circulation in the days prior to the flood events are evaluated based on the integrated moisture flux and its divergence field and storm track data, so that a better understanding of the relations between the flood magnitude and duration, strength of moisture convergence and role of regional moisture recycling or teleconnected moisture is established. Persistent patterns and anomalies in the sea surface temperature (SST) field in the Pacific and Atlantic oceans that may be associated with disturbances in the atmospheric circulation and with the flood dynamics are investigated through composite analysis. Finally, machine learning algorithms for nonlinear dimension reduction are employed to visualize and understand some of the spatio-temporal patterns of the dominated climate variables in a reduced dimensional space. Prospects for prediction are discussed.
-
Panunzio, M F; Caporizzi, R; Lagravinese, D; Conversano, M
2015-01-01
Every year the Italian Ministry of Health, on the basis of regional data, draws up the "Report on Official Controls" to be submitted to the Parliament. The report contains abundant data, diagrams and charts and illustrates the number and type of official controls (OC) performed by the pertinent Bodies (Ministry of Health, Regional and Local Health Authorities) over the previous year on Food Business Operators (FBO), in accordance with the EC Regulation 882/2004. The trend - which has consolidated over the years - relates to the multiplicity of OC and shows a decrease of such controls compared to an increase in "non-conformities". OC frequency is established by the Regional Authorities on the basis of the categorisation of both a "generic risk" for companies calculated taking into account the probability of occurrence of a "non-conformity", and a "specific" risk, assessed on the basis of the results of the OC actually performed on a given "Operatore del Settore Alimentare" (Food Sector Operator, in Italian: OSA). Thus, categorisation (i.e. the probability of occurrence of non-conformities) is the main driver of the OC scheduling and planning process. We have been asking ourselves whether the current OC planning/scheduling method is still suitable for ensuring food safety in the face of internalisation of the food supply chain. As a matter of fact, food safety is now becoming increasingly variable due to the globalization of consumption where "farm to fork", rather than "border to fork", food safety must be ensured. On the basis of these considerations, a different OC planning /scheduling method is being proposed based on the assessment of risks and the estimation of the occurrence of the same along the agro-food chain.
-
NASA Astrophysics Data System (ADS)
Massa, C.; Beilman, D. W.; Nichols, J. E.; Elison Timm, O.
2016-12-01
Holocene peat deposits from the Hawaiian Islands provide a unique opportunity to resolve millennial to centennial-scale climate variability over the central Pacific region, where data remain scarce. Because both extratropical and tropical modes of climate variability have a strong influence on modern rainfall over the archipelago, hydroclimate proxies from peat would provide valuable information about past Pacific climate changes. The few terrestrial records studied, based on pollen or leaf wax biomarkers, showed evidence for substantial vegetation changes that have been linked to a drying trend over the Holocene. Leaf wax n-alkanes, as well as their stable isotopic compositions (δ13C and δD), are indeed increasingly used to reconstruct past hydroclimate conditions. The interpretation of n-alkanes as biomarkers requires however a thorough knowledge of their distribution in modern plants that contribute to sediments, but in Hawaii the modern vegetation is understudied compared to proxy applications. Here we report results from a preliminary investigation of n-alkanes distributions in dominant modern plant litter collected at a bog site at the summit of the Waianae mountains on the Island of Oahu. We compared n-alkane distributions among species and plant groups in order to test whether taxa or plant functional types (mosses, ferns, woody plants, and sedges) can be discriminated from their n-alkane profiles. Results showed that general plant groups were difficult to distinguish based on individual n-alkanes abundances, chain lengths, or ratios. At the species level, the sedge Machaerina augustifolia, was largely dominated by n-C29 ( 60%), suggesting some chain lengths could be useful as proxies for identifying the contribution of sedges to sedimentary records. Woody plant average chain length was highly variable but overall was not shorter (even slightly higher) than in other terrestrial plants, as it is often assumed. A sedimentary profile from this site shows variation and an overall decrease in n-alkane chain length over the Holocene, but patterns across common modern plants suggest that caution should be exercised when ascribing n-alkane distribution parameters to a specific group of tropical vegetation.
-
NASA Astrophysics Data System (ADS)
Adam, Claudia; Vidal, Valerie; Escartín, Javier
2007-09-01
Walvis and St. Helena are the only long-lived hotspot chains in the South Atlantic. Therefore, their characterization is important to constrain the processes associated with mantle plume formation, their temporal evolution, and the interaction with plate and mantle dynamics in the region. We study the temporal evolution of plume buoyancy and magma production rate along both hotspot chains, which are constrained from the swell and volume of volcanic materials emplaced along the chain. The regional depth anomaly is calculated by correcting the 2' bathymetry grid of Smith and Sandwell [W.H.F. Smith, D.T. Sandwell, Global sea floor topography from satellite altimetry and ship depth soundings, Science 277 (1997) 1956-1962] for thermal subsidence and sediment loading. We separate the topography associated with volcanism and the swell surrounding the hotspot chains using the MiFil filtering method [C. Adam, V. Vidal, A. Bonneville, MiFil: A method to characterize seafloor swells with application to the south central Pacific, Geochem. Geophys. Geosyst. 6 (1) (2005) Q01003, doi: 10.1029/2004GC000814]. We then estimate the temporal variations associated with both parameters by computing volumes along the hotspot tracks. Neither Walvis nor St. Helena show a 'classical' hotspot behavior. We find that two plumes are at the origin of the St. Helena chain. This study also shows a swell associated with the Circe seamount, supporting the existence of a hotspot NW of the St. Helena trail. The variation in swell and volcanic fluxes suggests temporal variability in the plume behavior at time scales of 10-20 m.y. and 5 m.y., which may be related to oscillations and instabilities of the plume conduit, respectively. Cumulative fluxes in the area are largest for Walvis and weakest for Circe, and all are significantly lower than that reported for the Hawai'i hotspot.
-
Sunagar, Kartik; Undheim, Eivind A B; Scheib, Holger; Gren, Eric C K; Cochran, Chip; Person, Carl E; Koludarov, Ivan; Kelln, Wayne; Hayes, William K; King, Glenn F; Antunes, Agosthino; Fry, Bryan Grieg
2014-03-17
Due to the extreme variation of venom, which consequently results in drastically variable degrees of neutralization by CroFab antivenom, the management and treatment of envenoming by Crotalus oreganus helleri (the Southern Pacific Rattlesnake), one of the most medically significant snake species in all of North America, has been a clinician's nightmare. This snake has also been the subject of sensational news stories regarding supposed rapid (within the last few decades) evolution of its venom. This research demonstrates for the first time that variable evolutionary selection pressures sculpt the intraspecific molecular diversity of venom components in C. o. helleri. We show that myotoxic β-defensin peptides (aka: crotamines/small basic myotoxic peptides) are secreted in large amounts by all populations. However, the mature toxin-encoding nucleotide regions evolve under the constraints of negative selection, likely as a result of their non-specific mode of action which doesn't enforce them to follow the regime of the classic predator-prey chemical arms race. The hemorrhagic and tissue destroying snake venom metalloproteinases (SVMPs) were secreted in larger amounts by the Catalina Island and Phelan rattlesnake populations, in moderate amounts in the Loma Linda population and in only trace levels by the Idyllwild population. Only the Idyllwild population in the San Jacinto Mountains contained potent presynaptic neurotoxic phospholipase A2 complex characteristic of Mohave Rattlesnake (Crotalus scutulatus) and Neotropical Rattlesnake (Crotalus durissus terrificus). The derived heterodimeric lectin toxins characteristic of viper venoms, which exhibit a diversity of biological activities, including anticoagulation, agonism/antagonism of platelet activation, or procoagulation, appear to have evolved under extremely variable selection pressures. While most lectin α- and β-chains evolved rapidly under the influence of positive Darwinian selection, the β-chain lectin of the Catalina Island population appears to have evolved under the constraint of negative selection. Both lectin chains were conspicuously absent in both the proteomics and transcriptomics of the Idyllwild population. Thus, we not only highlight the tremendous biochemical diversity in C. o. helleri's venom-arsenal, but we also show that they experience remarkably variable strengths of evolutionary selection pressures, within each toxin class among populations and among toxin classes within each population. The mapping of geographical venom variation not only provides additional information regarding venom evolution, but also has direct medical implications by allowing prediction of the clinical effects of rattlesnake bites from different regions. Such information, however, also points to these highly variable venoms as being a rich source of novel toxins which may ultimately prove to be useful in drug design and development. These results have direct implications for the treatment of envenomed patients. The variable venom profile of Crotalus oreganus helleri underscores the biodiscovery potential of novel snake venoms. Copyright © 2014 Elsevier B.V. All rights reserved.
-
Discrete-Choice Modeling Of Non-Working Women’s Trip-Chaining Activity Based
NASA Astrophysics Data System (ADS)
Hayati, Amelia; Pradono; Purboyo, Heru; Maryati, Sri
2018-05-01
Start The urban developments of technology and economics are now changing the lifestyles of the urban societies. It is also changing their travel demand to meet their movement needs. Nowadays, urban women, especially in Bandung, West Java, have a high demand for their daily travel and tend to increase. They have the ease of accessibility to personal modes of transportation and freedom to go anywhere to meet their personal and family needs. This also happens to non-working women or as housewives in the city of Bandung. More than 50% of women’s mobility is outside the home, in the term of trip-chaining, from leaving to returning home in one day. It is based on their complex activities in order to meet the needs of family and home care. While less than 60% of male’s mobility is outdoors, it is a simple trip-chaining or only has a single trip. The trip-chaining has significant differences between non-working women and working-men. This illustrates the pattern of Mom and Dad’s mobility in a family with an activity-based approach for the same purpose, i.e. family welfare. This study explains how complex the trip-chaining of non-working urban women and as housewives, with an activity-based approach done outdoors in a week. Socio-economic and household demographic variables serve as the basis for measuring the independent variables affecting family welfare, as well as the variables of type, time and duration of activities performed by unemployed housewives. This study aims to examine the interrelationships between activity variables, especially the time of activity and travel, and socio-economic of household variables that can generate the complexity of women’s daily travel. Discrete Choice Modeling developed by Ben-Akiva, Chandra Bhat, etc., is used in this study to illustrate the relationship between activity and socio-economic demographic variables based on primary survey data in Bandung, West Java for 466 unemployed housewives. The results of the regression, by Seemingly Unrelated Regression approach methods, showed the interrelationship between all variables, including the complexity of trip chaining of housewives based on their daily activities. The type of mandatory and discretionary activities, and the duration of activities performed during the dismissal in the series of trip chains conducted are intended for the fulfillment of the welfare of all family member.
-
Segmental Versican Expression in the Trabecular Meshwork and Involvement in Outflow Facility
Keller, Kate E.; Bradley, John M.; Vranka, Janice A.
2011-01-01
Purpose. Versican is a large proteoglycan with numerous chondroitin sulfate (CS) glycosaminoglycan (GAG) side chains attached. To assess versican's potential contributions to aqueous humor outflow resistance, its segmental distribution in the trabecular meshwork (TM) and the effect on outflow facility of silencing the versican gene were evaluated. Methods. Fluorescent quantum dots (Qdots) were perfused to label outflow pathways of anterior segments. Immunofluorescence with confocal microscopy and quantitative RT-PCR were used to determine versican protein and mRNA distribution relative to Qdot-labeled regions. Lentiviral delivery of shRNA-silencing cassettes to TM cells in perfused anterior segment cultures was used to evaluate the involvement of versican and CS GAG chains in outflow facility. Results. Qdot uptake by TM cells showed considerable segmental variability in both human and porcine outflow pathways. Regional levels of Qdot labeling were inversely related to versican protein and mRNA levels; versican levels were relatively high in sparsely Qdot-labeled regions and low in densely labeled regions. Versican silencing decreased outflow facility in human and increased facility in porcine anterior segments. However, RNAi silencing of ChGn, an enzyme unique to CS GAG biosynthesis, increased outflow facility in both species. The fibrillar pattern of versican immunostaining in the TM juxtacanalicular region was disrupted after versican silencing in perfusion culture. Conclusions. Versican appears to be a central component of the outflow resistance, where it may organize GAGs and other ECM components to facilitate and control open flow channels in the TM. However, the exact molecular organization of this resistance appears to differ between human and porcine eyes. PMID:21596823
-
NASA Astrophysics Data System (ADS)
Gwal, A. K.; Shrivastava, A.
2006-11-01
ak_gwal@yahoo.co.in The scientists have found that the accumulation of tectonic energy is localized in certain places and is not universal. Taking into account this hypothesis the authors have studied the sequence of occurrence rate of the earthquakes (M≥5) in the South-East Asian region, as the chronological data related to the occurrence of earthquakes collected in that region for last five years i.e. from 2001 to 2005 have revealed that the disastrous tsunami events which took place on 26th December, 2004 as an effect of Sumatra earthquake( M=9) have increased the occurrence of earthquake frequency for a longer period (which might be due to adjustment of tectonic plates). Observing these facts i.e. sudden enhancement in occurrence rate of earthquakes, the authors have availed this opportunity to further explore the concept of seismoelectromagnetic-ionospheric phenomena, which still needs a lot of statistical evidences, comprising tremendous amount of data to establish it. In this paper the authors have tried to analyze the chain of observations made and data collected and stored month wise w.e.f. 26th December, 2004 to 31st March, 2005 in the region, using DEMETER satellite. Further, efforts have also been made to provide the statistical analysis of the ionospheric variability caused due to detected electromagnetic burst in ULF frequency ranges in the context of natural variability in order to distinguish the variability introduced by other sources. In brief, it could be concluded that there is possibility of getting the electromagnetic precursors in the ionosphere at different frequency ranges due to excess release of tectonic energy as a result of occurrence rate of the earthquakes in the region.
-
Customer involvement in greening the supply chain: an interpretive structural modeling methodology
NASA Astrophysics Data System (ADS)
Kumar, Sanjay; Luthra, Sunil; Haleem, Abid
2013-04-01
The role of customers in green supply chain management needs to be identified and recognized as an important research area. This paper is an attempt to explore the involvement aspect of customers towards greening of the supply chain (SC). An empirical research approach has been used to collect primary data to rank different variables for effective customer involvement in green concept implementation in SC. An interpretive structural-based model has been presented, and variables have been classified using matrice d' impacts croises- multiplication appliqué a un classement analysis. Contextual relationships among variables have been established using experts' opinions. The research may help practicing managers to understand the interaction among variables affecting customer involvement. Further, this understanding may be helpful in framing the policies and strategies to green SC. Analyzing interaction among variables for effective customer involvement in greening SC to develop the structural model in the Indian perspective is an effort towards promoting environment consciousness.
-
Martínez-Ruiz, Francisco José; Blas, Felipe J; Moreno-Ventas Bravo, A Ignacio; Míguez, José Manuel; MacDowell, Luis G
2017-05-17
The statistical associating fluid theory for attractive potentials of variable range (SAFT-VR) density functional theory (DFT) developed by [Gloor et al., J. Chem. Phys., 2004, 121, 12740-12759] is used to predict the interfacial behaviour of molecules modelled as fully-flexible square-well chains formed from tangentially-bonded monomers of diameter σ and potential range λ = 1.5σ. Four different model systems, comprising 4, 8, 12, and 16 monomers per molecule, are considered. In addition to that, we also compute a number of interfacial properties of molecular chains from direct simulation of the vapour-liquid interface. The simulations are performed in the canonical ensemble, and the vapour-liquid interfacial tension is evaluated using the wandering interface (WIM) method, a technique based on the thermodynamic definition of surface tension. Apart from surface tension, we also obtain density profiles, coexistence densities, vapour pressures, and critical temperature and density, paying particular attention to the effect of the chain length on these properties. According to our results, the main effect of increasing the chain length (at fixed temperature) is to sharpen the vapour-liquid interface and to increase the width of the biphasic coexistence region. As a result, the interfacial thickness decreases and the surface tension increases as the molecular chains get longer. The interfacial thickness and surface tension appear to exhibit an asymptotic limiting behaviour for long chains. A similar behaviour is also observed for the coexistence densities and critical properties. Agreement between theory and simulation results indicates that SAFT-VR DFT is only able to predict qualitatively the interfacial properties of the model. Our results are also compared with simulation data taken from the literature, including the vapour-liquid coexistence densities, vapour pressures, and surface tension.
-
Graded, Dynamically Routable Information Processing with Synfire-Gated Synfire Chains.
Wang, Zhuo; Sornborger, Andrew T; Tao, Louis
2016-06-01
Coherent neural spiking and local field potentials are believed to be signatures of the binding and transfer of information in the brain. Coherent activity has now been measured experimentally in many regions of mammalian cortex. Recently experimental evidence has been presented suggesting that neural information is encoded and transferred in packets, i.e., in stereotypical, correlated spiking patterns of neural activity. Due to their relevance to coherent spiking, synfire chains are one of the main theoretical constructs that have been appealed to in order to describe coherent spiking and information transfer phenomena. However, for some time, it has been known that synchronous activity in feedforward networks asymptotically either approaches an attractor with fixed waveform and amplitude, or fails to propagate. This has limited the classical synfire chain's ability to explain graded neuronal responses. Recently, we have shown that pulse-gated synfire chains are capable of propagating graded information coded in mean population current or firing rate amplitudes. In particular, we showed that it is possible to use one synfire chain to provide gating pulses and a second, pulse-gated synfire chain to propagate graded information. We called these circuits synfire-gated synfire chains (SGSCs). Here, we present SGSCs in which graded information can rapidly cascade through a neural circuit, and show a correspondence between this type of transfer and a mean-field model in which gating pulses overlap in time. We show that SGSCs are robust in the presence of variability in population size, pulse timing and synaptic strength. Finally, we demonstrate the computational capabilities of SGSC-based information coding by implementing a self-contained, spike-based, modular neural circuit that is triggered by streaming input, processes the input, then makes a decision based on the processed information and shuts itself down.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Oksenberg, J.R.; Cavalli-Sforza, L.L.; Steinman, L.
1989-02-01
Polymorphic markers in genes encoding the {alpha} chain of the human T-cell receptor (TcR) have been detected by Southern blot analysis in Pss I digests. Polymorphic bands were observed at 6.3 and 2.0 kilobases (kb) with frequencies of 0.30 and 0.44, respectively, in the general population. Using the polymerase chain reaction (PCR) method, the authors amplified selected sequences derived from the full-length TcR {alpha} cDNA probe. These PcR products were used as specific probes to demonstrate that the 6.3-kb polymorphic fragment hybridizes to the variable (V)-region probe and the 2.0-kb fragment hybridizes to the constant (C)-region probe. Segregation of themore » polymorphic bands was analyzed in family studies. To look for associations between these markers and autoimmune diseases, the authors have studied the restriction fragment length polymorphism distribution of the Pss I markers in patients with multiple sclerosis, myasthenia gravis, and Graves disease. Significant differences in the frequency of the polymorphic V{sub {alpha}} and C{sub {alpha}} markers were identified between patients and healthy individuals.« less
-
Dooley, Helen; Stanfield, Robyn L; Brady, Rebecca A; Flajnik, Martin F
2006-02-07
The cartilaginous fish are the oldest phylogenetic group in which Igs have been found. Sharks produce a unique Ig isotype, IgNAR, a heavy-chain homodimer that does not associate with light chains. Instead, the variable (V) regions of IgNAR bind antigen as soluble single domains. Our group has shown that IgNAR plays an integral part in the humoral response of nurse sharks (Ginglymostoma cirratum) upon antigen challenge. Here, we generated phage-displayed libraries of IgNAR V regions from an immunized animal and found a family of clones derived from the same rearrangement event but differentially mutated during expansion. Because of the cluster organization of shark Ig genes and the paucicopy nature of IgNAR, we were able to construct the putative ancestor of this family. By studying mutations in the context of clone affinities, we found evidence that affinity maturation occurs for this isotype. Subsequently, we were able to identify mutations important in the affinity improvement of this family. Because the family clones were all obtained after immunization, they provide insight into the in vivo maturation mechanisms, in general, and for single-domain antibody fragments.
-
Housset, D; Mazza, G; Grégoire, C; Piras, C; Malissen, B; Fontecilla-Camps, J C
1997-01-01
The crystal structure of a mouse T-cell antigen receptor (TCR) Fv fragment complexed to the Fab fragment of a specific anti-clonotypic antibody has been determined to 2.6 A resolution. The polypeptide backbone of the TCR V alpha domain is very similar to those of other crystallographically determined V alphas, whereas the V beta structure is so far unique among TCR V beta domains in that it displays a switch of the c" strand from the inner to the outer beta-sheet. The beta chain variable region of this TCR antigen-binding site is characterized by a rather elongated third complementarity-determining region (CDR3beta) that packs tightly against the CDR3 loop of the alpha chain, without leaving any intervening hydrophobic pocket. Thus, the conformation of the CDR loops with the highest potential diversity distinguishes the structure of this TCR antigen-binding site from those for which crystallographic data are available. On the basis of all these results, we infer that a significant conformational change of the CDR3beta loop found in our TCR is required for binding to its cognate peptide-MHC ligand. PMID:9250664
-
Santini, A C; Magalhães, J T; Cascardo, J C M; Corrêa, R X
2016-04-28
Chromobacterium violaceum is a free-living Gram-negative bacillus usually found in the water and soil in tropical regions, which causes infections in humans. Chromobacteriosis is characterized by rapid dissemination and high mortality. The aim of this study was to detect the genetic variability among C. violaceum type strain ATCC 12472, and seven isolates from the environment and one from a pulmonary secretion from a chromobacteriosis patient from Ilhéus, Bahia. The molecular characterization of all samples was performed by polymerase chain reaction (PCR) sequencing and 16S rDNA analysis. Primers specific for two ATCC 12472 pathogenicity genes, hilA and yscD, as well as random amplified polymorphic DNA (RAPD), were used for PCR amplification and comparative sequencing of the products. For a more specific approach, the PCR products of 16S rDNA were digested with restriction enzymes. Seven of the samples, including type-strain ATCC 12472, were amplified by the hilA primers; these were subsequently sequenced. Gene yscD was amplified only in type-strain ATCC 12472. MspI and AluI digestion revealed 16S rDNA polymorphisms. This data allowed the generation of a dendogram for each analysis. The isolates of C. violaceum have variability in random genomic regions demonstrated by RAPD. Also, these isolates have variability in pathogenicity genes, as demonstrated by sequencing and restriction enzyme digestion.
-
Feige, Matthias J; Gräwert, Melissa A; Marcinowski, Moritz; Hennig, Janosch; Behnke, Julia; Ausländer, David; Herold, Eva M; Peschek, Jirka; Castro, Caitlin D; Flajnik, Martin; Hendershot, Linda M; Sattler, Michael; Groll, Michael; Buchner, Johannes
2014-06-03
Sharks and other cartilaginous fish are the phylogenetically oldest living organisms that rely on antibodies as part of their adaptive immune system. They produce the immunoglobulin new antigen receptor (IgNAR), a homodimeric heavy chain-only antibody, as a major part of their humoral adaptive immune response. Here, we report the atomic resolution structure of the IgNAR constant domains and a structural model of this heavy chain-only antibody. We find that despite low sequence conservation, the basic Ig fold of modern antibodies is already present in the evolutionary ancient shark IgNAR domains, highlighting key structural determinants of the ubiquitous Ig fold. In contrast, structural differences between human and shark antibody domains explain the high stability of several IgNAR domains and allowed us to engineer human antibodies for increased stability and secretion efficiency. We identified two constant domains, C1 and C3, that act as dimerization modules within IgNAR. Together with the individual domain structures and small-angle X-ray scattering, this allowed us to develop a structural model of the complete IgNAR molecule. Its constant region exhibits an elongated shape with flexibility and a characteristic kink in the middle. Despite the lack of a canonical hinge region, the variable domains are spaced appropriately wide for binding to multiple antigens. Thus, the shark IgNAR domains already display the well-known Ig fold, but apart from that, this heavy chain-only antibody employs unique ways for dimerization and positioning of functional modules.
-
USDA-ARS?s Scientific Manuscript database
‘Ca. Liberibacter asiaticus’ is the causal agent of citrus huanglongbing, the most serious disease of citrus worldwide. We have developed and applied immunization and affinity screening methods to develop a primary library of recombinant single chain variable fragment (scFv) antibodies in an M13 vec...
-
Modeling Protoplanetary Disks to Characterize the Evolution of their Structure
NASA Astrophysics Data System (ADS)
Allen, Magdelena; van der Marel, Nienke; Williams, Jonathan
2018-01-01
Stars form from gravitationally collapsing clouds of gas and dust. Most young stars retain a protoplanetary disk for a few million years. This disk’s dust reemits stellar flux in the infrared, producing a spectral energy distribution (SED) observable by Spitzer and other telescopes. To understand the inner clearing of dust cavities and evolution in the SED, we used the Chiang & Goldreich two-layer approximation. We first wrote a python script based on refinements by Dullemond that includes a hot, puffed inner rim, shadowed mid region, flaring outer disk, and a variable inner cavity. This was then coupled with a Markov Chain Monte Carlo procedure to fit the observed SEDs of disks in the star forming Lupus region. The fitting procedure recovers physical characteristics of the disk including temperature, size, mass, and surface density. We compare the characteristics of circumstellar disks without holes and more evolved transition disks with cleared inner regions.
-
NASA Astrophysics Data System (ADS)
Guilbert, Justin; Betts, Alan K.; Rizzo, Donna M.; Beckage, Brian; Bomblies, Arne
2015-03-01
We present evidence of increasing persistence in daily precipitation in the northeastern United States that suggests that global circulation changes are affecting regional precipitation patterns. Meteorological data from 222 stations in 10 northeastern states are analyzed using Markov chain parameter estimates to demonstrate that a significant mode of precipitation variability is the persistence of precipitation events. We find that the largest region-wide trend in wet persistence (i.e., the probability of precipitation in 1 day and given precipitation in the preceding day) occurs in June (+0.9% probability per decade over all stations). We also find that the study region is experiencing an increase in the magnitude of high-intensity precipitation events. The largest increases in the 95th percentile of daily precipitation occurred in April with a trend of +0.7 mm/d/decade. We discuss the implications of the observed precipitation signals for watershed hydrology and flood risk.
-
Mouse Vk gene classification by nucleic acid sequence similarity.
Strohal, R; Helmberg, A; Kroemer, G; Kofler, R
1989-01-01
Analyses of immunoglobulin (Ig) variable (V) region gene usage in the immune response, estimates of V gene germline complexity, and other nucleic acid hybridization-based studies depend on the extent to which such genes are related (i.e., sequence similarity) and their organization in gene families. While mouse Igh heavy chain V region (VH) gene families are relatively well-established, a corresponding systematic classification of Igk light chain V region (Vk) genes has not been reported. The present analysis, in the course of which we reviewed the known extent of the Vk germline gene repertoire and Vk gene usage in a variety of responses to foreign and self antigens, provides a classification of mouse Vk genes in gene families composed of members with greater than 80% overall nucleic acid sequence similarity. This classification differed in several aspects from that of VH genes: only some Vk gene families were as clearly separated (by greater than 25% sequence dissimilarity) as typical VH gene families; most Vk gene families were closely related and, in several instances, members from different families were very similar (greater than 80%) over large sequence portions; frequently, classification by nucleic acid sequence similarity diverged from existing classifications based on amino-terminal protein sequence similarity. Our data have implications for Vk gene analyses by nucleic acid hybridization and describe potentially important differences in sequence organization between VH and Vk genes.
-
NASA Astrophysics Data System (ADS)
Thomas, R.; Connell, D.; Spears, T.; Leadbetter, A.; Burger, E. F.
2016-12-01
The scientific literature heavily features small-scale studies with the impact of the results extrapolated to regional/global importance. There are on-going initiatives (e.g. OA-ICC, GOA-ON, GEOTRACES, EMODNet Chemistry) aiming to assemble regional to global-scale datasets that are available for trend or meta-analyses. Assessing the quality and comparability of these data requires information about the processing chain from "sampling to spreadsheet". This provenance information needs to be captured and readily available to assess data fitness for purpose. The NOAA Ocean Acidification metadata template was designed in consultation with domain experts for this reason; the core carbonate chemistry variables have 23-37 metadata fields each and for scientists generating these datasets there could appear to be an ever increasing amount of metadata expected to accompany a dataset. While this provenance metadata should be considered essential by those generating or using the data, for those discovering data there is a sliding scale between what is considered discovery metadata (title, abstract, contacts, etc.) versus usage metadata (methodology, environmental setup, lineage, etc.), the split depending on the intended use of data. As part of the OA-ICC's activities, the metadata fields from the NOAA template relevant to the sample processing chain and QA criteria have been factored to develop profiles for, and extensions to, the OM-JSON encoding supported by the PROV ontology. While this work started focused on carbonate chemistry variable specific metadata, the factorization could be applied within the O&M model across other disciplines such as trace metals or contaminants. In a linked data world with a suitable high level model for sample processing and QA available, tools and support can be provided to link reproducible units of metadata (e.g. the standard protocol for a variable as adopted by a community) and simplify the provision of metadata and subsequent discovery.
-
Assessing climate change impacts on water resources in remote mountain regions
NASA Astrophysics Data System (ADS)
Buytaert, Wouter; De Bièvre, Bert
2013-04-01
From a water resources perspective, remote mountain regions are often considered as a basket case. They are often regions where poverty is often interlocked with multiple threats to water supply, data scarcity, and high uncertainties. In these environments, it is paramount to generate locally relevant knowledge about water resources and how they impact local livelihoods. This is often problematic. Existing environmental data collection tends to be geographically biased towards more densely populated regions, and prioritized towards strategic economic activities. Data may also be locked behind institutional and technological barriers. These issues create a "knowledge trap" for data-poor regions, which is especially acute in remote and hard-to-reach mountain regions. We present lessons learned from a decade of water resources research in remote mountain regions of the Andes, Africa and South Asia. We review the entire tool chain of assessing climate change impacts on water resources, including the interrogation and downscaling of global circulation models, translating climate variables in water availability and access, and assessing local vulnerability. In global circulation models, mountain regions often stand out as regions of high uncertainties and lack of agreement of future trends. This is partly a technical artifact because of the different resolution and representation of mountain topography, but it also highlights fundamental uncertainties in climate impacts on mountain climate. This problem also affects downscaling efforts, because regional climate models should be run in very high spatial resolution to resolve local gradients, which is computationally very expensive. At the same time statistical downscaling methods may fail to find significant relations between local climate properties and synoptic processes. Further uncertainties are introduced when downscaled climate variables such as precipitation and temperature are to be translated in hydrologically relevant variables such as streamflow and groundwater recharge. Fundamental limitations in both the understanding of hydrological processes in mountain regions (e.g., glacier melt, wetland attenuation, groundwater flows) and in data availability introduce large uncertainties. Lastly, assessing access to water resources is a major challenge. Topographical gradients and barriers, as well as strong spatiotemporal variations in hydrological processes, makes it particularly difficult to assess which parts of the mountain population is most vulnerable to future perturbations of the water cycle.
-
A flow cytometry-based strategy to identify and express IgM from VH1-69+ clonal peripheral B cells.
Charles, Edgar D; Orloff, Michael I M; Dustin, Lynn B
2011-01-05
Pathologic rheumatoid factor (RF) levels are hallmarks of several human diseases. Production of monoclonal RF in vitro is essential for studies of the antigenic specificities of RF, as well as for a dissection of the mechanisms of aberrant RF+ B cell activation. We have expanded upon previous methods to develop a flow cytometry-based method to efficiently clone monoclonal antibodies (mAbs) from humans with expansions of RF-like, immunoglobulin heavy chain variable region (IgVH) 1-69 gene segment-containing B cells. The cloned variable regions are expressed as IgM and produced during culture at concentrations between 5 and 20 μg/ml. Using this system, we show that clonal Igs from patients with HCV-related mixed cryoglobulinemia, when expressed as IgM, have RF activity. We anticipate that this system will be useful for the cloning and expression of mAbs partially encoded by VH1-69 and for determination of the reactivity patterns of polyspecific, low-affinity IgMs of human pathogenic importance. Copyright © 2010 Elsevier B.V. All rights reserved.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Fanning, Sean W.; Horn, James R.
2014-03-05
Conventional anti-hapten antibodies typically bind low-molecular weight compounds (haptens) in the crevice between the variable heavy and light chains. Conversely, heavy chain-only camelid antibodies, which lack a light chain, must rely entirely on a single variable domain to recognize haptens. While several anti-hapten VHHs have been generated, little is known regarding the underlying structural and thermodynamic basis for hapten recognition. Here, an anti-methotrexate VHH (anti-MTX VHH) was generated using grafting methods whereby the three complementarity determining regions (CDRs) were inserted onto an existing VHH framework. Thermodynamic analysis of the anti-MTX VHH CDR1-3 Graft revealed a micromolar binding affinity, while themore » crystal structure of the complex revealed a somewhat surprising noncanonical binding site which involved MTX tunneling under the CDR1 loop. Due to the close proximity of MTX to CDR4, a nonhypervariable loop, the CDR4 loop sequence was subsequently introduced into the CDR1-3 graft, which resulted in a dramatic 1000-fold increase in the binding affinity. Crystal structure analysis of both the free and complex anti-MTX CDR1-4 graft revealed CDR4 plays a significant role in both intermolecular contacts and binding site conformation that appear to contribute toward high affinity binding. Additionally, the anti-MTX VHH possessed relatively high specificity for MTX over closely related compounds aminopterin and folate, demonstrating that VHH domains are capable of binding low-molecular weight ligands with high affinity and specificity, despite their reduced interface.« less
-
Jarvi, S.I.; Tarr, C.L.; Mcintosh, C.E.; Atkinson, C.T.; Fleischer, R.C.
2004-01-01
The native Hawaiian honeycreepers represent a classic example of adaptive radiation and speciation, but currently face one the highest extinction rates in the world. Although multiple factors have likely influenced the fate of Hawaiian birds, the relatively recent introduction of avian malaria is thought to be a major factor limiting honeycreeper distribution and abundance. We have initiated genetic analyses of class II ?? chain Mhc genes in four species of honeycreepers using methods that eliminate the possibility of sequencing mosaic variants formed by cloning heteroduplexed polymerase chain reaction products. Phylogenetic analyses group the honeycreeper Mhc sequences into two distinct clusters. Variation within one cluster is high, with dN > d S and levels of diversity similar to other studies of Mhc (B system) genes in birds. The second cluster is nearly invariant and includes sequences from honeycreepers (Fringillidae), a sparrow (Emberizidae) and a blackbird (Emberizidae). This highly conserved cluster appears reminiscent of the independently segregating Rfp-Y system of genes defined in chickens. The notion that balancing selection operates at the Mhc in the honeycreepers is supported by transpecies polymorphism and strikingly high dN/dS ratios at codons putatively involved in peptide interaction. Mitochondrial DNA control region sequences were invariant in the i'iwi, but were highly variable in the 'amakihi. By contrast, levels of variability of class II ?? chain Mhc sequence codons that are hypothesized to be directly involved in peptide interactions appear comparable between i'iwi and 'amakihi. In the i'iwi, natural selection may have maintained variation within the Mhc, even in the face of what appears to a genetic bottleneck.
-
NASA Technical Reports Server (NTRS)
Ichoku, Charles M.
2010-01-01
The northern sub-Saharan African (NSSA) region, extending from the southern fringes of the Sahara to the Equator, and stretching west to east from the Atlantic to the Indian ocean coasts, plays a prominent role in the distribution of Saharan dust and other airborne matter around the region and to other parts of the world, the genesis of global atmospheric circulation, and the birth of such major (and often catastrophic) events as hurricanes. Therefore, this NSSA region represents a critical variable in the global climate change equation. Recent satellite-based studies have revealed that the NSSA region has one of the highest biomass-burning rates per unit land area among all regions of the world. Because of the high concentration and frequency of fires in this region, with the associated abundance of heat release and gaseous and particulate smoke emissions, biomass-burning activity is believed to be a major driver of the regional carbon, energy, and water cycles. We acknowledge that the rainy season in the NSSA region is from April to September while biomass burning occurs mainly during the dry season (October to March). Nevertheless, these two phenomena are indirectly coupled to each other through a chain of complex processes and conditions, including land-cover and surface-albedo changes, the carbon cycle, evapotranspiration, drought, desertification, surface water runoff, ground water recharge, and variability in atmospheric composition, heating rates, and circulation. In this presentation, we will examine the theoretical linkages between these processes, discuss the preliminary results based on satellite data analysis, and provide an overview of plans for more integrated research to be conducted over the next few years.
-
René, Céline; Prat, Nathalie; Thuizat, Audrey; Broctawik, Mélanie; Avinens, Odile; Eliaou, Jean-François
2014-01-01
Previous studies have suggested a geographical pattern of immunoglobulin rearrangement in chronic lymphocytic leukaemia (CLL), which could be as a result of a genetic background or an environmental antigen. However, the characteristics of Ig rearrangements in the population from the South of France have not yet been established. Here, we studied CLL B-cell repertoire and mutational pattern in a Southern French cohort of patients using an in-house protocol for whole sequencing of the rearranged immunoglobulin heavy-chain genes. Described biased usage of variable, diversity and joining genes between the mutated and unmutated groups was found in our population. However, variable gene frequencies are more in accordance with those observed in the Mediterranean patients. We found that the third complementary-determining region (CDR) length was higher in unmutated sequences, because of bias in the diversity and joining genes usage and not due to the N diversity. Mutations found in CLL followed the features of canonical somatic hypermutation mechanism: preference of targeting for activation-induced cytidine deaminase and polymerase motifs, base change bias for transitions and more replacement mutations occurring in CDRs than in framework regions. Surprisingly, localization of activation-induced cytidine deaminase motifs onto the variable gene showed a preference for framework regions. The study of the characteristics at the age of diagnosis showed no difference in clinical outcome, but suggested a tendency of increased replacement and transition-over-transversion mutations and a longer third CDR length in older patients. PMID:24725733
-
Ordóñez, Adriana; Pérez, Juan; Tan, Lu; Dickens, Jennifer A.; Motamedi-Shad, Neda; Irving, James A.; Haq, Imran; Ekeowa, Ugo; Marciniak, Stefan J.; Miranda, Elena; Lomas, David A.
2015-01-01
Mutant Z α1-antitrypsin (E342K) accumulates as polymers within the endoplasmic reticulum (ER) of hepatocytes predisposing to liver disease, whereas low levels of circulating Z α1-antitrypsin lead to emphysema by loss of inhibition of neutrophil elastase. The ideal therapy should prevent polymer formation while preserving inhibitory activity. Here we used mAb technology to identify interactors with Z α1-antitrypsin that comply with both requirements. We report the generation of an mAb (4B12) that blocked α1-antitrypsin polymerization in vitro at a 1:1 molar ratio, causing a small increase of the stoichiometry of inhibition for neutrophil elastase. A single-chain variable fragment (scFv) intrabody was generated based on the sequence of mAb4B12. The expression of scFv4B12 within the ER (scFv4B12KDEL) and along the secretory pathway (scFv4B12) reduced the intracellular polymerization of Z α1-antitrypsin by 60%. The scFv4B12 intrabody also increased the secretion of Z α1-antitrypsin that retained inhibitory activity against neutrophil elastase. MAb4B12 recognized a discontinuous epitope probably located in the region of helices A/C/G/H/I and seems to act by altering protein dynamics rather than binding preferentially to the native state. This novel approach could reveal new target sites for small-molecule intervention that may block the transition to aberrant polymers without compromising the inhibitory activity of Z α1-antitrypsin.—Ordóñez, A., Pérez, J., Tan, L., Dickens, J. A., Motamedi-Shad, N., Irving, J. A., Haq, I., Ekeowa, U., Marciniak, S. J., Miranda, E., Lomas, D. A. A single-chain variable fragment intrabody prevents intracellular polymerization of Z α1-antitrypsin while allowing its antiproteinase activity. PMID:25757566
-
Atassi, M Zouhair; Jankovic, Joseph; Steward, Lance E; Aoki, K Roger; Dolimbek, Behzod Z
2012-01-01
We recently mapped the regions on the heavy (H) chain of botulinum neurotoxin, type B (BoNT/B) recognized by blocking antibodies (Abs) from cervical dystonia (CD) patients who develop immunoresistance during toxin treatment. Since blocking could also be effected by Abs directed against regions on the light (L) chain, we have mapped here the L chain, using the same 30 CD antisera. We synthesized, purified and characterized 32 19-residue L chain peptides that overlapped successively by 5 residues (peptide L32 overlapped with peptide N1 of the H chain by 12 residues). In a given patient, Abs against the L chain seemed less intense than those against H chain. Most sera recognized a limited set of L chain peptides. The levels of Abs against a given region varied with the patient, consistent with immune responses to each epitope being under separate MHC control. The peptides most frequently recognized were: L13, by 30 of 30 antisera (100%); L22, by 23 of 30 (76.67%); L19, by 15 of 30 (50.00%); L26, by 11 of 30 (36.70%); and L14, by 12 of 30 (40.00%). The activity of L14 probably derives from its overlap with L13. The levels of Ab binding decreased in the following order: L13 (residues 169-187), L22 (295-313), L19 (253-271), and L26 (351-369). Peptides L12 (155-173), L18 (239-257), L15 (197-215), L1 (1-19) and L23 (309-327) exhibited very low Ab binding. The remaining peptides had little or no Ab-binding activity. The antigenic regions are analyzed in terms of their three-dimensional locations and the enzyme active site. With the previous localization of the antigenic regions on the BoNT/B H chain, the human Ab recognition of the entire BoNT/B molecule is presented and compared to the recognition of BoNT/A by human blocking Abs. Copyright © 2011. Published by Elsevier GmbH.
-
Plevova, Karla; Francova, Hana Skuhrova; Burckova, Katerina; Brychtova, Yvona; Doubek, Michael; Pavlova, Sarka; Malcikova, Jitka; Mayer, Jiri; Tichy, Boris; Pospisilova, Sarka
2014-01-01
In chronic lymphocytic leukemia, usually a monoclonal disease, multiple productive immunoglobulin heavy chain gene rearrangements are identified sporadically. Prognostication of such cases based on immunoglobulin heavy variable gene mutational status can be problematic, especially if the different rearrangements have discordant mutational status. To gain insight into the possible biological mechanisms underlying the origin of the multiple rearrangements, we performed a comprehensive immunogenetic and immunophenotypic characterization of 31 cases with the multiple rearrangements identified in a cohort of 1147 patients with chronic lymphocytic leukemia. For the majority of cases (25/31), we provide evidence of the co-existence of at least two B lymphocyte clones with a chronic lymphocytic leukemia phenotype. We also identified clonal drifts in serial samples, likely driven by selection forces. More specifically, higher immunoglobulin variable gene identity to germline and longer complementarity determining region 3 were preferred in persistent or newly appearing clones, a phenomenon more pronounced in patients with stereotyped B-cell receptors. Finally, we report that other factors, such as TP53 gene defects and therapy administration, influence clonal selection. Our findings are relevant to clonal evolution in the context of antigen stimulation and transition of monoclonal B-cell lymphocytosis to chronic lymphocytic leukemia. PMID:24038023
-
Geoghegan, James C.; Fleming, Ryan; Damschroder, Melissa; Bishop, Steven M.; Sathish, Hasige A.; Esfandiary, Reza
2016-01-01
ABSTRACT Undesired solution behaviors such as reversible self-association (RSA), high viscosity, and liquid-liquid phase separation can introduce substantial challenges during development of monoclonal antibody formulations. Although a global mechanistic understanding of RSA (i.e., native and reversible protein-protein interactions) is sufficient to develop robust formulation controls, its mitigation via protein engineering requires knowledge of the sites of protein-protein interactions. In the study reported here, we coupled our previous hydrogen-deuterium exchange mass spectrometry findings with structural modeling and in vitro screening to identify the residues responsible for RSA of a model IgG1 monoclonal antibody (mAb-C), and rationally engineered variants with improved solution properties (i.e., reduced RSA and viscosity). Our data show that mutation of either solvent-exposed aromatic residues within the heavy and light chain variable regions or buried residues within the heavy chain/light chain interface can significantly mitigate RSA and viscosity by reducing the IgG's surface hydrophobicity. The engineering strategy described here highlights the utility of integrating complementary experimental and in silico methods to identify mutations that can improve developability, in particular, high concentration solution properties, of candidate therapeutic antibodies. PMID:27050875
-
A mutagenesis study of a catalytic antibody
DOE Office of Scientific and Technical Information (OSTI.GOV)
Jackson, D.Y.; Prudent, J.R.; Baldwin, E.P.
1991-01-01
The authors have generated seven site-specific mutations in the genes encoding the variable region of the heavy chain domain (V{sub H}) of the phosphocholine-binding antibody S107.S107 is a member of a family of well-characterized highly homologous antibodies that bind phosphorylcholine mono- and diesters. Two of these antibodies, MOPC-167 and T15, have previously been shown to catalyze the hydrolysis of 4-nitrophenyl N-trimethylammonioethyl carbonate. Two conserved heavy-chain residues, Tyr-33 and Arg-52, were postulated to be involved in binding and hydrolysis of 4-nitrophenylcholine carbonate esters. To more precisely define the catalytic roles of these residues, three Arg-52 mutants (R52K, R52Q, R52C) and fourmore » Tyr-33 mutants (Y33H, Y33F, Y33E, Y33D) of antibody S107 were generated. The genes encoding the V{sub H} binding domain of S107 were inserted into plasmid pUC-fl, and in vitro mutagenesis was performed. These results not only demonstrate the importance of electrostatic interactions in catalysis by antibody S107 but also show that catalytic side chains can be introduced into antibodies to enhance their catalytic efficiency.« less
-
Smirnov, Ivan; Carletti, Eugénie; Kurkova, Inna; Nachon, Florian; Nicolet, Yvain; Mitkevich, Vladimir A.; Débat, Hélène; Avalle, Bérangère; Belogurov, Alexey A.; Kuznetsov, Nikita; Reshetnyak, Andrey; Masson, Patrick; Tonevitsky, Alexander G.; Ponomarenko, Natalia; Makarov, Alexander A.; Friboulet, Alain; Tramontano, Alfonso; Gabibov, Alexander
2011-01-01
Igs offer a versatile template for combinatorial and rational design approaches to the de novo creation of catalytically active proteins. We have used a covalent capture selection strategy to identify biocatalysts from within a human semisynthetic antibody variable fragment library that uses a nucleophilic mechanism. Specific phosphonylation at a single tyrosine within the variable light-chain framework was confirmed in a recombinant IgG construct. High-resolution crystallographic structures of unmodified and phosphonylated Fabs display a 15-Å-deep two-chamber cavity at the interface of variable light (VL) and variable heavy (VH) fragments having a nucleophilic tyrosine at the base of the site. The depth and structure of the pocket are atypical of antibodies in general but can be compared qualitatively with the catalytic site of cholinesterases. A structurally disordered heavy chain complementary determining region 3 loop, constituting a wall of the cleft, is stabilized after covalent modification by hydrogen bonding to the phosphonate tropinol moiety. These features and presteady state kinetics analysis indicate that an induced fit mechanism operates in this reaction. Mutations of residues located in this stabilized loop do not interfere with direct contacts to the organophosphate ligand but can interrogate second shell interactions, because the H3 loop has a conformation adjusted for binding. Kinetic and thermodynamic parameters along with computational docking support the active site model, including plasticity and simple catalytic components. Although relatively uncomplicated, this catalytic machinery displays both stereo- and chemical selectivity. The organophosphate pesticide paraoxon is hydrolyzed by covalent catalysis with rate-limiting dephosphorylation. This reactibody is, therefore, a kinetically selected protein template that has enzyme-like catalytic attributes. PMID:21896761
-
Assessing Climate Change Within Lake Champlain
NASA Astrophysics Data System (ADS)
Leibensperger, E. M.; Pierce, W.; Mihuc, T.; Myers, L.
2016-12-01
Lake Champlain is experiencing environmental stresses that have caused statistically significant biological, chemical, and physical trends. Such trends have already impacted management strategies within the Lake Champlain basin, which lies within the states of New York and Vermont and province of Quebec. A long-term monitoring program initiated in 1992 has revealed warming of upwards of 0.7°C per decade within certain regions of the lake; much faster than observed local atmospheric warming. Here we analyze the observed lake warming in the context of atmospheric variability and assess its uncertainty given monitoring frequency (biweekly to monthly), variable seasonal and hourly observation timing, and synoptic variability of lake dynamics. To address these issues, we use observations from a June-October 2016 deployment of a data buoy on Lake Champlain containing a 1-meter spaced thermistor chain and surface weather station. These new observations, and reanalysis of intensive monitoring during a campaign in 1993, indicate that synoptic variability of lake thermal structure lowers confidence in trends derived from infrequent observations. However, principal component analysis of lake thermal structure reveals two primary modes of variability that are predictable from atmospheric conditions, presenting an opportunity to improve interpretation of existing and future observations.
-
Hayhurst, Andrew; Happe, Scott; Mabry, Robert; Koch, Zephyr; Iverson, Brent L; Georgiou, George
2003-05-01
Brucella melitensis is a highly infectious animal pathogen able to cause a recurring debilitating disease in humans and is therefore high on the list of biological warfare agents. Immunoglobulin genes from mice immunized with gamma-irradiated B. melitensis strain 16M were used to construct a library that was screened by phage display against similarly prepared bacteria. The selected phage particles afforded a strong enzyme-linked immunosorbent assay (ELISA) signal against gamma-irradiated B. melitensis cells. However, extensive efforts to express the respective single chain antibody variable region fragment (scFv) in soluble form failed due to: (i) poor solubility and (ii) in vivo degradation of the c-myc tag used for the detection of the recombinant antibodies. Both problems could be addressed by: (i) fusing a human kappa light chain constant domain (Ck) chain to the scFv to generate single chain antibody fragment (scAb) antibody fragments and (ii) by co-expression of the periplasmic chaperone Skp. While soluble, functional antibodies could be produced in this manner, phage-displaying scFvs or scAbs were still found to be superior ELISA reagents for immunoassays, due to the large signal amplification afforded by anti-phage antibodies. The isolated phage antibodies were shown to be highly specific to B. melitensis and did not recognize Yersinia pseudotuberculosis in contrast to the existing diagnostic monoclonal YST 9.2.1.
-
Basic Properties of Strong Mixing Conditions.
1985-06-01
H. Dehling and W. Philipp. Almost sure invariance principles for weakly dependent vector-valued random variables. Ann. Probab. 10 (1982) 689-701. 34...Harris chains will not be discussed here.) It is well known that every stationary Harris chain has a well defined " period " p E {1,2,3,... (the chain is...chain is absolutely regular. (ii) More generally, for any strictly stationary real Harris chain, lim n (o in) = 1 - 1/p where p is the period . A
-
Removing the regional level from the Niger vaccine supply chain.
Assi, Tina-Marie; Brown, Shawn T; Kone, Souleymane; Norman, Bryan A; Djibo, Ali; Connor, Diana L; Wateska, Angela R; Rajgopal, Jayant; Slayton, Rachel B; Lee, Bruce Y
2013-06-10
Since many of the world's vaccine supply chains contain multiple levels, the question remains of whether removing a level could bring efficiencies. We utilized HERMES to generate a detailed discrete-event simulation model of Niger's vaccine supply chain and compared the current four-tier (central, regional, district, and integrated health center levels) with a modified three-tier structure (removing the regional level). Different scenarios explored various accompanying shipping policies and frequencies. Removing the regional level and implementing a collection-based shipping policy from the district stores increases vaccine availability from a mean of 70-100% when districts could collect vaccines at least weekly. Alternatively, implementing a delivery-based shipping policy from the central store monthly in three-route and eight-route scenarios only increases vaccine availability to 87%. Restricting central-to district vaccine shipments to a quarterly schedule for three-route and eight-route scenarios reduces vaccine availability to 49%. The collection-based shipping policy from district stores reduces supply chain logistics cost per dose administered from US$0.14 at baseline to US$0.13 after removing the regional level. Removing the regional level from Niger's vaccine supply chain can substantially improve vaccine availability as long as certain concomitant adjustments to shipping policies and frequencies are implemented. Copyright © 2013 Elsevier Ltd. All rights reserved.
-
Removing the Regional Level from the Niger Vaccine Supply Chain
Assi, Tina-Marie; Brown, Shawn T.; Kone, Souleymane; Norman, Bryan A.; Djibo, Ali; Connor, Diana L.; Wateska, Angela R.; Rajgopal, Jayant; Slayton, Rachel B.; Lee, Bruce Y.
2013-01-01
Objective Since many of the world’s vaccine supply chains contain multiple levels, the question remains of whether removing a level could bring efficiencies. Methods We utilized HERMES to generate a detailed discrete-event simulation model of Niger’s vaccine supply chain and compare the current four-tier (central, regional, district and integrated health center levels) with a modified three-tier structure (removing the regional level). Different scenarios explored various accompanying shipping policies and frequencies. Findings Removing the regional level and implementing a collection-based shipping policy from the district stores increases vaccine availability from a mean of 70% to 100% when districts could collect vaccines at least weekly. Alternatively, implementing a delivery-based shipping policy from the central store monthly in three-route and eight-route scenarios only increases vaccine availability to 87%. Restricting central-to district vaccine shipments to a quarterly schedule for three-route and eight-route scenarios reduces vaccine availability to 49%. The collection-based shipping policy from district stores reduces supply chain logistics cost per dose administered from US$0.14 at baseline to US$0.13 after removing the regional level. Conclusion Removing the regional level from Niger’s vaccine supply chain can substantially improve vaccine availability as long as certain concomitant adjustments to shipping policies and frequencies are implemented. PMID:23602666
-
Ghan, Steven; Wang, Minghuai; Zhang, Shipeng; Ferrachat, Sylvaine; Gettelman, Andrew; Griesfeller, Jan; Kipling, Zak; Lohmann, Ulrike; Morrison, Hugh; Neubauer, David; Partridge, Daniel G; Stier, Philip; Takemura, Toshihiko; Wang, Hailong; Zhang, Kai
2016-05-24
A large number of processes are involved in the chain from emissions of aerosol precursor gases and primary particles to impacts on cloud radiative forcing. Those processes are manifest in a number of relationships that can be expressed as factors dlnX/dlnY driving aerosol effects on cloud radiative forcing. These factors include the relationships between cloud condensation nuclei (CCN) concentration and emissions, droplet number and CCN concentration, cloud fraction and droplet number, cloud optical depth and droplet number, and cloud radiative forcing and cloud optical depth. The relationship between cloud optical depth and droplet number can be further decomposed into the sum of two terms involving the relationship of droplet effective radius and cloud liquid water path with droplet number. These relationships can be constrained using observations of recent spatial and temporal variability of these quantities. However, we are most interested in the radiative forcing since the preindustrial era. Because few relevant measurements are available from that era, relationships from recent variability have been assumed to be applicable to the preindustrial to present-day change. Our analysis of Aerosol Comparisons between Observations and Models (AeroCom) model simulations suggests that estimates of relationships from recent variability are poor constraints on relationships from anthropogenic change for some terms, with even the sign of some relationships differing in many regions. Proxies connecting recent spatial/temporal variability to anthropogenic change, or sustained measurements in regions where emissions have changed, are needed to constrain estimates of anthropogenic aerosol impacts on cloud radiative forcing.
-
Ghan, Steven; Wang, Minghuai; Zhang, Shipeng; Ferrachat, Sylvaine; Gettelman, Andrew; Griesfeller, Jan; Kipling, Zak; Lohmann, Ulrike; Morrison, Hugh; Neubauer, David; Partridge, Daniel G.; Stier, Philip; Takemura, Toshihiko; Wang, Hailong; Zhang, Kai
2016-01-01
A large number of processes are involved in the chain from emissions of aerosol precursor gases and primary particles to impacts on cloud radiative forcing. Those processes are manifest in a number of relationships that can be expressed as factors dlnX/dlnY driving aerosol effects on cloud radiative forcing. These factors include the relationships between cloud condensation nuclei (CCN) concentration and emissions, droplet number and CCN concentration, cloud fraction and droplet number, cloud optical depth and droplet number, and cloud radiative forcing and cloud optical depth. The relationship between cloud optical depth and droplet number can be further decomposed into the sum of two terms involving the relationship of droplet effective radius and cloud liquid water path with droplet number. These relationships can be constrained using observations of recent spatial and temporal variability of these quantities. However, we are most interested in the radiative forcing since the preindustrial era. Because few relevant measurements are available from that era, relationships from recent variability have been assumed to be applicable to the preindustrial to present-day change. Our analysis of Aerosol Comparisons between Observations and Models (AeroCom) model simulations suggests that estimates of relationships from recent variability are poor constraints on relationships from anthropogenic change for some terms, with even the sign of some relationships differing in many regions. Proxies connecting recent spatial/temporal variability to anthropogenic change, or sustained measurements in regions where emissions have changed, are needed to constrain estimates of anthropogenic aerosol impacts on cloud radiative forcing. PMID:26921324
-
Duret, Steven; Guillier, Laurent; Hoang, Hong-Minh; Flick, Denis; Laguerre, Onrawee
2014-06-16
Deterministic models describing heat transfer and microbial growth in the cold chain are widely studied. However, it is difficult to apply them in practice because of several variable parameters in the logistic supply chain (e.g., ambient temperature varying due to season and product residence time in refrigeration equipment), the product's characteristics (e.g., pH and water activity) and the microbial characteristics (e.g., initial microbial load and lag time). This variability can lead to different bacterial growth rates in food products and has to be considered to properly predict the consumer's exposure and identify the key parameters of the cold chain. This study proposes a new approach that combines deterministic (heat transfer) and stochastic (Monte Carlo) modeling to account for the variability in the logistic supply chain and the product's characteristics. The model generates a realistic time-temperature product history , contrary to existing modeling whose describe time-temperature profile Contrary to existing approaches that use directly a time-temperature profile, the proposed model predicts product temperature evolution from the thermostat setting and the ambient temperature. The developed methodology was applied to the cold chain of cooked ham including, the display cabinet, transport by the consumer and the domestic refrigerator, to predict the evolution of state variables, such as the temperature and the growth of Listeria monocytogenes. The impacts of the input factors were calculated and ranked. It was found that the product's time-temperature history and the initial contamination level are the main causes of consumers' exposure. Then, a refined analysis was applied, revealing the importance of consumer behaviors on Listeria monocytogenes exposure. Copyright © 2014. Published by Elsevier B.V.
-
Menezes, C A S; Sullivan, A K; Falta, M T; Mack, D G; Freed, B M; Rocha, M O C; Gollob, K J; Fontenot, A P; Dutra, W O
2012-01-01
Human infection with Trypanosoma cruzi leads to Chagas disease, which presents as several different clinical conditions ranging from an asymptomatic form to a severe dilated cardiomyopathy. Several studies have demonstrated that T cells play a critical role in the development of cardiac pathology, as well as in immunoregulation during chronic disease. However, the mechanisms that drive protective or pathogenic T cell response are not known. We have shown that CD4+ T cells from chagasic patients preferentially express T cell receptor (TCR) β-chain variable region (Vβ) 5. The aim of this work was to determine whether T cells expressing this particular Vβ region displayed variable or restricted CDR3 sequences, as an indicator of the nature of the stimulus leading to the activation of these T cells in vivo. Additionally, we aimed to evaluate phenotypic characteristics of these cells that might be associated with pathology. CDR3 junctional region sequencing of Vβ5·1 expressing CD4+ T cells revealed the occurrence of a highly homologous CDR3 region with conserved TCR Jβ region usage among patients with cardiac, but not indeterminate, Chagas disease. Moreover, correlation analysis indicated that the frequency of CD4+Vβ5·1+ cells is associated with granzyme A expression, suggesting that these cells might display cytotoxic function. Together these results provide new insight into T cell recognition of antigens involved in Chagas disease and suggest that these cells may be implicated in the pathogenesis of chagasic cardiomyopathy. PMID:22774985
-
NASA Astrophysics Data System (ADS)
Cho, Won Sang; Gainer, James S.; Kim, Doojin; Matchev, Konstantin T.; Moortgat, Filip; Pape, Luc; Park, Myeonghun
2014-08-01
We consider a class of on-shell constrained mass variables that are 3+1 dimensional generalizations of the Cambridge M T2 variable and that automatically incorporate various assumptions about the underlying event topology. The presence of additional on-shell constraints causes their kinematic distributions to exhibit sharper endpoints than the usual M T2 distribution. We study the mathematical properties of these new variables, e.g., the uniqueness of the solution selected by the minimization over the invisible particle 4-momenta. We then use this solution to reconstruct the masses of various particles along the decay chain. We propose several tests for validating the assumed event topology in missing energy events from new physics. The tests are able to determine: 1) whether the decays in the event are two-body or three-body, 2) if the decay is two-body, whether the intermediate resonances in the two decay chains are the same, and 3) the exact sequence in which the visible particles are emitted from each decay chain.
-
Clinical and molecular predictors of disease severity and survival in chronic lymphocytic leukemia.
Weinberg, J Brice; Volkheimer, Alicia D; Chen, Youwei; Beasley, Bethany E; Jiang, Ning; Lanasa, Mark C; Friedman, Daphne; Vaccaro, Gina; Rehder, Catherine W; Decastro, Carlos M; Rizzieri, David A; Diehl, Louis F; Gockerman, Jon P; Moore, Joseph O; Goodman, Barbara K; Levesque, Marc C
2007-12-01
Several parameters may predict disease severity and overall survival in chronic lymphocytic leukemia (CLL). The purpose of our study of 190 CLL patients was to compare immunoglobulin heavy chain variable region (IgV(H)) mutation status, cytogenetic abnormalities, and leukemia cell CD38 and Zap-70 to older, traditional parameters. We also wanted to construct a simple, inexpensive prognosis score that would significantly predict TTT and survival in patients at the time of diagnosis and help practicing clinicians. In univariate analyses, patients with higher clinical stage, higher leukocyte count at diagnosis, shorter leukocyte doubling time, elevated serum lactate dehydrogenase (LDH), unmutated immunoglobulin heavy chain variable region (IgV(H)) genes, and higher CD38 had a shorter overall survival and time-to-treatment (TTT). CLL cell Zap-70 expression was higher in patients with unmutated IgV(H), and those with higher Zap-70 tended to have shorter survival. IgV(H)4-34 or IgV(H)1-69 was the most common IgV(H) genes used (16 and 12%, respectively). Of those with IgV(H)1-69, 86% had unmutated IgV(H) and had a significantly shorter TTT. A cytogenetic abnormality was noted in 71% of the patients tested. Patients with 11q22 del and 17p13 del or complex abnormalities were significantly more likely to have unmutated IgV(H). We found that a prognostic score constructed using modified Rai stage, cellular CD38, and serum LDH (parameters easily obtained clinically) significantly predicted TTT and survival in patients at the time of diagnosis and performed as well or better than models using the newer markers.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Proia, R.L.
1988-03-01
Lysosomal {beta}-hexosaminidase is composed of two structurally similar chains, {alpha} and {beta}, that are the products of different genes. Mutations in either gene causing {beta}-hexosaminidase deficiency result in the lysosomal storage disease GM2-gangliosidosis. To enable the investigation of the molecular lesions in this disorder and to study the evolutionary relationship between the {alpha} and {beta} chains, the {beta}-chain gene was isolated, and its organization was characterized. The {beta}-chain coding region is divided into 14 exons distributed over {approx}40 kilobases of DNA. Comparison with the {alpha}-chain gene revealed that 12 of the 13 introns interrupt the coding regions at homologous positions.more » This extensive sharing of intron placement demonstrates that the {alpha} and {beta} chains evolved by way of the duplication of a common ancestor.« less
-
Schmetterer, Klaus G; Haiderer, Daniela; Leb-Reichl, Victoria M; Neunkirchner, Alina; Jahn-Schmid, Beatrice; Küng, Hans J; Schuch, Karina; Steinberger, Peter; Bohle, Barbara; Pickl, Winfried F
2011-01-01
Regulatory T (Treg) cells establish and maintain tolerance to self-antigens and many foreign antigens, such as allergens, by suppressing effector T-cell proliferation and function. We have previously shown that human T-cell receptor (TCR) αβ-chains specific for allergen-derived epitopes confer allergen specificity on peripheral blood T cells of individuals with and without allergy. To study the feasibility of generating allergen-specific human Treg cells by retroviral transduction of a transcription unit encoding forkhead box protein 3 (FOXP3) and allergen-specific TCR αβ-chains. cDNAs encoding the α and β-chains of a Bet v 1(142-153)-specific TCR (TCR alpha variable region 6/TCR beta variable region 20) and human FOXP3 were linked via picornaviral 2A sequences and expressed as single translational unit from an internal ribosomal entry site-green fluorescence protein-containing retroviral vector. Retrovirally transduced peripheral blood T cells were tested for expression of transgenes, Treg phenotype, and regulatory capacity toward allergen-specific effector T cells. Transduced T cells displayed a Treg phenotype with clear-cut upregulation of CD25, CD39, and cytotoxic T-lymphocyte antigen 4. The transduced cells were hyporesponsive in cytokine production and secretion and, like naturally occurring Treg cells, did not proliferate after antigen-specific or antigen-mimetic stimulation. However, proliferation was inducible upon exposure to exogenous IL-2. In coculture experiments, TRAV6(+)TRBV20(+)FOXP3(+) transgenic T cells, unlike FOXP3(+) single transgenic T cells or naturally occurring Treg cells, highly significantly suppressed T cell cytokine production and proliferation of corresponding allergen-specific effector T cells in an allergen-specific, dose-dependent manner. We demonstrate a transgenic approach to engineer human allergen-specific Treg cells that exert their regulatory function in an activation-dependent manner. Customized Treg cells might become useful for tolerance induction therapies in individuals with allergic and other immune-mediated diseases. Copyright © 2010 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.
-
NASA Astrophysics Data System (ADS)
Pickman, Yishai; Dunn-Walters, Deborah; Mehr, Ramit
2013-10-01
Complementarity-determining region 3 (CDR3) is the most hyper-variable region in B cell receptor (BCR) and T cell receptor (TCR) genes, and the most critical structure in antigen recognition and thereby in determining the fates of developing and responding lymphocytes. There are millions of different TCR Vβ chain or BCR heavy chain CDR3 sequences in human blood. Even now, when high-throughput sequencing becomes widely used, CDR3 length distributions (also called spectratypes) are still a much quicker and cheaper method of assessing repertoire diversity. However, distribution complexity and the large amount of information per sample (e.g. 32 distributions of the TCRα chain, and 24 of TCRβ) calls for the use of machine learning tools for full exploration. We have examined the ability of supervised machine learning, which uses computational models to find hidden patterns in predefined biological groups, to analyze CDR3 length distributions from various sources, and distinguish between experimental groups. We found that (a) splenic BCR CDR3 length distributions are characterized by low standard deviations and few local maxima, compared to peripheral blood distributions; (b) healthy elderly people's BCR CDR3 length distributions can be distinguished from those of the young; and (c) a machine learning model based on TCR CDR3 distribution features can detect myelodysplastic syndrome with approximately 93% accuracy. Overall, we demonstrate that using supervised machine learning methods can contribute to our understanding of lymphocyte repertoire diversity.
-
Evidence for the interior evolution of Ceres from geologic analysis of fractures
Scully, Jennifer E. C.; Buczkowski, Debra; Schmedemann, Nico; Raymond, Carol A.; Castillo-Rogez, Julie C.; Scott King,; Bland, Michael T.; Ermakov, Anton; O'Brien, D.P.; Marchi, S.; Longobardo, A.; Russell, C.T.; Fu, R.R.; Neveu, M.
2017-01-01
Ceres is the largest asteroid belt object, and the Dawn spacecraft observed Ceres since 2015. Dawn observed two morphologically distinct linear features on Ceres's surface: secondary crater chains and pit chains. Pit chains provide unique insights into Ceres's interior evolution. We interpret pit chains called the Samhain Catenae as the surface expression of subsurface fractures. Using the pit chains' spacings, we estimate that the localized thickness of Ceres's fractured, outer layer is approximately ≥58 km, at least ~14 km greater than the global average. We hypothesize that extensional stresses, induced by a region of upwelling material arising from convection/diapirism, formed the Samhain Catenae. We derive characteristics for this upwelling material, which can be used as constraints in future interior modeling studies. For example, its predicted location coincides with Hanami Planum, a high-elevation region with a negative residual gravity anomaly, which may be surficial evidence for this proposed region of upwelling material.
-
USDA-ARS?s Scientific Manuscript database
A single-chain variable fragment (scFv) and alkaline phosphatase (AP) fusion protein for detection of O, O-diethyl organophosphorus pesticides (OPs) was produced and characterized. The scFv gene was prepared by cloning VL and VH genes from a hybridoma cell secreting monoclonal antibody with broad-s...
-
Steen Magnussen
2009-01-01
Areas burned annually in 29 Canadian forest fire regions show a patchy and irregular correlation structure that significantly influences the distribution of annual totals for Canada and for groups of regions. A binary Monte Carlo Markov Chain (MCMC) is constructed for the purpose of joint simulation of regional areas burned in forest fires. For each year the MCMC...
-
Exact relativistic Toda chain eigenfunctions from Separation of Variables and gauge theory
NASA Astrophysics Data System (ADS)
Sciarappa, Antonio
2017-10-01
We provide a proposal, motivated by Separation of Variables and gauge theory arguments, for constructing exact solutions to the quantum Baxter equation associated to the N-particle relativistic Toda chain and test our proposal against numerical results. Quantum Mechanical non-perturbative corrections, essential in order to obtain a sensible solution, are taken into account in our gauge theory approach by considering codimension two defects on curved backgrounds (squashed S 5 and degenerate limits) rather than flat space; this setting also naturally incorporates exact quantization conditions and energy spectrum of the relativistic Toda chain as well as its modular dual structure.
-
NASA Astrophysics Data System (ADS)
Kusrini, Elisa; Subagyo; Aini Masruroh, Nur
2016-01-01
This research is a sequel of the author's earlier conducted researches in the fields of designing of integrated performance measurement between supply chain's actors and regulator. In the previous paper, the design of performance measurement is done by combining Balanced Scorecard - Supply Chain Operation Reference - Regulator Contribution model and Data Envelopment Analysis. This model referred as B-S-Rc-DEA model. The combination has the disadvantage that all the performance variables have the same weight. This paper investigates whether by giving weight to performance variables will produce more sensitive performance measurement in detecting performance improvement. Therefore, this paper discusses the development of the model B-S-Rc-DEA by giving weight to its performance'variables. This model referred as Scale B-S-Rc-DEA model. To illustrate the model of development, some samples from small medium enterprises of leather craft industry supply chain in province of Yogyakarta, Indonesia are used in this research. It is found that Scale B-S-Rc-DEA model is more sensitive to detecting performance improvement than B-S- Rc-DEA model.
-
Samad, Manar D; Ulloa, Alvaro; Wehner, Gregory J; Jing, Linyuan; Hartzel, Dustin; Good, Christopher W; Williams, Brent A; Haggerty, Christopher M; Fornwalt, Brandon K
2018-06-09
The goal of this study was to use machine learning to more accurately predict survival after echocardiography. Predicting patient outcomes (e.g., survival) following echocardiography is primarily based on ejection fraction (EF) and comorbidities. However, there may be significant predictive information within additional echocardiography-derived measurements combined with clinical electronic health record data. Mortality was studied in 171,510 unselected patients who underwent 331,317 echocardiograms in a large regional health system. We investigated the predictive performance of nonlinear machine learning models compared with that of linear logistic regression models using 3 different inputs: 1) clinical variables, including 90 cardiovascular-relevant International Classification of Diseases, Tenth Revision, codes, and age, sex, height, weight, heart rate, blood pressures, low-density lipoprotein, high-density lipoprotein, and smoking; 2) clinical variables plus physician-reported EF; and 3) clinical variables and EF, plus 57 additional echocardiographic measurements. Missing data were imputed with a multivariate imputation by using a chained equations algorithm (MICE). We compared models versus each other and baseline clinical scoring systems by using a mean area under the curve (AUC) over 10 cross-validation folds and across 10 survival durations (6 to 60 months). Machine learning models achieved significantly higher prediction accuracy (all AUC >0.82) over common clinical risk scores (AUC = 0.61 to 0.79), with the nonlinear random forest models outperforming logistic regression (p < 0.01). The random forest model including all echocardiographic measurements yielded the highest prediction accuracy (p < 0.01 across all models and survival durations). Only 10 variables were needed to achieve 96% of the maximum prediction accuracy, with 6 of these variables being derived from echocardiography. Tricuspid regurgitation velocity was more predictive of survival than LVEF. In a subset of studies with complete data for the top 10 variables, multivariate imputation by chained equations yielded slightly reduced predictive accuracies (difference in AUC of 0.003) compared with the original data. Machine learning can fully utilize large combinations of disparate input variables to predict survival after echocardiography with superior accuracy. Copyright © 2018 American College of Cardiology Foundation. Published by Elsevier Inc. All rights reserved.
-
A non-affine micro-macro approach to strain-crystallizing rubber-like materials
NASA Astrophysics Data System (ADS)
Rastak, Reza; Linder, Christian
2018-02-01
Crystallization can occur in rubber materials at large strains due to a phenomenon called strain-induced crystallization. We propose a multi-scale polymer network model to capture this process in rubber-like materials. At the microscopic scale, we present a chain formulation by studying the thermodynamic behavior of a polymer chain and its crystallization mechanism inside a stretching polymer network. The chain model accounts for the thermodynamics of crystallization and presents a rate-dependent evolution law for crystallization based on the gradient of the free energy with respect to the crystallinity variables to ensures the dissipation is always non-negative. The multiscale framework allows the anisotropic crystallization of rubber which has been observed experimentally. Two different approaches for formulating the orientational distribution of crystallinity are studied. In the first approach, the algorithm tracks the crystallization at a finite number of orientations. In contrast, the continuous distribution describes the crystallization for all polymer chain orientations and describes its evolution with only a few distribution parameters. To connect the deformation of the micro with that of the macro scale, our model combines the recently developed maximal advance path constraint with the principal of minimum average free energy, resulting in a non-affine deformation model for polymer chains. Various aspects of the proposed model are validated by existing experimental results, including the stress response, crystallinity evolution during loading and unloading, crystallinity distribution, and the rotation of the principal crystallization direction. As a case study, we simulate the formation of crystalline regions around a pre-existing notch in a 3D rubber block and we compare the results with experimental data.
-
Damage, Self-Healing, and Hysteresis in Spider Silks
De Tommasi, D.; Puglisi, G.; Saccomandi, G.
2010-01-01
Abstract In this article, we propose a microstructure-based continuum model to describe the material behavior of spider silks. We suppose that the material is composed of a soft fraction with entropic elasticity and a hard, damageable fraction. The hard fraction models the presence of stiffer, crystal-rich, oriented regions and accounts for the effect of softening induced by the breaking of hydrogen bonds. To describe the observed presence of crystals with different size, composition, and orientation, this hard fraction is modeled as a distribution of materials with variable properties. The soft fraction describes the remaining regions of amorphous material and is here modeled as a wormlike chain. During stretching, we consider the effect of bond-breaking as a transition from the hard- to the soft-material phase. As we demonstrate, a crucial effect of bond-breaking that accompanies the softening of the material is an increase in contour length associated with chains unraveling. The model describes also the self-healing properties of the material by assuming partial bond reconnection upon unloading. Despite its simplicity, the proposed mechanical system reproduces the main experimental effects observed in cyclic loading of spider silks. Moreover, our approach is amenable to two- or three-dimensional extensions and may prove to be a useful tool in the field of microstructure optimization for bioinspired materials. PMID:20441758
-
NASA Astrophysics Data System (ADS)
Ma, Qiyun; Zhang, Jiquan; Sun, Caiyun; Zhang, Feng; Wu, Rina; Wu, Lan
2017-06-01
In this paper, spatiotemporal variability of drought in Xilingol grassland during pasture growing season (from April to September) was investigated, using 52 years (1961-2012) of precipitation data recorded at 14 rain gauge stations in the study area. The Standardized Precipitation Index was used to compute the severity of drought. The Mann-Kendall test, the linear trend, and the sequential Mann-Kendall test were applied to standardized precipitation index (SPI) time series. The results indicate that drought has become increasingly serious on the region scale during pasture growing season, and the rate of SPI decreases ranged from -0.112 to -0.013 per decade. As for the MK test, most of the stations, the Z value range is from -1.081 to -0.005 and Kendall's τ varies from -0.104 to -0.024. Meanwhile, drought is increased obviously from the northwest to the southeast region. Meanwhile, the occurrence probability of each severity class, times for reaching different drought class from any drought severity state, and residence times in each drought class have been obtained with Markov chain. Furthermore, the drought severities during pasture growing season in 2013-2016 are predicted depending on the weighted Markov chain. The results may provide a scientific basis for preventing and mitigating drought disaster.
-
Azuara, Cyril; Lindahl, Erik; Koehl, Patrice; Orland, Henri; Delarue, Marc
2006-07-01
We describe a new way to calculate the electrostatic properties of macromolecules which eliminates the assumption of a constant dielectric value in the solvent region, resulting in a Generalized Poisson-Boltzmann-Langevin equation (GPBLE). We have implemented a web server (http://lorentz.immstr.pasteur.fr/pdb_hydro.php) that both numerically solves this equation and uses the resulting water density profiles to place water molecules at preferred sites of hydration. Surface atoms with high or low hydration preference can be easily displayed using a simple PyMol script, allowing for the tentative prediction of the dimerization interface in homodimeric proteins, or lipid binding regions in membrane proteins. The web site includes options that permit mutations in the sequence as well as reconstruction of missing side chain and/or main chain atoms. These tools are accessible independently from the electrostatics calculation, and can be used for other modeling purposes. We expect this web server to be useful to structural biologists, as the knowledge of solvent density should prove useful to get better fits at low resolution for X-ray diffraction data and to computational biologists, for whom these profiles could improve the calculation of interaction energies in water between ligands and receptors in docking simulations.
-
Verifying the Hanging Chain Model
ERIC Educational Resources Information Center
Karls, Michael A.
2013-01-01
The wave equation with variable tension is a classic partial differential equation that can be used to describe the horizontal displacements of a vertical hanging chain with one end fixed and the other end free to move. Using a web camera and TRACKER software to record displacement data from a vibrating hanging chain, we verify a modified version…
-
Harris, Katherine E; Aldred, Shelley Force; Davison, Laura M; Ogana, Heather Anne N; Boudreau, Andrew; Brüggemann, Marianne; Osborn, Michael; Ma, Biao; Buelow, Benjamin; Clarke, Starlynn C; Dang, Kevin H; Iyer, Suhasini; Jorgensen, Brett; Pham, Duy T; Pratap, Payal P; Rangaswamy, Udaya S; Schellenberger, Ute; van Schooten, Wim C; Ugamraj, Harshad S; Vafa, Omid; Buelow, Roland; Trinklein, Nathan D
2018-01-01
We created a novel transgenic rat that expresses human antibodies comprising a diverse repertoire of heavy chains with a single common rearranged kappa light chain (IgKV3-15-JK1). This fixed light chain animal, called OmniFlic, presents a unique system for human therapeutic antibody discovery and a model to study heavy chain repertoire diversity in the context of a constant light chain. The purpose of this study was to analyze heavy chain variable gene usage, clonotype diversity, and to describe the sequence characteristics of antigen-specific monoclonal antibodies (mAbs) isolated from immunized OmniFlic animals. Using next-generation sequencing antibody repertoire analysis, we measured heavy chain variable gene usage and the diversity of clonotypes present in the lymph node germinal centers of 75 OmniFlic rats immunized with 9 different protein antigens. Furthermore, we expressed 2,560 unique heavy chain sequences sampled from a diverse set of clonotypes as fixed light chain antibody proteins and measured their binding to antigen by ELISA. Finally, we measured patterns and overall levels of somatic hypermutation in the full B-cell repertoire and in the 2,560 mAbs tested for binding. The results demonstrate that OmniFlic animals produce an abundance of antigen-specific antibodies with heavy chain clonotype diversity that is similar to what has been described with unrestricted light chain use in mammals. In addition, we show that sequence-based discovery is a highly effective and efficient way to identify a large number of diverse monoclonal antibodies to a protein target of interest.
-
The actin multigene family and livestock speciation using the polymerase chain reaction.
Fairbrother, K S; Hopwood, A J; Lockley, A K; Bardsley, R G
1998-01-01
Actins constitute a family of highly-conserved multifunctional intracellular proteins, best known as myofibrillar components in striated muscle fibres. Most vertebrate genomes contain numerous actin genes with high sequence homology in protein coding regions but considerable variability in intron number and sizes. This genetic diversity can be utilised for livestock speciation purposes. The high sequence conservation has enabled a single pair of oligonucleotides to be used to prime the polymerase chain reaction (PCR) with DNA extracted from all animals so far studied. Multiple amplification products were obtained which on gel electrophoresis constituted characteristic species-specific 'fingerprints'. The patterns were reproducible, did not vary between individuals of the same breed or between different breeds within a species, and could be generated even from heat-processed muscle held at 120 degrees C for one hour. Given the capacity of PCR to amplify relatively short sequences in highly-degraded DNA, this approach may be suitable for authentication of processed meat products.
-
A soft pillow for hard times? Economic insecurity, food intake and body weight in Russia.
Staudigel, Matthias
2016-12-01
This study investigates causal effects of economic insecurity on subjective anxiety, food intake, and weight outcomes. A review of psychological and nutrition studies highlights the complexity of processes at work on each stage of this causal chain. Econometric analyses trace the effects along the hypothesized pathway using detailed household panel data from the Russia Longitudinal Monitoring Survey from 1994 to 2005. Economic insecurity measures serve as key explanatory variables in regressions and are instrumented by exogenous regional indicators. Results support a causal chain from economic insecurity to weight outcomes for some population subgroups. In contrast to the leading hypothesis that economic insecurity increases body weight, I find strong evidence of a decreasing effect among women. Results suggest further that consumption of foods rich in sugar responds strongly to higher levels of economic insecurity. Heterogeneous impacts of economic insecurity on body weight call for individual-level interventions rather than large-scale action. Copyright © 2016 Elsevier B.V. All rights reserved.
-
Skvortsov, A M; Leermakers, F A M; Fleer, G J
2013-08-07
In the melt polymer conformations are nearly ideal according to Flory's ideality hypothesis. Silberberg generalized this statement for chains in the interfacial region. We check the Silberberg argument by analyzing the conformations of a probe chain end-grafted at a solid surface in a sea of floating free chains of concentration φ by the self-consistent field (SCF) method. Apart from the grafting, probe chain and floating chains are identical. Most of the results were obtained for a standard SCF model with freely jointed chains on a six-choice lattice, where immediate step reversals are allowed. A few data were generated for a five-choice lattice, where such step reversals are forbidden. These coarse-grained models describe the equilibrium properties of flexible atactic polymer chains at the scale of the segment length. The concentration was varied over the whole range from φ = 0 (single grafted chain) to φ = 1 (probe chain in the melt). The number of contacts with the surface, average height of the free end and its dispersion, average loop and train length, tail size distribution, end-point and overall segment distributions were calculated for a grafted probe chain as a function of φ, for several chain lengths and substrate∕polymer interactions, which were varied from strong repulsion to strong adsorption. The computations show that the conformations of the probe chain in the melt do not depend on substrate∕polymer interactions and are very similar to the conformations of a single end-grafted chain under critical conditions, and can thus be described analytically. When the substrate∕polymer interaction is fixed at the value corresponding to critical conditions, all equilibrium properties of a probe chain are independent of φ, over the whole range from a dilute solution to the melt. We believe that the conformations of all flexible chains in the surface region of the melt are close to those of an appropriate single chain in critical conditions, provided that one end of the single chain is fixed at the same point as a chain in the melt.
-
Canadian High Arctic Ionospheric Network (CHAIN)
NASA Astrophysics Data System (ADS)
Jayachandran, P. T.; Langley, R. B.; MacDougall, J. W.; Mushini, S. C.; Pokhotelov, D.; Hamza, A. M.; Mann, I. R.; Milling, D. K.; Kale, Z. C.; Chadwick, R.; Kelly, T.; Danskin, D. W.; Carrano, C. S.
2009-02-01
Polar cap ionospheric measurements are important for the complete understanding of the various processes in the solar wind-magnetosphere-ionosphere system as well as for space weather applications. Currently, the polar cap region is lacking high temporal and spatial resolution ionospheric measurements because of the orbit limitations of space-based measurements and the sparse network providing ground-based measurements. Canada has a unique advantage in remedying this shortcoming because it has the most accessible landmass in the high Arctic regions, and the Canadian High Arctic Ionospheric Network (CHAIN) is designed to take advantage of Canadian geographic vantage points for a better understanding of the Sun-Earth system. CHAIN is a distributed array of ground-based radio instruments in the Canadian high Arctic. The instrument components of CHAIN are 10 high data rate Global Positioning System ionospheric scintillation and total electron content monitors and six Canadian Advanced Digital Ionosondes. Most of these instruments have been sited within the polar cap region except for two GPS reference stations at lower latitudes. This paper briefly overviews the scientific capabilities, instrument components, and deployment status of CHAIN. This paper also reports a GPS signal scintillation episode associated with a magnetospheric impulse event. More details of the CHAIN project and data can be found at http://chain.physics.unb.ca/chain.
-
Structural and affinity studies of IgM polyreactive natural autoantibodies.
Diaw, L; Magnac, C; Pritsch, O; Buckle, M; Alzari, P M; Dighiero, G
1997-01-15
Natural polyreactive autoantibodies (NAA) are an important component of the normal B cell repertoire. One intriguing characteristic of these Abs is their binding to various dissimilar Ags. It has been generally assumed that these Abs bind the Ags with low affinity, and are encoded by germline genes. We have used surface plasmon resonance to determine binding of avidities, and conducted a structural analysis of five murine monoclonal natural autoantibodies displaying a typical polyreactive binding pattern against cytoskeleton Ags and DNA. We show that 1) all the five Abs bind the different Ags with kinetic constants similar to those observed for immune Abs; 2) they express a restricted set of V(H) and V(L) genes, since the same V(H) gene is expressed by three out of the five, and one particular Vkappa gene was expressed twice. In addition, a single D gene segment was used by three of the five Abs; and 3) they express, in most cases, genes in a close germline configuration. Our amino acid sequence and modeling studies show that the distribution of exposed side chains in the NAA paratopes is close to the general pattern observed in the complementarity-determining regions (CDRs) of variable domains from immune Abs. Although CDR3 regions of the heavy chain have been postulated to play a major role in determining polyreactivity on the basis of recombinatorial experiments, our results failed to show any distinctive particularity of this region in terms of length or charge when compared with classical immune Abs.
-
Fera, Daniela; Schmidt, Aaron G; Haynes, Barton F; Gao, Feng; Liao, Hua-Xin; Kepler, Thomas B; Harrison, Stephen C
2014-07-15
Rapidly evolving pathogens, such as human immunodeficiency and influenza viruses, escape immune defenses provided by most vaccine-induced antibodies. Proposed strategies to elicit broadly neutralizing antibodies require a deeper understanding of antibody affinity maturation and evolution of the immune response to vaccination or infection. In HIV-infected individuals, viruses and B cells evolve together, creating a virus-antibody "arms race." Analysis of samples from an individual designated CH505 has illustrated the interplay between an antibody lineage, CH103, and autologous viruses at various time points. The CH103 antibodies, relatively broad in their neutralization spectrum, interact with the CD4 binding site of gp120, with a contact dominated by CDRH3. We show by analyzing structures of progenitor and intermediate antibodies and by correlating them with measurements of binding to various gp120s that there was a shift in the relative orientation of the light- and heavy-chain variable domains during evolution of the CH103 lineage. We further show that mutations leading to this conformational shift probably occurred in response to insertions in variable loop 5 (V5) of the HIV envelope. The shift displaced the tips of the light chain away from contact with V5, making room for the inserted residues, which had allowed escape from neutralization by the progenitor antibody. These results, which document the selective mechanism underlying this example of a virus-antibody arms race, illustrate the functional significance of affinity maturation by mutation outside the complementarity determining region surface of the antibody molecule.
-
Fab-based bispecific antibody formats with robust biophysical properties and biological activity.
Wu, Xiufeng; Sereno, Arlene J; Huang, Flora; Lewis, Steven M; Lieu, Ricky L; Weldon, Caroline; Torres, Carina; Fine, Cody; Batt, Micheal A; Fitchett, Jonathan R; Glasebrook, Andrew L; Kuhlman, Brian; Demarest, Stephen J
2015-01-01
A myriad of innovative bispecific antibody (BsAb) platforms have been reported. Most require significant protein engineering to be viable from a development and manufacturing perspective. Single-chain variable fragments (scFvs) and diabodies that consist only of antibody variable domains have been used as building blocks for making BsAbs for decades. The drawback with Fv-only moieties is that they lack the native-like interactions with CH1/CL domains that make antibody Fab regions stable and soluble. Here, we utilize a redesigned Fab interface to explore 2 novel Fab-based BsAbs platforms. The redesigned Fab interface designs limit heavy and light chain mixing when 2 Fabs are co-expressed simultaneously, thus allowing the use of 2 different Fabs within a BsAb construct without the requirement of one or more scFvs. We describe the stability and activity of a HER2×HER2 IgG-Fab BsAb, and compare its biophysical and activity properties with those of an IgG-scFv that utilizes the variable domains of the same parental antibodies. We also generated an EGFR × CD3 tandem Fab protein with a similar format to a tandem scFv (otherwise known as a bispecific T cell engager or BiTE). We show that the Fab-based BsAbs have superior biophysical properties compared to the scFv-based BsAbs. Additionally, the Fab-based BsAbs do not simply recapitulate the activity of their scFv counterparts, but are shown to possess unique biological activity.
-
Fab-based bispecific antibody formats with robust biophysical properties and biological activity
Wu, Xiufeng; Sereno, Arlene J; Huang, Flora; Lewis, Steven M; Lieu, Ricky L; Weldon, Caroline; Torres, Carina; Fine, Cody; Batt, Micheal A; Fitchett, Jonathan R; Glasebrook, Andrew L; Kuhlman, Brian; Demarest, Stephen J
2015-01-01
A myriad of innovative bispecific antibody (BsAb) platforms have been reported. Most require significant protein engineering to be viable from a development and manufacturing perspective. Single-chain variable fragments (scFvs) and diabodies that consist only of antibody variable domains have been used as building blocks for making BsAbs for decades. The drawback with Fv-only moieties is that they lack the native-like interactions with CH1/CL domains that make antibody Fab regions stable and soluble. Here, we utilize a redesigned Fab interface to explore 2 novel Fab-based BsAbs platforms. The redesigned Fab interface designs limit heavy and light chain mixing when 2 Fabs are co-expressed simultaneously, thus allowing the use of 2 different Fabs within a BsAb construct without the requirement of one or more scFvs. We describe the stability and activity of a HER2×HER2 IgG-Fab BsAb, and compare its biophysical and activity properties with those of an IgG-scFv that utilizes the variable domains of the same parental antibodies. We also generated an EGFR × CD3 tandem Fab protein with a similar format to a tandem scFv (otherwise known as a bispecific T cell engager or BiTE). We show that the Fab-based BsAbs have superior biophysical properties compared to the scFv-based BsAbs. Additionally, the Fab-based BsAbs do not simply recapitulate the activity of their scFv counterparts, but are shown to possess unique biological activity. PMID:25774965
-
Possible Effects of Seasonal Fires on Drought Across the Northern Sub-Saharan African Region
NASA Technical Reports Server (NTRS)
Ichoku, Charles
2010-01-01
Recent satellite-based studies have revealed that the northern sub-Saharan African (NSSA) region has one of the highest biomass-burning rates per unit land area among all regions of the world. Because of the high concentration and frequency of fires in this region, with the associated abundance of heat release and gaseous and particulate smoke emissions, biomass-burning activity is believed to be a major driver of the regional carbon, energy, and water cycles. We acknowledge that the rainy season in the NSSA region is from April to September while biomass burning occurs mainly during the dry season (October to March). Nevertheless, these two phenomena are indirectly coupled to each other through a chain of complex processes and conditions, including land-cover and surface-albedo changes, the carbon cycle, evapotranspiration, drought, desertification, surface water runoff, ground water recharge, and variability in atmospheric composition, heating rates, and circulation. In this presentation, we will examine the theoretical linkages between these processes, discuss the preliminary results based on satellite data analysis, and provide an overview of plans for more integrated research to be conducted over the next few years.
-
Kim, Sangkyu; Park, Insoo; Park, Seung Gu; Cho, Seulki; Kim, Jin Hong; Ipper, Nagesh S.; Choi, Sun Shim; Lee, Eung Suk; Hong, Hyo Jeong
2017-01-01
We constructed a large naïve human Fab library (3 × 1010 colonies) from the lymphocytes of 809 human donors, assessed available diversities of the heavy-chain variable (VH) and κ light-chain variable (VK) domain repertoires, and validated the library by selecting Fabs against 10 therapeutically relevant antigens by phage display. We obtained a database of unique 7,373 VH and 41,804 VK sequences by 454 pyrosequencing, and analyzed the repertoires. The distribution of VH and VK subfamilies and germline genes in our antibody repertoires slightly differed from those in earlier published natural antibody libraries. The frequency of somatic hypermutations (SHMs) in heavy-chain complementarity determining region (HCDR)1 and HCDR2 are higher compared with the natural IgM repertoire. Analysis of position-specific SHMs in CDRs indicates that asparagine, threonine, arginine, aspartate and phenylalanine are the most frequent non-germline residues on the antibody-antigen interface and are converted mostly from the germline residues, which are highly represented in germline SHM hotspots. The amino acid composition and length-dependent changes in amino acid frequencies of HCDR3 are similar to those in previous reports, except that frequencies of aspartate and phenylalanine are a little higher in our repertoire. Taken together, the results show that this antibody library shares common features of natural antibody repertoires and also has unique features. The antibody library will be useful in the generation of human antibodies against diverse antigens, and the information about the diversity of natural antibody repertoires will be valuable in the future design of synthetic human antibody libraries with high functional diversity. PMID:28927259
-
Kim, Sangkyu; Park, Insoo; Park, Seung Gu; Cho, Seulki; Kim, Jin Hong; Ipper, Nagesh S; Choi, Sun Shim; Lee, Eung Suk; Hong, Hyo Jeong
2017-09-30
We constructed a large naïve human Fab library (3 × 10 10 colonies) from the lymphocytes of 809 human donors, assessed available diversities of the heavy-chain variable (VH) and κ light-chain variable (VK) domain repertoires, and validated the library by selecting Fabs against 10 therapeutically relevant antigens by phage display. We obtained a database of unique 7,373 VH and 41,804 VK sequences by 454 pyrosequencing, and analyzed the repertoires. The distribution of VH and VK subfamilies and germline genes in our antibody repertoires slightly differed from those in earlier published natural antibody libraries. The frequency of somatic hypermutations (SHMs) in heavy-chain complementarity determining region (HCDR)1 and HCDR2 are higher compared with the natural IgM repertoire. Analysis of position-specific SHMs in CDRs indicates that asparagine, threonine, arginine, aspartate and phenylalanine are the most frequent non-germline residues on the antibody-antigen interface and are converted mostly from the germline residues, which are highly represented in germline SHM hotspots. The amino acid composition and length-dependent changes in amino acid frequencies of HCDR3 are similar to those in previous reports, except that frequencies of aspartate and phenylalanine are a little higher in our repertoire. Taken together, the results show that this antibody library shares common features of natural antibody repertoires and also has unique features. The antibody library will be useful in the generation of human antibodies against diverse antigens, and the information about the diversity of natural antibody repertoires will be valuable in the future design of synthetic human antibody libraries with high functional diversity.
-
Zhu, Guijie; Zhao, Peng; Deng, Nan; Tao, Dingyin; Sun, Liangliang; Liang, Zhen; Zhang, Lihua; Zhang, Yukui
2012-09-18
Single chain variable fragment (scFv) displaying the M13 phage library was covalently immobilized on magnetic microspheres and used as a protein equalizer for the treatment of human serum. First, scFv displaying M13 phage library functionalized magnetic microspheres (scFv@M13@MM) was incubated with a human serum sample. Second, captured proteins on scFv@M13@MM were eluted with 2 M NaCl, 50 mM glycine-hydrochloric acid (Gly-HCl), and 20% (v/v) acetonitrile with 0.5% (v/v) trifluoroacetic acid in sequence. Finally, the tightly bonded proteins were released by the treatment with thrombin. The eluates were first analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with silver staining. Results indicated that the difference of protein concentration was reduced obviously in NaCl and Gly-HCl fractions compared with untreated human serum sample. The eluates were also digested with trypsin, followed by online 2D-strong cation exchange (SCX)-RPLC-ESI-MS/MS analysis. Results demonstrated that the number of proteins identified from an scFv@M13@MM treated human serum sample was improved 100% compared with that from the untreated sample. In addition, the spectral count of 10 high abundance proteins (serum albumin, serotransferrin, α-2-macroglobulin, α-1-antitrypsin, apolipoprotein B-100, Ig γ-2 chain C region, haptoglobin, hemopexin, α-1-acid glycoprotein 1, and α-2-HS-glycoprotein) decreased evidently after scFv@M13@MM treatment. All these results demonstrate that scFv@M13@MM could efficiently remove high-abundance proteins, reduce the protein concentration difference of human serum, and result in more protein identification.
-
Lavania, Mallika; Jadhav, Rupendra; Turankar, Ravindra P; Singh, Itu; Nigam, Astha; Sengupta, U
2015-12-01
Leprosy is still a major health problem in India which has the highest number of cases. Multiple locus variable number of tandem repeat analysis (MLVA) and single nucleotide polymorphism (SNP) have been proposed as tools of strain typing for tracking the transmission of leprosy. However, empirical data for a defined population from scale and duration were lacking for studying the transmission chain of leprosy. Seventy slit skin scrapings were collected from Purulia (West Bengal), Miraj (Maharashtra), Shahdara (Delhi), and Naini (UP) hospitals of The Leprosy Mission (TLM). SNP subtyping and MLVA on 10 VNTR loci were applied for the strain typing of Mycobacterium leprae. Along with the strain typing conventional epidemiological investigation was also performed to trace the transmission chain. In addition, phylogenetic analysis was done on variable number of tandem repeat (VNTR) data sets using sequence type analysis and recombinational tests (START) software. START software performs analyses to aid in the investigation of bacterial population structure using multilocus sequence data. These analyses include data summary, lineage assignment, and tests for recombination and selection. Diversity was observed in the cross-sectional survey of isolates obtained from 70 patients. Similarity in fingerprinting profiles observed in specimens of cases from the same family or neighborhood locations indicated a possible common source of infection. The data suggest that these VNTRs including subtyping of SNPs can be used to study the sources and transmission chain in leprosy, which could be very important in monitoring of the disease dynamics in high endemic foci. The present study strongly indicates that multi-case families might constitute epidemic foci and the main source of M. leprae in villages, causing the predominant strain or cluster infection leading to the spread of leprosy in the community. Copyright © 2015 Elsevier B.V. All rights reserved.
-
Structural classification of CDR-H3 revisited: a lesson in antibody modeling.
Kuroda, Daisuke; Shirai, Hiroki; Kobori, Masato; Nakamura, Haruki
2008-11-15
Among the six complementarity-determining regions (CDRs) in the variable domains of an antibody, the third CDR of the heavy chain (CDR-H3), which lies in the center of the antigen-binding site, plays a particularly important role in antigen recognition. CDR-H3 shows significant variability in its length, sequence, and structure. Although difficult, model building of this segment is the most critical step in antibody modeling. Since our first proposal of the "H3-rules," which classify CDR-H3 structure based on amino acid sequence, the number of experimentally determined antibody structures has increased. Here, we revise these H3-rules and propose an improved classification scheme for CDR-H3 structure modeling. In addition, we determine the common features of CDR-H3 in antibody drugs as well as discuss the concept of "antibody druggability," which can be applied as an indicator of antibody evaluation during drug discovery.
-
Should we trust build-up/wash-off water quality models at the scale of urban catchments?
Bonhomme, Céline; Petrucci, Guido
2017-01-01
Models of runoff water quality at the scale of an urban catchment usually rely on build-up/wash-off formulations obtained through small-scale experiments. Often, the physical interpretation of the model parameters, valid at the small-scale, is transposed to large-scale applications. Testing different levels of spatial variability, the parameter distributions of a water quality model are obtained in this paper through a Monte Carlo Markov Chain algorithm and analyzed. The simulated variable is the total suspended solid concentration at the outlet of a periurban catchment in the Paris region (2.3 km 2 ), for which high-frequency turbidity measurements are available. This application suggests that build-up/wash-off models applied at the catchment-scale do not maintain their physical meaning, but should be considered as "black-box" models. Copyright © 2016 Elsevier Ltd. All rights reserved.
-
Scheel, Ida; Ferkingstad, Egil; Frigessi, Arnoldo; Haug, Ola; Hinnerichsen, Mikkel; Meze-Hausken, Elisabeth
2013-01-01
Climate change will affect the insurance industry. We develop a Bayesian hierarchical statistical approach to explain and predict insurance losses due to weather events at a local geographic scale. The number of weather-related insurance claims is modelled by combining generalized linear models with spatially smoothed variable selection. Using Gibbs sampling and reversible jump Markov chain Monte Carlo methods, this model is fitted on daily weather and insurance data from each of the 319 municipalities which constitute southern and central Norway for the period 1997–2006. Precise out-of-sample predictions validate the model. Our results show interesting regional patterns in the effect of different weather covariates. In addition to being useful for insurance pricing, our model can be used for short-term predictions based on weather forecasts and for long-term predictions based on downscaled climate models. PMID:23396890
-
NASA Astrophysics Data System (ADS)
Contreras, S.; Werne, J. P.; Araneda, A.; Conejero, C. A.
2015-12-01
Sedimentary carbon isotope values (δ13C) of bulk organic matter and long chain (C25 to C35) n-alkanes are among the most long-lived and widely utilized proxies of organic matter and vegetation source. The carbon distribution (e.g. average carbon chain length, ACL) and isotope signature from long chain n-alkanes had been intensively used on paleoclimate studies because they are less influenced by diagenesis, differential preservation of compound classes, and changes in the sources of organic matter than bulk δ13C values. Recently, studies of modern plant n-alkanes have challenged the use of carbon distribution and carbon isotope signature from sedimentary n-alkanes as reliable indicators of vegetation and climate change. The climate in central-south western South America (SA) is projected to become significantly warmer and drier over the next several decades to centuries in response to anthropogenically driven warming. Paleolimnological studies along western SA are critical to obtain more realistic and reliable regional reconstructions of past climate and environments, including vegetation and water budget variability. Here we discuss bulk δ13C, distribution and δ13C in long chain n-alkanes from a suite of ~40 lake surface sediment (core-top) samples spanning the transition from a Mediterranean climate with a patchwork of cultivated vegetation, pastureland, conifers in central Chile to a rainy temperate climate dominated by broadleaf deciduous and evergreen forest. Data are compared to the latitudinal and orographic climatic trends of the Andes based on the climatology (e.g. precipitation and temperature) of the locations of all lakes involved in this study, using monthly gridded reanalysis products of the Climate Forecast System Reanalysis (CFSR), based on the NCEP global forecast model and meteorological stations available in the region, from January 1979 to December 2010 with a 0.5° horizontal resolution.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Dong, Jianbo; Thompson, Aaron A.; Fan, Yongfeng
2010-08-13
Ingestion or inhalation of botulinum neurotoxin (BoNT) results in botulism, a severe and frequently fatal disease. Current treatments rely on antitoxins, which, while effective, cannot reverse symptoms once BoNT has entered the neuron. For treatments that can reverse intoxication, interest has focused on developing inhibitors of the enzymatic BoNT light chain (BoNT Lc). Such inhibitors typically mimic substrate and bind in or around the substrate cleavage pocket. To explore the full range of binding sites for serotype A light chain (BoNT/A Lc) inhibitors, we created a library of non-immune llama single-domain VHH (camelid heavy-chain variable region derived from heavy-chain-only antibody)more » antibodies displayed on the surface of the yeast Saccharomyces cerevisiae. Library selection on BoNT/A Lc yielded 15 yeast-displayed VHH with equilibrium dissociation constants (K{sub d}) from 230 to 0.03 nM measured by flow cytometry. Eight of 15 VHH inhibited the cleavage of substrate SNAP25 (synaptosome-associated protein of 25,000 Da) by BoNT/A Lc. The most potent VHH (Aa1) had a solution K{sub d} for BoNT/A Lc of 1.47 x 10{sup -10} M and an IC{sub 50} (50% inhibitory concentration) of 4.7 x 10{sup -10} M and was resistant to heat denaturation and reducing conditions. To understand the mechanism by which Aa1 inhibited catalysis, we solved the X-ray crystal structure of the BoNT/A Lc-Aa1 VHH complex at 2.6 {angstrom} resolution. The structure reveals that the Aa1 VHH binds in the {alpha}-exosite of the BoNT/A Lc, far from the active site for catalysis. The study validates the utility of non-immune llama VHH libraries as a source of enzyme inhibitors and identifies the BoNT/A Lc {alpha}-exosite as a target for inhibitor development.« less
-
... major national fast-food chains and casual-dining restaurant chains have announced they will no longer use ... deep-fry foods. Many smaller local and regional restaurant chains have made similar announcements. In late 2013, ...
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Wall, J.S.; Gupta, V.; Wilkerson, M.
2004-04-01
Primary (AL) amyloidosis results from the pathologic deposition of monoclonal light chains as amyloid fibrils. Studies of recombinant-derived variable region (V{sub L}) fragments of these proteins have shown an inverse relationship between thermodynamic stability and fibrillogenic potential. Further, ionic interactions within the V{sub L} domain were predicted to influence the kinetics of light chain fibrillogenicity, as evidenced from our analyses of a relatively stable V{sub {lambda}}6 protein (Jto) with a long range electrostatic interaction between Asp and Arg side chains at position 29 and 68, respectively, and an unstable, highly fibrillogenic V{sub {lambda}}6 protein (Wil) that had neutral amino acidsmore » at these locations. To test this hypothesis, we have generated two Jto-related mutants designed to disrupt the interaction between Asp 29 and Arg 68 (JtoD29A and JtoR68S). Although the thermodynamic stabilities of unfolding for these two molecules were identical, they exhibited very different kinetics of fibril formation: the rate of JtoD29A fibrillogenesis was slow and comparable to the parent molecule, whereas that of JtoR68S was significantly faster. High-resolution X-ray diffraction analyses of crystals prepared from the two mutants having the same space group and unit cell dimensions revealed no significant main-chain conformational changes. However, several notable side-chain alterations were observed in JtoR68S, as compared with JtoD29A, that resulted in the solvent exposure of a greater hydrophobic surface and modifications in the electrostatic potential surface. We posit that these differences contributed to the enhanced fibrillogenic potential of the Arg 68 mutant, since both Jto mutants lacked the intrachain ionic interaction and were equivalently unstable. The information gleaned from our studies has provided insight into structural parameters that in addition to overall thermodynamic stability, contribute to the fibril forming propensity of immunoglobulin light chains.« less
-
Immunoglobulin from Antarctic fish species of Rajidae family.
Coscia, Maria Rosaria; Cocca, Ennio; Giacomelli, Stefano; Cuccaro, Fausta; Oreste, Umberto
2012-03-01
Immunoglobulins (Ig) of Chondroichthyes have been extensively studied in sharks; in contrast, in skates investigations on Ig remain scarce and fragmentary despite the high occurrence of skates in all of the major oceans of the world. To focus on Rajidae Igμ, the most abundant heavy chain isotype, we have chosen the Antarctic species Bathyraja eatonii, Bathyraja albomaculata, Bathyraja brachyurops, and Amblyraja georgiana which live at high latitudes in the Southern Ocean, and at very low temperatures. We prepared mRNA from the spleen of individuals of each species and performed RT-PCR experiments using two oligonucleotides designed on the alignment of various elasmobranch Igμ heavy chain sequences available in GenBank. The PCR products, about 1400-nt long, were cloned and sequenced. Nucleotide sequence identities calculated for the constant region domains ranged from 88.5% to 97.5% between species, and from 91.1% to 99.7% within species. In a distance tree, including also Raja erinacea sequences, two major branches were obtained, one containing Arhynchobatinae sequences, the other one Rajinae sequences. Four presumptive D gene segments were identified in the region of the VH/D/JH recombination; two different D segments were often found in the same sequence. Moreover, 5-15 genomic fragments of different lengths, carrying the gene locus encoding Igμ chain were revealed by Southern blotting analysis. B. eatonii amino acid sequences were analyzed for the positional diversity by Shannon entropy analysis, showing CH4 as the most conserved domain, and CH3 as the most variable one. B. eatonii CDR3 region length varied between 11 and 15 amino acid residues; the mean length (13.4 aa) was greater than that of Leucoraja eglanteria sequences (7.7 aa). An alignment of representative sequences of Antarctic species and R. erinacea showed that more cysteine residues not involved in the intradomain disulfide bridges were present in Antarctic species. Copyright © 2011 Elsevier B.V. All rights reserved.
-
Forte, Esther; Llovell, Felix; Vega, Lourdes F; Trusler, J P Martin; Galindo, Amparo
2011-04-21
An accurate prediction of phase behavior at conditions far and close to criticality cannot be accomplished by mean-field based theories that do not incorporate long-range density fluctuations. A treatment based on renormalization-group (RG) theory as developed by White and co-workers has proven to be very successful in improving the predictions of the critical region with different equations of state. The basis of the method is an iterative procedure to account for contributions to the free energy of density fluctuations of increasing wavelengths. The RG method has been combined with a number of versions of the statistical associating fluid theory (SAFT), by implementing White's earliest ideas with the improvements of Prausnitz and co-workers. Typically, this treatment involves two adjustable parameters: a cutoff wavelength L for density fluctuations and an average gradient of the wavelet function Φ. In this work, the SAFT-VR (variable range) equation of state is extended with a similar crossover treatment which, however, follows closely the most recent improvements introduced by White. The interpretation of White's latter developments allows us to establish a straightforward method which enables Φ to be evaluated; only the cutoff wavelength L then needs to be adjusted. The approach used here begins with an initial free energy incorporating only contributions from short-wavelength fluctuations, which are treated locally. The contribution from long-wavelength fluctuations is incorporated through an iterative procedure based on attractive interactions which incorporate the structure of the fluid following the ideas of perturbation theories and using a mapping that allows integration of the radial distribution function. Good agreement close and far from the critical region is obtained using a unique fitted parameter L that can be easily related to the range of the potential. In this way the thermodynamic properties of a square-well (SW) fluid are given by the same number of independent intermolecular model parameters as in the classical equation. Far from the critical region the approach provides the correct limiting behavior reducing to the classical equation (SAFT-VR). In the critical region the β critical exponent is calculated and is found to take values close to the universal value. In SAFT-VR the free energy of an associating chain fluid is obtained following the thermodynamic perturbation theory of Wertheim from the knowledge of the free energy and radial distribution function of a reference monomer fluid. By determining L for SW fluids of varying well width a unique equation of state is obtained for chain and associating systems without further adjustment of critical parameters. We use computer simulation data of the phase behavior of chain and associating SW fluids to test the accuracy of the new equation.
-
The complex influence of ENSO on droughts in Ecuador
NASA Astrophysics Data System (ADS)
Vicente-Serrano, S. M.; Aguilar, E.; Martínez, R.; Martín-Hernández, N.; Azorin-Molina, C.; Sanchez-Lorenzo, A.; El Kenawy, A.; Tomás-Burguera, M.; Moran-Tejeda, E.; López-Moreno, J. I.; Revuelto, J.; Beguería, S.; Nieto, J. J.; Drumond, A.; Gimeno, L.; Nieto, R.
2017-01-01
In this study, we analyzed the influence of El Niño-Southern Oscillation (ENSO) on the spatio-temporal variability of droughts in Ecuador for a 48-year period (1965-2012). Droughts were quantified from 22 high-quality and homogenized time series of precipitation and air temperature by means of the Standardized Precipitation Evapotranspiration Index. In addition, the propagation of two different ENSO indices (El Niño 3.4 and El Niño 1 + 2 indices) and other atmospheric circulation processes (e.g., vertical velocity) on different time-scales of drought severity were investigated. The results showed a very complex influence of ENSO on drought behavior across Ecuador, with two regional patterns in the evolution of droughts: (1) the Andean chain with no changes in drought severity, and (2) the Western plains with less severe and frequent droughts. We also detected that drought variability in the Andes mountains is explained by the El Niño 3.4 index [sea surface temperature (SST) anomalies in the central Pacific], whereas the Western plains are much more driven by El Niño 1 + 2 index (SST anomalies in the eastern Pacific). Moreover, it was also observed that El Niño and La Niña phases enhance droughts in the Andes and Western plains regions, respectively. The results of this work could be crucial for predicting and monitoring drought variability and intensity in Ecuador.
-
Lundqvist, M L; Middleton, D L; Hazard, S; Warr, G W
2001-12-14
The region of the duck IgH locus extending from upstream of the proximal diversity (D) segment to downstream of the constant gene cluster has been cloned and mapped. A sequence contig of 48,796 base pairs established that the organization of the genes is D-J(H)-mu-alpha-upsilon. No evidence for a functional homologue (or remnant) of a delta gene was found. The alpha gene is in inverted transcriptional orientation; class switch to IgA expression thus requires inversion of the approximately 27-kilobase pair region that includes both mu and alpha genes. The secreted forms of duck alpha and mu are each encoded by 4 constant region exons, and the hydrophobic C-terminal regions of the membrane receptor forms of alpha and mu are encoded by one and two transmembrane exons, respectively. Putative switch (S) regions were identified for duck mu and upsilon by comparison with chicken Smu and Supsilon sequences and for duck alpha by comparison with mouse Salpha. The duck IgH locus is rich in complex variable number tandem repeats, which occupy approximately 60% of the sequenced region, and occur at a much higher frequency in the IgH locus than in other sequenced regions of the duck genome.
-
[A monitor for the emergency psychiatric service chain].
Salden, M E F H; van Kemenade, J F L M; van Dam, A; Mulder, C L
2014-01-01
Many organisations are involved in the delivery of psychiatric emergency care to patients in crisis. We therefore refer to these organisations as the Psychiatric Emergency Chain (PEC). The quality of the PEC determines the quality of the psychiatric emergency care in a region. In order to measure the quality of this type of care a monitor has been developed in collaboration with the main stakeholders in the region Western North Brabant (WNB). To develop a monitor to measure the quality of a PEC in a region, identify weak spots in the chain and feed back the results to the chain. By searching the literature, studying the primary process in the chain and by having interviews with stakeholders we compiled a questionnaire for monitoring the pec. The monitor was tested in the WNB region and adjusted on the basis of experiences with the respondents. Then the monitor was generalised and tested in another region (Eindhoven and the Kempen) in order to find out whether this instrument might also be applicable to other pecs in the Netherlands. The monitor provided the PEC with a common language for evaluating the chain. The monitor was able to pinpoint particular weak spots and collaboration problems such as the response time required to reach the emergency, communication between and within organisations and domain discussions between different stakeholders. Chain partners used the results of the monitor to negotiate better collaborative agreements and to improve their care. The monitor is able to pinpoint problem areas in the PEC and subsequently to generate feedback to its stakeholders. This creates opportunities for improvement in the PEC. Therefore the monitor is a useful instrument for evaluating a local PEC periodically.
-
Quiet Time Depression of the Equatorial Electrojet and Dynamics of the F-layer Ionosphere
NASA Astrophysics Data System (ADS)
Khadka, S.; Valladares, C. E.; Doherty, P.
2017-12-01
The depression of the equatorial electrojet (EEJ) is marked by a westward current due to streaming movement of laterally limited (±3°) charged particles in the ionospheric E region during the day along the magnetic equator. It is a complex low-latitude phenomenon and driven by various sources of electric fields associated with global neutral wind, solar tidal force, Interplanetary magnetic Field (IMF), etc. This unique physical property of the equatorial ionosphere holds a great promise for sorting out the governing mechanism of the dayside ionospheric electrodynamics and the onset of the enigmatic plasma structures in the ionospheric layers. Present study provides an overview of the special sequence of the longitudinal, seasonal, and occurrence rate variability of the depression of the EEJ, including its temporal variation, using data from an excellent chain of magnetic and ionospheric observatories along the low-latitude regions. A case and statistical study of the geomagnetically quiet time depression of EEJ strengths is presented using a pair of magnetometers, one located at the dip equator and another off the dip equator (±6° to ±9° away) in the American low-latitude regions. The significance of the variability of the depression of the EEJ current observed in the scenario of vertical drifts, sporadic E-layer, the equatorial F region plasma fountain, and height of the peak ionization in the F-layer, as well as GPS-TEC distributions, will be investigated.
-
Raue, R; Hess, M
1998-08-01
Three different polymerase chain reactions (PCRs), two of them combined with restriction enzyme analysis (REA), were developed for detection and differentiation of all 12 fowl adenovirus (FAV) serotypes and the egg drop syndrome (EDS) virus. For primer construction FAV1, FAV10 and EDS virus hexon proteins were aligned and conserved and variable regions were determined. Two primer sets (H1/H2 and H3/H4) for single use were constructed which hybridize in three conserved regions of hexon genes. Each primer pair amplifies approximately half of the hexon gene including two loop regions. An amplification product was detected with both primer sets using purified DNA from all FAV1-12 reference strains. Viral EDS DNA was negative using the H1/H2 or H3/H4 primer pair. HaeII digestion of the H1/H2 amplification products differentiates between all viruses except FAV4 and FAV5. In comparison, much more clustering among genomic closely related FAV serotypes was seen after HpaII digestion of the H3/H4 PCR products. Oligonucleotides H5/H6 located in the variable regions of EDS virus hexon gene do not detect any of the FAV serotypes. The PCRs and REA described are suitable to detect all avian adenoviruses infecting chickens, to distinguish all 12 FAV reference strains and to differentiate FAVs from the EDS virus.
-
Nursing home financial performance: the role of ownership and chain affiliation.
Weech-Maldonado, Robert; Laberge, Alex; Pradhan, Rohit; Johnson, Christopher E; Yang, Zhou; Hyer, Kathryn
2012-01-01
The nursing home industry serves one of the most vulnerable populations, and its financial sustainability is a matter of public concern. However, limited empirical evidence exists on the impact of ownership and chain affiliation on nursing home financial performance. The aim of this study was to examine the joint effects of ownership and chain affiliation on the financial performance of the nursing home industry for the study period 1999-2004 on a national sample of 11,236 nursing homes per year. Data included the Medicare Cost Reports; the Online Survey, Certification, and Reporting file; and the Area Resource File. Dependent variables included operating and total margins. Independent variables included four ownership/chain affiliation combinations: for-profit chain, for-profit independent, not-for-profit chain, and not-for-profit independent. Random effects generalized least square regressions were performed. Results show that for-profit nursing homes delivered better financial performance than not-for-profit facilities did across both operating and total margins. However, the relationship between chain affiliation and financial performance was more nuanced. In the case of operating margin, chain-affiliated facilities delivered superior financial performance irrespective of ownership type; however, in the case of total margin, independents outperformed chain-affiliated facilities among for-profits. Our findings show an interactive effect of ownership and chain affiliation on nursing home financial performance, suggesting the pursuit of different organizational strategies by different ownership/chain affiliation subgroups (for-profit chain, for-profit independent, not-for-profit chain, and not-for-profit independent), with implications for financial performance. For-profit independent nursing homes managed to be the top performing group in terms of overall financial despite the operating financial advantage of for-profit chain-affiliated nursing homes. Similarly, not-for-profit independent nursing homes and not-for-profit chain homes had comparable overall financial performance despite the operating financial advantage of chain homes.
-
An Integer Programming Model for Multi-Echelon Supply Chain Decision Problem Considering Inventories
NASA Astrophysics Data System (ADS)
Harahap, Amin; Mawengkang, Herman; Siswadi; Effendi, Syahril
2018-01-01
In this paper we address a problem that is of significance to the industry, namely the optimal decision of a multi-echelon supply chain and the associated inventory systems. By using the guaranteed service approach to model the multi-echelon inventory system, we develop a mixed integer; programming model to simultaneously optimize the transportation, inventory and network structure of a multi-echelon supply chain. To solve the model we develop a direct search approach using a strategy of releasing nonbasic variables from their bounds, combined with the “active constraint” method. This strategy is used to force the appropriate non-integer basic variables to move to their neighbourhood integer points.
-
New construction of eigenstates and separation of variables for SU( N) quantum spin chains
NASA Astrophysics Data System (ADS)
Gromov, Nikolay; Levkovich-Maslyuk, Fedor; Sizov, Grigory
2017-09-01
We conjecture a new way to construct eigenstates of integrable XXX quantum spin chains with SU( N) symmetry. The states are built by repeatedly acting on the vacuum with a single operator B good( u) evaluated at the Bethe roots. Our proposal serves as a compact alternative to the usual nested algebraic Bethe ansatz. Furthermore, the roots of this operator give the separated variables of the model, explicitly generalizing Sklyanin's approach to the SU( N) case. We present many tests of the conjecture and prove it in several special cases. We focus on rational spin chains with fundamental representation at each site, but expect many of the results to be valid more generally.
-
Fabrication of ZnS nanoparticle chains on a protein template
Hulleman, J.; Kim, S. M.; Tumkur, T.; Rochet, J.-C.; Stach, E.; Stanciu, L.
2011-01-01
In the present study, we have exploited the properties of a fibrillar protein for the template synthesis of zinc sulfide (ZnS) nanoparticle chains. The diameter of the ZnS nanoparticle chains was tuned in range of ~30 to ~165 nm by varying the process variables. The nanoparticle chains were characterized by field emission scanning electron microscopy, UV–Visible spectroscopy, transmission electron microscopy, electron energy loss spectroscopy, and high-resolution transmission electron microscopy. The effect of incubation temperature on the morphology of the nanoparticle chains was also studied. PMID:21804765
-
Nanoparticle stability in semidilute and concentrated polymer solutions.
Dutta, Nupur; Green, David
2008-05-20
The wetting of PDMS-grafted silica spheres (PDMS- g-silica) is connected to their depletion restabilization in semidilute and concentrated PDMS/cyohexane polymer solutions. Specifically, we found that a wetting diagram of chemically identical graft and free homopolymers predicts stability of hard, semisoft, and soft spheres as a function of the bulk free polymer volume fraction, graft density, and the graft and free polymer chain lengths. The transition between stable and aggregated regions is determined optically and with dynamic light scattering. The point of demarcation between the regions occurs when the graft and free polymer chains are equal in length. When graft chains are longer than free chains, the particles are stable; in contrast, the particles are unstable when the opposite is true. The regions of particle stability and instability are corroborated with theoretical self-consistent mean-field calculations, which not only show that the grafted brush is responsible for particle dispersion in the complete wetting region but also aggregation in the incomplete wetting region. Ultimately, our results indicate that depletion restabilization depends on the interfacial properties of the nanoparticles in semidilute and concentrated polymer solutions.
-
Atassi, M Zouhair; Dolimbek, Behzod Z; Jankovic, Joseph; Steward, Lance E; Aoki, K Roger
2011-07-01
This work was aimed at determining the BoNT/A L-chain antigenic regions recognized by blocking antibodies in human antisera from cervical dystonia patients who had become immunoresistant to BoNT/A treatment. Antisera from 28 immunoresistant patients were analyzed for binding to each of 32 overlapping synthetic peptides that spanned the entire L-chain. A mixture of the antisera showed that antibodies bound to three peptides, L11 (residues 141-159), L14 (183-201) and L18 (239-257). When mapped separately, the antibodies were bound only by a limited set of peptides. No peptide bound antibodies from all the patients and amounts of antibodies bound to a given peptide varied with the patient. Peptides L11, L14 and L18 were recognized predominantly. A small but significant number of patients had antibodies to peptides L27 (365-383) and L29 (379-397). Other peptides were recognized at very low and perhaps insignificant antibody levels by a minority (15% or less) of patients or had no detectable antibody with any of the sera. In the 3-dimensional structure, antibody-binding regions L11, L14 and L18 of the L-chain occupy surface areas and did not correlate with electrostatic potential, hydrophilicity/hydrophobicity, or temperature factor. These three antigenic regions reside in close proximity to the belt of the heavy chain. The regions L11 and L18 are accessible in both the free light chain and the holotoxin forms, while L14 appears to be less accessible in the holotoxin. Antibodies against these regions could prevent delivery of the L-chain into the neurons by inhibition of the translocation. Copyright © 2011 Elsevier GmbH. All rights reserved.
-
Neelly, Kurt R; Terry, Joseph G; Morris, Martin J
2010-01-01
A relatively new and scarcely researched technique to increase strength is the use of supplemental heavy chain resistance (SHCR) in conjunction with plate weights to provide variable resistance to free weight exercises. The purpose of this case study was to determine the actual resistance being provided by a double-looped versus a linear hung SHCR to the back squat exercise. The linear technique simply hangs the chain directly from the bar, whereas the double-looped technique uses a smaller chain to adjust the height of the looped chain. In both techniques, as the squat descends, chain weight is unloaded onto the floor, and as the squat ascends, chain weight is progressively loaded back as resistance. One experienced and trained male weight lifter (age = 33 yr; height = 1.83 m; weight = 111.4 kg) served as the subject. Plate weight was set at 84.1 kg, approximately 50% of the subject's 1 repetition maximum. The SHCR was affixed to load cells, sampling at a frequency of 500 Hz, which were affixed to the Olympic bar. Data were collected as the subject completed the back squat under the following conditions: double-looped 1 chain (9.6 kg), double-looped 2 chains (19.2 kg), linear 1 chain, and linear 2 chains. The double-looped SHCR resulted in a 78-89% unloading of the chain weight at the bottom of the squat, whereas the linear hanging SHCR resulted in only a 36-42% unloading. The double-looped technique provided nearly 2 times the variable resistance at the top of the squat compared with the linear hanging technique, showing that attention must be given to the technique used to hang SHCR.
-
A residue-specific shift in stability and amyloidogenicity of antibody variable domains.
Nokwe, Cardine N; Zacharias, Martin; Yagi, Hisashi; Hora, Manuel; Reif, Bernd; Goto, Yuji; Buchner, Johannes
2014-09-26
Variable (V) domains of antibodies are essential for antigen recognition by our adaptive immune system. However, some variants of the light chain V domains (VL) form pathogenic amyloid fibrils in patients. It is so far unclear which residues play a key role in governing these processes. Here, we show that the conserved residue 2 of VL domains is crucial for controlling its thermodynamic stability and fibril formation. Hydrophobic side chains at position 2 stabilize the domain, whereas charged residues destabilize and lead to amyloid fibril formation. NMR experiments identified several segments within the core of the VL domain to be affected by changes in residue 2. Furthermore, molecular dynamic simulations showed that hydrophobic side chains at position 2 remain buried in a hydrophobic pocket, and charged side chains show a high flexibility. This results in a predicted difference in the dissociation free energy of ∼10 kJ mol(-1), which is in excellent agreement with our experimental values. Interestingly, this switch point is found only in VL domains of the κ family and not in VLλ or in VH domains, despite a highly similar domain architecture. Our results reveal novel insight into the architecture of variable domains and the prerequisites for formation of amyloid fibrils. This might also contribute to the rational design of stable variable antibody domains. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.
-
Structural re-alignment in an immunologic surface region of ricin A chain
DOE Office of Scientific and Technical Information (OSTI.GOV)
Zemla, A T; Zhou, C E
2007-07-24
We compared structure alignments generated by several protein structure comparison programs to determine whether existing methods would satisfactorily align residues at a highly conserved position within an immunogenic loop in ribosome inactivating proteins (RIPs). Using default settings, structure alignments generated by several programs (CE, DaliLite, FATCAT, LGA, MAMMOTH, MATRAS, SHEBA, SSM) failed to align the respective conserved residues, although LGA reported correct residue-residue (R-R) correspondences when the beta-carbon (Cb) position was used as the point of reference in the alignment calculations. Further tests using variable points of reference indicated that points distal from the beta carbon along a vector connectingmore » the alpha and beta carbons yielded rigid structural alignments in which residues known to be highly conserved in RIPs were reported as corresponding residues in structural comparisons between ricin A chain, abrin-A, and other RIPs. Results suggest that approaches to structure alignment employing alternate point representations corresponding to side chain position may yield structure alignments that are more consistent with observed conservation of functional surface residues than do standard alignment programs, which apply uniform criteria for alignment (i.e., alpha carbon (Ca) as point of reference) along the entirety of the peptide chain. We present the results of tests that suggest the utility of allowing user-specified points of reference in generating alternate structural alignments, and we present a web server for automatically generating such alignments.« less
-
Booker, Anthony; Frommenwiler, Debora; Johnston, Deborah; Umealajekwu, Chinenye; Reich, Eike; Heinrich, Michael
2014-03-14
Herbal medicine value chains have generally been overlooked compared with food commodities. Not surprisingly, revenue generation tends to be weighted towards the end of the chain and consequently the farmers and producers are the lowest paid beneficiaries. Value chains have an impact both on the livelihood of producers and on the composition and quality of products commonly sold locally and globally and consequently on the consumers. In order to understand the impact of value chains on the composition of products, we studied the production conditions for turmeric (Curcuma longa) and the metabolomic composition of products derived from it. We aimed at integrating these two components in order to gain a better understanding of the effect of different value chains on the livelihoods of some producers. This interdisciplinary project uses a mixed methods approach. Case studies were undertaken on two separate sites in India. Data was initially gathered on herbal medicine value chains by means of semi-structured interviews and non-participant observations. Samples were collected from locations in India, Europe and the USA and analysed using (1)H NMR spectroscopy coupled with multivariate analysis software and with high performance thin layer chromatography (HPTLC). We investigate medicinal plant value chains and interpret the impact different value chains have on some aspects of the livelihoods of producers in India and, for the first time, analytically assess the chemical variability and quality implications that different value chains may have on the products available to end users in Europe. There are benefits to farmers that belonged to an integrated chain and the resulting products were subject to a higher standard of processing and storage. By using analytical methods, including HPTLC and (1)H NMR spectroscopy, it has been possible to correlate some variations in product composition for selected producers and identify strengths and weaknesses of some types of value chains. The two analytical techniques provide different and complementary data and together they can be used to effectively differentiate between a wide variety of crude drug powders and herbal medicinal products. This project demonstrates that there is a need to study the links between producers and consumers of commodities produced in so-called 'provider countries' and that metabolomics offer a novel way of assessing the chemical variability along a value chain. This also has implications for understanding the impact this has on the livelihood of those along the value chain. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.
-
Day, Ryan; Joo, Hyun; Chavan, Archana; Lennox, Kristin P.; Chen, Ann; Dahl, David B.; Vannucci, Marina; Tsai, Jerry W.
2012-01-01
As an alternative to the common template based protein structure prediction methods based on main-chain position, a novel side-chain centric approach has been developed. Together with a Bayesian loop modeling procedure and a combination scoring function, the Stone Soup algorithm was applied to the CASP9 set of template based modeling targets. Although the method did not generate as large of perturbations to the template structures as necessary, the analysis of the results gives unique insights into the differences in packing between the target structures and their templates. Considerable variation in packing is found between target and template structures even when the structures are close, and this variation is found due to 2 and 3 body packing interactions. Outside the inherent restrictions in packing representation of the PDB, the first steps in correctly defining those regions of variable packing have been mapped primarily to local interactions, as the packing at the secondary and tertiary structure are largely conserved. Of the scoring functions used, a loop scoring function based on water structure exhibited some promise for discrimination. These results present a clear structural path for further development of a side-chain centered approach to template based modeling. PMID:23266765
-
Abruzzo, Lynne V; Barron, Lynn L; Anderson, Keith; Newman, Rachel J; Wierda, William G; O'brien, Susan; Ferrajoli, Alessandra; Luthra, Madan; Talwalkar, Sameer; Luthra, Rajyalakshmi; Jones, Dan; Keating, Michael J; Coombes, Kevin R
2007-09-01
To develop a model incorporating relevant prognostic biomarkers for untreated chronic lymphocytic leukemia patients, we re-analyzed the raw data from four published gene expression profiling studies. We selected 88 candidate biomarkers linked to immunoglobulin heavy-chain variable region gene (IgV(H)) mutation status and produced a reliable and reproducible microfluidics quantitative real-time polymerase chain reaction array. We applied this array to a training set of 29 purified samples from previously untreated patients. In an unsupervised analysis, the samples clustered into two groups. Using a cutoff point of 2% homology to the germline IgV(H) sequence, one group contained all 14 IgV(H)-unmutated samples; the other contained all 15 mutated samples. We confirmed the differential expression of 37 of the candidate biomarkers using two-sample t-tests. Next, we constructed 16 different models to predict IgV(H) mutation status and evaluated their performance on an independent test set of 20 new samples. Nine models correctly classified 11 of 11 IgV(H)-mutated cases and eight of nine IgV(H)-unmutated cases, with some models using three to seven genes. Thus, we can classify cases with 95% accuracy based on the expression of as few as three genes.
-
Cocca, Brian A.; Seal, Samarendra N.; D'Agnillo, Paolo; Mueller, Yvonne M.; Katsikis, Peter D.; Rauch, Joyce; Weigert, Martin; Radic, Marko Z.
2001-01-01
Apoptotic cells contain nuclear autoantigens that may initiate a systemic autoimmune response. To explore the mechanism of antibody binding to apoptotic cells, 3H9, a murine autoantibody with dual specificity for phospholipids and DNA, was used. H chain mutants of 3H9 were constructed, expressed as single-chain Fv (scFv) in Escherichia coli, and assessed for binding to phosphatidylserine, an antigen expressed on apoptotic cells. Both 3H9 and its germline revertant bound to dioleoyl phosphatidylserine in ELISA, and binding was enhanced by β2 glycoprotein I (β2GPI), a plasma protein that selectively binds to apoptotic cells. Higher relative affinity for DOPS-β2GPI was achieved by the introduction of Arg residues into the 3H9 H chain variable region at positions previously shown to mediate DNA binding. Specificity of the two structurally most diverse scFv for apoptotic cells was shown by flow cytometry, and two populations of scFv-bound cells were identified by differences in propidium iodide staining. The results suggest that, in autoimmunity, B cells with Ig receptors for apoptotic cells and DNA are positively selected, and that the antibodies they produce have the potential to affect the clearance and processing of apoptotic cells. PMID:11717440
-
Day, Ryan; Joo, Hyun; Chavan, Archana C; Lennox, Kristin P; Chen, Y Ann; Dahl, David B; Vannucci, Marina; Tsai, Jerry W
2013-02-01
As an alternative to the common template based protein structure prediction methods based on main-chain position, a novel side-chain centric approach has been developed. Together with a Bayesian loop modeling procedure and a combination scoring function, the Stone Soup algorithm was applied to the CASP9 set of template based modeling targets. Although the method did not generate as large of perturbations to the template structures as necessary, the analysis of the results gives unique insights into the differences in packing between the target structures and their templates. Considerable variation in packing is found between target and template structures even when the structures are close, and this variation is found due to 2 and 3 body packing interactions. Outside the inherent restrictions in packing representation of the PDB, the first steps in correctly defining those regions of variable packing have been mapped primarily to local interactions, as the packing at the secondary and tertiary structure are largely conserved. Of the scoring functions used, a loop scoring function based on water structure exhibited some promise for discrimination. These results present a clear structural path for further development of a side-chain centered approach to template based modeling. Copyright © 2012 Elsevier Ltd. All rights reserved.
-
NASA Astrophysics Data System (ADS)
Olson, R.; Evans, J. P.; Fan, Y.
2015-12-01
NARCliM (NSW/ACT Regional Climate Modelling Project) is a regional climate project for Australia and the surrounding region. It dynamically downscales 4 General Circulation Models (GCMs) using three Regional Climate Models (RCMs) to provide climate projections for the CORDEX-AustralAsia region at 50 km resolution, and for south-east Australia at 10 km resolution. The project differs from previous work in the level of sophistication of model selection. Specifically, the selection process for GCMs included (i) conducting literature review to evaluate model performance, (ii) analysing model independence, and (iii) selecting models that span future temperature and precipitation change space. RCMs for downscaling the GCMs were chosen based on their performance for several precipitation events over South-East Australia, and on model independence.Bayesian Model Averaging (BMA) provides a statistically consistent framework for weighing the models based on their likelihood given the available observations. These weights are used to provide probability distribution functions (pdfs) for model projections. We develop a BMA framework for constructing probabilistic climate projections for spatially-averaged variables from the NARCliM project. The first step in the procedure is smoothing model output in order to exclude the influence of internal climate variability. Our statistical model for model-observations residuals is a homoskedastic iid process. Comparing RCMs with Australian Water Availability Project (AWAP) observations is used to determine model weights through Monte Carlo integration. Posterior pdfs of statistical parameters of model-data residuals are obtained using Markov Chain Monte Carlo. The uncertainty in the properties of the model-data residuals is fully accounted for when constructing the projections. We present the preliminary results of the BMA analysis for yearly maximum temperature for New South Wales state planning regions for the period 2060-2079.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Nishiyama, Yasuhiro; Planque, Stephanie; Mitsuda, Yukie
2009-11-23
We describe murine monoclonal antibodies (mAbs) raised by immunization with an electrophilic gp120 analog (E-gp120) expressing the rare ability to neutralize genetically heterologous human immunodeficiency virus (HIV) strains. Unlike gp120, E-gp120 formed covalent oligomers. The reactivity of gp120 and E-gp120 with mAbs to reference neutralizing epitopes was markedly different, indicating their divergent structures. Epitope mapping with synthetic peptides and electrophilic peptide analogs indicated binary recognition of two distinct gp120 regions by anti-E-gp120 mAbs, the 421-433 and 288-306 peptide regions. Univalent Fab and single chain Fv fragments expressed the ability to recognize both peptides. X-ray crystallography of an anti-E-gp120 Fab fragmentmore » revealed two neighboring cavities, the typical antigen-binding cavity formed by the complementarity determining regions (CDRs) and another cavity dominated by antibody heavy chain variable (VH) domain framework (FR) residues. Substitution of the FR cavity VH Lys-19 residue by an Ala residue resulted in attenuated binding of the 421-433 region peptide probe. The CDRs and VH FR replacement/silent mutation ratios exceeded the ratio for a random mutation process, suggesting adaptive development of both putative binding sites. All mAbs studied were derived from VH1 family genes, suggesting biased recruitment of the V gene germ line repertoire by E-gp120. The conserved 421-433 region of gp120 is essential for HIV binding to host CD4 receptors. This region is recognized weakly by the FR of antibodies produced without exposure to HIV, but it usually fails to induce adaptive synthesis of neutralizing antibodies. We present models accounting for improved CD4-binding site recognition and broad HIV neutralizing activity of the mAbs, long sought goals in HIV vaccine development.« less
-
Bowers, Peter M.; Neben, Tamlyn Y.; Tomlinson, Geoffery L.; Dalton, Jennifer L.; Altobell, Larry; Zhang, Xue; Macomber, John L.; Wu, Betty F.; Toobian, Rachelle M.; McConnell, Audrey D.; Verdino, Petra; Chau, Betty; Horlick, Robert A.; King, David J.
2013-01-01
A method for simultaneous humanization and affinity maturation of monoclonal antibodies has been developed using heavy chain complementarity-determining region (CDR) 3 grafting combined with somatic hypermutation in vitro. To minimize the amount of murine antibody-derived antibody sequence used during humanization, only the CDR3 region from a murine antibody that recognizes the cytokine hβNGF was grafted into a nonhomologous human germ line V region. The resulting CDR3-grafted HC was paired with a CDR-grafted light chain, displayed on the surface of HEK293 cells, and matured using in vitro somatic hypermutation. A high affinity humanized antibody was derived that was considerably more potent than the parental antibody, possessed a low pm dissociation constant, and demonstrated potent inhibition of hβNGF activity in vitro. The resulting antibody contained half the heavy chain murine donor sequence compared with the same antibody humanized using traditional methods. PMID:23355464
-
Haikonen, Tuuli; Rajamäki, Minna-Liisa; Tian, Yan-Ping; Valkonen, Jari P T
2013-07-01
Helper component proteinase (HCpro) is a multifunctional protein of potyviruses (genus Potyvirus). HCpro of Potato virus A (PVA) interacts with the microtubule-associated protein HIP2 in host cells, and depletion of HIP2 reduces virus accumulation. This study shows that HCpro of Potato virus Y and Tobacco etch virus also interact with HIP2. The C-proximal portion of PVA HCpro determines the interaction with HIP2 and was found to contain a stretch of six residues comprising a highly variable region (HVR) in potyviruses. Mutations in HVR reduced PVA accumulation in tobacco plants and induced necrotic symptoms novel to PVA. Microarray and quantitative reverse transcription polymerase chain reaction analyses revealed induction of many defense-related genes including ethylene- and jasmonic acid-inducible pathways in systemically infected leaves at necrosis onset. Salicylic acid-mediated signaling was dispensable for the response. Genes related to microtubule functions were down-regulated. Structural modeling of HCpro suggested that all mutations in HVR caused conformational changes in adjacent regions containing functionally important motifs conserved in potyviruses. Those mutations, which also caused conformational changes in HVR, led to the greatest reduction of fitness. Our results implicate HVR in the regulation of HCpro conformation and virus-host interactions and suggest that mutation of HVR induces host defense.
-
Shark Ig light chain junctions are as diverse as in heavy chains.
Fleurant, Marshall; Changchien, Lily; Chen, Chin-Tung; Flajnik, Martin F; Hsu, Ellen
2004-11-01
We have characterized a small family of four genes encoding one of the three nurse shark Ig L chain isotypes, called NS5. All NS5 cDNA sequences are encoded by three loci, of which two are organized as conventional clusters, each consisting of a V and J gene segment that can recombine and one C region exon; the third contains a germline-joined VJ in-frame and the fourth locus is a pseudogene. This is the second nurse shark L chain type where both germline-joined and split V-J organizations have been found. Since there are only two rearranging Ig loci, it was possible for the first time to examine junctional diversity in defined fish Ig genes, comparing productive vs nonproductive rearrangements. N region addition was found to be considerably more extensive in length and in frequency than any other vertebrate L chain so far reported and rivals that in H chain. We put forth the speculation that the unprecedented efficiency of N region addition (87-93% of NS5 sequences) may be a result not only of simultaneous H and L chain rearrangement in the shark but also of processing events that afford greater accessibility of the V or J gene coding ends to terminal deoxynucleotidyltransferase.
-
Value of Conditioned Reinforcers as a Function of Temporal Context
ERIC Educational Resources Information Center
O'Daly, Matthew; Meyer, Steven; Fantino, Edmund
2005-01-01
In two experiments, pigeons were trained on a multiple-chain schedule, in which the initial link for one chain was a variable-interval (VI) 100s schedule and for the other chain a VI 10s schedule. The terminal links were both fixed-time 30s schedules signaled by differently colored stimuli. Following training, the pigeons had their preference for…
-
Piva, S; Gariano, G R; Bonilauri, P; Giacometti, F; Decastelli, L; Florio, D; Massella, E; Serraino, A
2017-04-01
This comparative study investigated the occurrence of cadF, cj1349, ciaB, pldA, tlyA, hecA, hecB, mviN, irgA and IroE genes in 212 Arcobacter butzleri isolated from three different environmental sites linked to the dairy chain (farms, industrial and artisanal dairy plants) located in three Italian regions (Lombardy, Emilia-Romagna and Calabria). According to the presence of these genes, different pathotypes (P-types) were determined. The main genes detected were ciaB, mviN, tlyA, cj1349, pldA and cadF, while the least common genes were iroE, hecA, hecB and irgA. TlyA, irgA, hecA, hecB and iroE, which were significantly more frequent in isolates recovered in industrial dairy plants. Twelve P-types were detected. The occurrence of the most frequently detected P-types (P-types 1, 2, 3 and 5) differed significantly (P < 0·001) in relation to both the environmental site and geographical area of isolation. The highest diversity in P-types was observed in industrial dairy plants and in the Calabria region. The results of this study show a correlation between the occurrence of putative virulence genes and virulence genotype variability depending on the environmental site and geographical origin of the isolates. The present study provides insights into the similar distribution of putative virulence genes in a dairy chain and other sources' isolates and also into a geographical distribution of some P-types. We have shown that industrial dairy plants may represent an environmental site favouring a selection of the isolates with a higher pathogenetic pattern. © 2017 The Society for Applied Microbiology.
-
Synthetic Fab Fragments that Bind the HIV-1 gp41 Heptad Repeat Regions
Liu, Yanyun; Regula, Lauren K.; Stewart, Alex; Lai, Jonathan R.
2011-01-01
Recent work has demonstrated that antibody phage display libraries containing restricted diversity in the complementarity determining regions (CDRs) can be used to target a wide variety of antigens with high affinity and specificity. In the most extreme case, antibodies whose combining sites are comprised of only two residues – tyrosine and serine – have been identified against several protein antigens. [F. A. Fellouse, B. Li, D. M. Compaan, A. A. Peden, S. G. Hymowitz, and S. S. Sidhu, J. Mol. Biol., 348 (2005) 1153–1162.] Here, we report the isolation and characterization of antigen-binding fragments (Fabs) from such “minimalist” diversity synthetic antibody libraries that bind the heptad repeat regions of human immunodeficiency virus type 1 (HIV-1) gp41. We show that these Fabs are highly specific for the HIV-1 epitope and comparable in affinity to a single chain variable fragment (scFv) derived from a natural antibody repertoire that targets the same region. Since the heptad repeat regions of HIV-1 gp41 are required for viral entry, these Fabs have potential for use in therapeutic, research, or diagnostic applications. PMID:21925149
-
Persistent drying in the tropics linked to natural forcing.
Winter, Amos; Zanchettin, Davide; Miller, Thomas; Kushnir, Yochanan; Black, David; Lohmann, Gerrit; Burnett, Allison; Haug, Gerald H; Estrella-Martínez, Juan; Breitenbach, Sebastian F M; Beaufort, Luc; Rubino, Angelo; Cheng, Hai
2015-07-14
Approximately half of the world's population lives in the tropics, and future changes in the hydrological cycle will impact not just the freshwater supplies but also energy production in areas dependent upon hydroelectric power. It is vital that we understand the mechanisms/processes that affect tropical precipitation and the eventual surface hydrological response to better assess projected future regional precipitation trends and variability. Paleo-climate proxies are well suited for this purpose as they provide long time series that pre-date and complement the present, often short instrumental observations. Here we present paleo-precipitation data from a speleothem located in Mesoamerica that reveal large multi-decadal declines in regional precipitation, whose onset coincides with clusters of large volcanic eruptions during the nineteenth and twentieth centuries. This reconstruction provides new independent evidence of long-lasting volcanic effects on climate and elucidates key aspects of the causal chain of physical processes determining the tropical climate response to global radiative forcing.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Guard-Petter, J.; Parker, C.T.; Asokan, K.
1999-05-01
Twelve human and chicken isolates of Salmonella enterica serovar Enteritidis belonging to phage types 4, 8, 13a, and 23 were characterized for variability in lipopolysaccharide (LPS) composition. Isolates were differentiated into two groups, i.e., those that lacked immunoreactive O-chain, termed rough isolates, and those that had immunoreactive O-chain, termed smooth isolates. Isolates within these groups could be further differentiated by LPS compositional differences as detected by gel electrophoresis and gas liquid chromatography of samples extracted with water, which yielded significantly more LPS in comparison to phenol-chloroform extraction. The rough isolates were of two types, the O-antigen synthesis mutants and themore » O-antigen polymerization (wzy) mutants. Smooth isolates were also of two types, one producing low-molecular-weight (LMW) LPS and the other producing high-molecular-weight (HMW) LPS. To determine the genetic basis for the O-chain variability of the smooth isolates, the authors analyzed the effects of a null mutation in the O-chain length determinant gene, wzz (cld) of serovar Typhimurium. This mutation results in a loss of HMW LPS; however, the LMW LPS of this mutant was longer and more glucosylated than that from clinical isolates of serovar Enteritidis. Cluster analysis of these data and of those from two previously characterized isogenic strains of serovar Enteritidis that had different virulence attributes indicated that glucosylation of HMW LPS (via oafR function) is variable and results in two types of HMW structures, one that is highly glucosylated and one that is minimally glucosylated. These results strongly indicate that naturally occurring variability in wzy, wzz, and oafR function can be used to subtype isolates of serovar Enteritidis during epidemiological investigations.« less
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Berenstein, David; Kavli Institute for Theoretical Physics, University of California at Santa Barbara, California 93106; Correa, Diego H.
We study an XXX open spin chain with variable number of sites, where the variability is introduced only at the boundaries. This model arises naturally in the study of giant gravitons in the anti-de Sitter-space/conformal field-theory correspondence. We show how to quantize the spin chain by mapping its states to a bosonic lattice of finite length with sources and sinks of particles at the boundaries. Using coherent states, we show how the Hamiltonian for the bosonic lattice gives the correct description of semiclassical open strings ending on giant gravitons.
-
Xu, X-M; Nicholson, P; Thomsett, M A; Simpson, D; Cooke, B M; Doohan, F M; Brennan, J; Monaghan, S; Moretti, A; Mule, G; Hornok, L; Beki, E; Tatnell, J; Ritieni, A; Edwards, S G
2008-01-01
ABSTRACT Over 4 years, the environmental conditions and the causal agents of Fusarium head blight (FHB) disease of wheat were determined in field sites in four European countries: Hungary, Ireland, Italy, and the United Kingdom. Polymerase chain reaction-based methods were used to detect each species causing FHB and quantify its DNA (as a measurement of fungal abundance) in the samples. Canonical correspondence analysis (CCA) was used to determine the relationship of the incidence and abundance of each species with weather variables. CCA indicated that little variability in the species prevalence data was explained by the weather variables. In contrast, a greater proportion of variability in abundance data was accounted for by the weather variables. Most samples contained two or more species and statistical analysis suggested that these species tended to coexist at field sites. CCA also indicated that there were differences in the relationships of the prevalence and abundance of the six FHB species with environmental variables. Fusarium poae was associated with relatively drier and warmer conditions, whereas F. graminearum was associated with warmer/humid conditions. F. avenaceum and F. culmorum were both associated with niches of cooler/wet/humid conditions. Two Microdochium species were associated with regions of relatively cool/moderate temperatures and frequent rainfalls of short duration. The results also suggested that environmental conditions differentially affect the infection and colonization processes, and the comparative abundance of the six species.
-
Tomanov, Konstantin; Nehlin, Lilian; Ziba, Ionida
2018-01-01
The small ubiquitin-related modifier (SUMO) conjugation apparatus usually attaches single SUMO moieties to its substrates, but SUMO chains have also been identified. To better define the biochemical requirements and characteristics of SUMO chain formation, mutations in surface-exposed Lys residues of Arabidopsis SUMO-conjugating enzyme (SCE) were tested for in vitro activity. Lys-to-Arg changes in the amino-terminal region of SCE allowed SUMO acceptance from SUMO-activating enzyme and supported substrate mono-sumoylation, but these mutations had significant effects on SUMO chain assembly. We found no indication that SUMO modification of SCE promotes chain formation. A substrate was identified that is modified by SUMO chain addition, showing that SCE can distinguish substrates for either mono-sumoylation or SUMO chain attachment. It is also shown that SCE with active site Cys mutated to Ser can accept SUMO to form an oxyester, but cannot transfer this SUMO moiety onto substrates, explaining a previously known dominant negative effect of this mutation. PMID:29133528
-
Ma, Quan-Ping; Su, Liang; Liu, Jing-Wen; Yao, Ming-Xiao; Yuan, Guang-Ying
2018-06-01
The aim of the present study was to investigate the correlation between the multi‑drug resistance of Shigella flexneri and the drug‑resistant gene cassette carried by integrons; in the meanwhile, to detect the associations between drug‑resistance and gene mutations of the active efflux pump acrAB‑tolC gene and its regulatory genes, including marOR, acrR and soxS. A total of 158 isolates were isolated from the stool samples of 1,026 children with diarrhoea aged 14 years old between May 2012 and October 2015 in Henan. The K‑B method was applied for the determination of drug resistance of Shigella flexneri, and polymerase chain reaction amplification was used for class 1, 2 and 3 integrase genes. Enzyme digestion and sequence analysis were performed for the variable regions of positive strains. Based on the drug sensitivity assessment, multi‑drug resistant strains that were resistant to five or more antibiotics, and sensitive strains were selected for amplification. Their active efflux pump genes, acrA and acrB, and regulatory genes, marOR, acrR and soxS, were selected for sequencing. The results revealed that 91.1% of the 158 strains were multi‑resistant to ampicillin, chloramphenicol, tetracycline and streptomycin, and 69.6% of the strains were multi‑resistant to sulfamethoxazole/trimethoprim. The resistance to ceftazidime, ciprofloxacin and levofloxacin was <32.9%. All strains (100%) were sensitive to cefoxitin, cefoperazone/sulbactam and imipenem. The rate of the class 1 integron positivity was 91.9% (144/158). Among these class 1 integron‑positive strains, 18 strains exhibited the resistance gene cassette dfrV in the variable region of the strain, four strains exhibited dfrA17‑aadA5 in the variable region and 140 strains exhibited blaOXA‑30‑aadA1 in the variable region. Four strains showed no resistance gene in the variable regions. The rate of class 2 integron positivity was 86.1% (136/158), and all positive strains harboured the dfrA1‑sat1‑aadA resistance gene cassette in the variable region. The class 3 integrase gene was not detected in these strains. The gene sequencing showed the deletion of base CATT in the 36, 37, 38, 39 site in the marOR gene, which is a regulatory gene of the active efflux pump, AcrAB‑TolC. Taken together, the multi‑drug resistance of Shigella flexneri was closely associated with gene mutations of class 1 and 2 integrons and the marOR gene.
-
Lemma, Habtamu Regassa; Singh, Rajwinder; Kaur, Navjot
2015-01-01
Coordination of different business activities among units becomes vital as organizations pay much attention to their core activities. Thus, their fruitfulness constantly relies on their capacity to coordinate their internal and external activities in the supply chain outside their own boundaries. Giving consideration to these obvious reasons, this paper aims to investigate the determinants of supply chain coordination of milk and dairy industries. The data were collected from 330 milk suppliers, processors, and retailers in the central part of Ethiopia. The structural equation modeling has been employed to develop the structural relationship between key constructs and measured variables. In total, 15 measured variables for coordination in the supply chain have been identified. These are further grouped into four factors namely, non-price coordination, price coordination, relationships and product development decision. It has been observed that the implementation of these factors could maximize the coordination linkage among supply chain members. Thus, dairy sectors should take the identified coordination factors into account in each of their business dealings.
-
The role of discharge variation in scaling of drainage area and food chain length in rivers
Sabo, John L.; Finlay, Jacques C.; Kennedy, Theodore A.; Post, David M.
2010-01-01
Food chain length (FCL) is a fundamental component of food web structure. Studies in a variety of ecosystems suggest that FCL is determined by energy supply, environmental stability, and/or ecosystem size, but the nature of the relationship between environmental stability and FCL, and the mechanism linking ecosystem size to FCL, remain unclear. Here we show that FCL increases with drainage area and decreases with hydrologic variability and intermittency across 36 North American rivers. Our analysis further suggests that hydrologic variability is the mechanism underlying the correlation between ecosystem size and FCL in rivers. Ecosystem size lengthens river food chains by integrating and attenuating discharge variation through stream networks, thereby enhancing environmental stability in larger river systems.
-
The role of discharge variation in scaling of drainage area and food chain length in rivers.
Sabo, John L; Finlay, Jacques C; Kennedy, Theodore; Post, David M
2010-11-12
Food chain length (FCL) is a fundamental component of food web structure. Studies in a variety of ecosystems suggest that FCL is determined by energy supply, environmental stability, and/or ecosystem size, but the nature of the relationship between environmental stability and FCL, and the mechanism linking ecosystem size to FCL, remain unclear. Here we show that FCL increases with drainage area and decreases with hydrologic variability and intermittency across 36 North American rivers. Our analysis further suggests that hydrologic variability is the mechanism underlying the correlation between ecosystem size and FCL in rivers. Ecosystem size lengthens river food chains by integrating and attenuating discharge variation through stream networks, thereby enhancing environmental stability in larger river systems.
-
Quantifying side-chain conformational variations in protein structure
Miao, Zhichao; Cao, Yang
2016-01-01
Protein side-chain conformation is closely related to their biological functions. The side-chain prediction is a key step in protein design, protein docking and structure optimization. However, side-chain polymorphism comprehensively exists in protein as various types and has been long overlooked by side-chain prediction. But such conformational variations have not been quantitatively studied and the correlations between these variations and residue features are vague. Here, we performed statistical analyses on large scale data sets and found that the side-chain conformational flexibility is closely related to the exposure to solvent, degree of freedom and hydrophilicity. These analyses allowed us to quantify different types of side-chain variabilities in PDB. The results underscore that protein side-chain conformation prediction is not a single-answer problem, leading us to reconsider the assessment approaches of side-chain prediction programs. PMID:27845406
-
Quantifying side-chain conformational variations in protein structure
NASA Astrophysics Data System (ADS)
Miao, Zhichao; Cao, Yang
2016-11-01
Protein side-chain conformation is closely related to their biological functions. The side-chain prediction is a key step in protein design, protein docking and structure optimization. However, side-chain polymorphism comprehensively exists in protein as various types and has been long overlooked by side-chain prediction. But such conformational variations have not been quantitatively studied and the correlations between these variations and residue features are vague. Here, we performed statistical analyses on large scale data sets and found that the side-chain conformational flexibility is closely related to the exposure to solvent, degree of freedom and hydrophilicity. These analyses allowed us to quantify different types of side-chain variabilities in PDB. The results underscore that protein side-chain conformation prediction is not a single-answer problem, leading us to reconsider the assessment approaches of side-chain prediction programs.
-
Quantifying side-chain conformational variations in protein structure.
Miao, Zhichao; Cao, Yang
2016-11-15
Protein side-chain conformation is closely related to their biological functions. The side-chain prediction is a key step in protein design, protein docking and structure optimization. However, side-chain polymorphism comprehensively exists in protein as various types and has been long overlooked by side-chain prediction. But such conformational variations have not been quantitatively studied and the correlations between these variations and residue features are vague. Here, we performed statistical analyses on large scale data sets and found that the side-chain conformational flexibility is closely related to the exposure to solvent, degree of freedom and hydrophilicity. These analyses allowed us to quantify different types of side-chain variabilities in PDB. The results underscore that protein side-chain conformation prediction is not a single-answer problem, leading us to reconsider the assessment approaches of side-chain prediction programs.
-
Changing precipitation in western Europe, climate change or natural variability?
NASA Astrophysics Data System (ADS)
Aalbers, Emma; Lenderink, Geert; van Meijgaard, Erik; van den Hurk, Bart
2017-04-01
Multi-model RCM-GCM ensembles provide high resolution climate projections, valuable for among others climate impact assessment studies. While the application of multiple models (both GCMs and RCMs) provides a certain robustness with respect to model uncertainty, the interpretation of differences between ensemble members - the combined result of model uncertainty and natural variability of the climate system - is not straightforward. Natural variability is intrinsic to the climate system, and a potentially large source of uncertainty in climate change projections, especially for projections on the local to regional scale. To quantify the natural variability and get a robust estimate of the forced climate change response (given a certain model and forcing scenario), large ensembles of climate model simulations of the same model provide essential information. While for global climate models (GCMs) a number of such large single model ensembles exists and have been analyzed, for regional climate models (RCMs) the number and size of single model ensembles is limited, and the predictability of the forced climate response at the local to regional scale is still rather uncertain. We present a regional downscaling of a 16-member single model ensemble over western Europe and the Alps at a resolution of 0.11 degrees (˜12km), similar to the highest resolution EURO-CORDEX simulations. This 16-member ensemble was generated by the GCM EC-EARTH, which was downscaled with the RCM RACMO for the period 1951-2100. This single model ensemble has been investigated in terms of the ensemble mean response (our estimate of the forced climate response), as well as the difference between the ensemble members, which measures natural variability. We focus on the response in seasonal mean and extreme precipitation (seasonal maxima and extremes with a return period up to 20 years) for the near to far future. For most precipitation indices we can reliably determine the climate change signal, given the applied model chain and forcing scenario. However, the analysis also shows how limited the information in single ensemble members is on the local scale forced climate response, even for high levels of global warming when the forced response has emerged from natural variability. Analysis and application of multi-model ensembles like EURO-CORDEX should go hand-in-hand with single model ensembles, like the one presented here, to be able to correctly interpret the fine-scale information in terms of a forced signal and random noise due to natural variability.
-
From Chains for Mean Value Inequalities to Mitrinovic's Problem II
ERIC Educational Resources Information Center
Zhu, Ling
2005-01-01
In this note, an integrated form of some significant means with two variables is provided, and some chains for mean value inequalities are obtained. At the same time, a concise family of algebraic functions appears, which satisfy Mitrinovic's requirements.
-
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
-
Yang, Tao; Yang, Lijun; Chai, Weiran; Li, Renke; Xie, Jun; Niu, Bo
2011-03-01
A phage display single-chain variable fragment (scFv) library against TNFα was constructed using a recombinant phage antibody system (RPAS). The cloned scFv gene was introduced into the phage display vector pCANTAB 5E and expressed in Escherichia coli (E. coli) with a yield of up to 0.15 mg/l of total protein. With the attempt to improve the expression level of TNF-scFv, a strategy was established for subcloning the scFv gene from pCANTAB 5E into the plasmid pBV220. Under the control of a highly efficient tandem P(R)P(L) promoter system, scFv production was increased to 30% of total protein as inclusion bodies. After extraction from the cell pellet by sonication, the inclusion bodies were solubilized and denatured in the presence of 8M urea. Purification of denatured scFv was performed using nickel column chromatography followed by renaturation. The purity and activity of the refolded scFv were confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Western blotting and by an enzyme-linked immunoabsorbent assay (ELISA). The results reveal that the overall yield of bioactive TNF-scFv from E. coli flask cultures was more than 45 mg/l culture medium and 15 mg/g wet weight cells. The renatured scFv exhibited binding activity similarly to soluble scFv. In conclusion we developed a method to over-express TNF-scFv, which have biological function after purification and renaturation. Copyright © 2010 Elsevier Inc. All rights reserved.
-
Allegrucci, M; Young-Cooper, G O; Alexander, C B; Newman, B A; Mage, R G
1991-02-01
The rabbit is unique in having well-defined allotypes in the variable region of the heavy chain. Products of the VHa locus, (with alleles a1, a2, and a3), account for the majority of the serum immunoglobulins. A small percentage of the serum immunoglobulins are a-negative. In 1986, Kelus and Weiss described a mutation that depressed the expression of the Ig VH a2 genes in an a1/a2 rabbit. From this animal the Alicia rabbit strain was developed and the mutation was termed ali. We previously showed, using Southern analysis and the transverse alternating field electrophoresis technique, that the difference between the ali rabbit and normal is a relatively small deletion including some of the most 3' VH genes. The most JH proximal 3' VH1 genes in DNA from normal rabbits of a1, a2 and a3 haplotypes encode a1, a2 and a3 molecules respectively, and it has been suggested that these genes are responsible for allelic inheritance of VHa allotypes. The present study suggests that the 3' end of the VH locus probably plays a key role in regulation of VH gene expression in rabbits because VH gene(s) in this region are the target(s) of preferential VDJ rearrangements. This raises the possibility that mechanisms such as somatic gene conversion and hypermutation are at work to generate the antibody repertoire in this species. Our data support the view that the 3' VH1 gene may be the preferred target for rearrangement in normal rabbits, and for the normal chromosome in heterozygous ali animals. However, homozygous ali rabbits with a deletion that removed the a2-encoding VH1 on both chromosomes do survive, rearrange other VH genes and produce normal levels of immunoglobulins as well as a significant percentage of B cells which bear the a2 allotype. This challenges the view that one VH gene, VH1, is solely responsible for the inheritance pattern of VHa allotypes.
-
Imai, Kohsuke; Catalan, Nadia; Plebani, Alessandro; Maródi, László; Sanal, Özden; Kumaki, Satoru; Nagendran, Vasantha; Wood, Philip; Glastre, Catherine; Sarrot-Reynauld, Françoise; Hermine, Olivier; Forveille, Monique; Revy, Patrick; Fischer, Alain; Durandy, Anne
2003-01-01
Hyper-IgM syndrome (HIGM) is a heterogeneous condition characterized by impaired Ig class-switch recombination (CSR). The molecular defects that have so far been associated with this syndrome — which affect the CD40 ligand in HIGM type 1 (HIGM1), CD40 in HIGM3, and activation-induced cytidine deaminase (AID) in HIGM2 — do not account for all cases. We investigated the clinical and immunological characteristics of 15 patients with an unidentified form of HIGM. Although the clinical manifestations were similar to those observed in HIGM2, these patients exhibited a slightly milder HIGM syndrome with residual IgG production. We found that B cell CSR was intrinsically impaired. However, the generation of somatic hypermutations was observed in the variable region of the Ig heavy chain gene, as in control B lymphocytes. In vitro studies showed that the molecular defect responsible for this new HIGM entity (HIGM4) occurs downstream of the AID activity, as the AID gene was induced normally and AID-induced DNA double-strand breaks in the switch μ region of the Ig heavy chain locus were detected during CSR as normal. Thus, HIGM4 is probably the consequence of a selective defect either in a CSR-specific factor of the DNA repair machinery or in survival signals delivered to switched B cells. PMID:12840068
-
Obwaller, A; Duchêne, M; Bruhn, H; Steipe, B; Tripp, C; Kraft, D; Wiedermann, G; Auer, H; Aspöck, H
2001-05-01
Myosins from nematode parasites elicit strong humoral and cellular immune responses and have been investigated as vaccine candidates. In this study we cloned and sequenced a cDNA coding for myosin heavy chain from Toxocara canis, a nematode parasite of canids which may also infect humans and cause various unspecific symptoms. To determine the major antigenic regions the myosin heavy chain was systematically dissected into ten overlapping recombinant fusion polypeptides which were purified by metal chelate chromatography. Single fragments were then tested for their IgG reactivity in sera from toxocarosis patients and healthy probands. Two regions, one region at the mid to carboxy-terminal end of the head domain and one region in the rod domain, were identified as major antigens, which in combination were positive with 86% of the sera. The other domains were less reactive. This shows that the patients' IgG reactivity was not directed evenly against all parts of the molecule, but was rather clustered in few regions.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Castillo-Villar, Krystel K.; Eksioglu, Sandra; Taherkhorsandi, Milad
The production of biofuels using second-generation feedstocks has been recognized as an important alternative source of sustainable energy and its demand is expected to increase due to regulations such as the Renewable Fuel Standard. However, the pathway to biofuel industry maturity faces unique, unaddressed challenges. Here, to address this challenges, this article presents an optimization model which quantifies and controls the impact of biomass quality variability on supply chain related decisions and technology selection. We propose a two-stage stochastic programming model and associated efficient solution procedures for solving large-scale problems to (1) better represent the random nature of the biomassmore » quality (defined by moisture and ash contents) in supply chain modeling, and (2) assess the impact of these uncertainties on the supply chain design and planning. The proposed model is then applied to a case study in the state of Tennessee. Results show that high moisture and ash contents negatively impact the unit delivery cost since poor biomass quality requires the addition of quality control activities. Experimental results indicate that supply chain cost could increase as much as 27%–31% when biomass quality is poor. We assess the impact of the biomass quality on the topological supply chain. Our case study indicates that biomass quality impacts supply chain costs; thus, it is important to consider the impact of biomass quality in supply chain design and management decisions.« less
-
Castillo-Villar, Krystel K.; Eksioglu, Sandra; Taherkhorsandi, Milad
2017-02-20
The production of biofuels using second-generation feedstocks has been recognized as an important alternative source of sustainable energy and its demand is expected to increase due to regulations such as the Renewable Fuel Standard. However, the pathway to biofuel industry maturity faces unique, unaddressed challenges. Here, to address this challenges, this article presents an optimization model which quantifies and controls the impact of biomass quality variability on supply chain related decisions and technology selection. We propose a two-stage stochastic programming model and associated efficient solution procedures for solving large-scale problems to (1) better represent the random nature of the biomassmore » quality (defined by moisture and ash contents) in supply chain modeling, and (2) assess the impact of these uncertainties on the supply chain design and planning. The proposed model is then applied to a case study in the state of Tennessee. Results show that high moisture and ash contents negatively impact the unit delivery cost since poor biomass quality requires the addition of quality control activities. Experimental results indicate that supply chain cost could increase as much as 27%–31% when biomass quality is poor. We assess the impact of the biomass quality on the topological supply chain. Our case study indicates that biomass quality impacts supply chain costs; thus, it is important to consider the impact of biomass quality in supply chain design and management decisions.« less
-
STCRDab: the structural T-cell receptor database
de Oliveira, Saulo H P; Krawczyk, Konrad
2018-01-01
Abstract The Structural T–cell Receptor Database (STCRDab; http://opig.stats.ox.ac.uk/webapps/stcrdab) is an online resource that automatically collects and curates TCR structural data from the Protein Data Bank. For each entry, the database provides annotations, such as the α/β or γ/δ chain pairings, major histocompatibility complex details, and where available, antigen binding affinities. In addition, the orientation between the variable domains and the canonical forms of the complementarity-determining region loops are also provided. Users can select, view, and download individual or bulk sets of structures based on these criteria. Where available, STCRDab also finds antibody structures that are similar to TCRs, helping users explore the relationship between TCRs and antibodies. PMID:29087479
-
Human anti-CD30 recombinant antibodies by guided phage antibody selection using cell panning
Klimka, A; Matthey, B; Roovers, R C; Barth, S; Arends, J-W; Engert, A; Hoogenboom, H R
2000-01-01
In various clinical studies, Hodgkin’s patients have been treated with anti-CD30 immunotherapeutic agents and have shown promising responses. One of the problems that appeared from these studies is the development of an immune response against the non-human therapeutics, which limits repeated administration and reduces efficacy. We have set out to make a recombinant, human anti-CD30 single-chain variable fragment (scFv) antibody, which may serve as a targeting moiety with reduced immunogenicity and more rapid tumour penetration in similar clinical applications. Rather than selecting a naive phage antibody library on recombinant CD30 antigen, we used guided selection of a murine antibody in combination with panning on the CD30-positive cell line L540. The murine monoclonal antibody Ki-4 was chosen as starting antibody, because it inhibits the shedding of the extracellular part of the CD30 antigen. This makes the antibody better suited for CD30-targeting than most other anti-CD30 antibodies. We have previously isolated the murine Ki-4 scFv by selecting a mini-library of hybridoma-derived phage scFv-antibodies via panning on L540 cells. Here, we report that phage display technology was successfully used to obtain a human Ki-4 scFv version by guided selection. The murine variable heavy (VH) and light (VL) chain genes of the Ki-4 scFv were sequentially replaced by human V gene repertoires, while retaining only the major determinant for epitope-specificity: the heavy-chain complementarity determining region 3 (CDR3) of murine Ki-4. After two rounds of chain shuffling and selection by panning on L540 cells, a fully human anti-CD30 scFv was selected. It competes with the parental monoclonal antibody Ki-4 for binding to CD30, inhibits the shedding of the extracellular part of the CD30 receptor from L540 cells and is thus a promising candidate for the generation of anti-CD30 immunotherapeutics. © 2000 Cancer Research Campaign PMID:10901379
-
Muchima, Kaname; Todaka, Taro; Shinchi, Hiroyuki; Sato, Ayaka; Tazoe, Arisa; Aramaki, Rikiya; Kakitsubata, Yuhei; Yokoyama, Risa; Arima, Naomichi; Baba, Masanori; Wakao, Masahiro; Ito, Yuji; Suda, Yasuo
2018-04-01
Adult T-cell leukemia (ATL) is an intractable blood cancer caused by the infection of human T-cell leukemia virus type-1, and effective medical treatment is required. It is known that the structure and expression levels of cell surface sugar chains vary depending on cell states such as inflammation and cancer. Thus, it is expected that the antibody specific for ATL cell surface sugar chain would be an effective diagnostic tool and a strong candidate for the development of an anti-ATL drug. Here, we developed a stable sugar chain-binding single-chain variable fragment antibody (scFv) that can bind to ATL cells using a fibre-type Sugar Chip and phage display method. The fiber-type Sugar Chips were prepared using O-glycans released from ATL cell lines. The scFv-displaying phages derived from human B cells (diversity: 1.04 × 108) were then screened using the fiber-type Sugar Chips, and an O-glycan-binding scFv was obtained. The flow cytometry analysis revealed that the scFv predominantly bound to ATL cell lines. The sugar chain-binding properties of the scFv was evaluated by array-type Sugar Chip immobilized with a library of synthetic glycosaminoglycan disaccharide structures. Highly sulphated disaccharide structures were found to have high affinity to scFv.
-
Quantifying Short-Chain Chlorinated Paraffin Congener Groups.
Yuan, Bo; Bogdal, Christian; Berger, Urs; MacLeod, Matthew; Gebbink, Wouter A; Alsberg, Tomas; de Wit, Cynthia A
2017-09-19
Accurate quantification of short-chain chlorinated paraffins (SCCPs) poses an exceptional challenge to analytical chemists. SCCPs are complex mixtures of chlorinated alkanes with variable chain length and chlorination level; congeners with a fixed chain length (n) and number of chlorines (m) are referred to as a "congener group" C n Cl m . Recently, we resolved individual C n Cl m by mathematically deconvolving soft ionization high-resolution mass spectra of SCCP mixtures. Here we extend the method to quantifying C n Cl m by introducing C n Cl m specific response factors (RFs) that are calculated from 17 SCCP chain-length standards with a single carbon chain length and variable chlorination level. The signal pattern of each standard is measured on APCI-QTOF-MS. RFs of each C n Cl m are obtained by pairwise optimization of the normal distribution's fit to the signal patterns of the 17 chain-length standards. The method was verified by quantifying SCCP technical mixtures and spiked environmental samples with accuracies of 82-123% and 76-109%, respectively. The absolute differences between calculated and manufacturer-reported chlorination degrees were -0.9 to 1.0%Cl for SCCP mixtures of 49-71%Cl. The quantification method has been replicated with ECNI magnetic sector MS and ECNI-Q-Orbitrap-MS. C n Cl m concentrations determined with the three instruments were highly correlated (R 2 > 0.90) with each other.
-
The Canadian High Arctic Ionospheric Network (CHAIN)
NASA Astrophysics Data System (ADS)
Jayachandran, P. T.; Langley, R. B.; MacDougall, J. W.; Mushini, S. C.; Pokhotelov, D.; Chadwick, R.; Kelly, T.
2009-05-01
Polar cap ionospheric measurements are important for the complete understanding of the various processes in the solar wind - magnetosphere - ionosphere (SW-M-I) system as well as for space weather applications. Currently the polar cap region is lacking high temporal and spatial resolution ionospheric measurements because of the orbit limitations of space-based measurements and the sparse network providing ground- based measurements. Canada has a unique advantage in remedying this shortcoming because it has the most accessible landmass in the high Arctic regions and the Canadian High Arctic Ionospheric Network (CHAIN) is designed to take advantage of Canadian geographic vantage points for a better understanding of the Sun-Earth system. CHAIN is a distributed array of ground-based radio instruments in the Canadian high Arctic. The instruments components of CHAIN are ten high data-rate Global Positioning System ionospheric scintillation and total electron content monitors and six Canadian Advanced Digital Ionosondes. Most of these instruments have been sited within the polar cap region except for two GPS reference stations at lower latitudes. This paper briefly overviews the scientific capabilities, instrument components, and deployment status of CHAIN.
-
Breathers and rogue waves in a Heisenberg ferromagnetic spin chain or an alpha helical protein
NASA Astrophysics Data System (ADS)
Yang, Jin-Wei; Gao, Yi-Tian; Su, Chuan-Qi; Wang, Qi-Min; Lan, Zhong-Zhou
2017-07-01
In this paper, a fourth-order variable-coefficient nonlinear Schrödinger equation for a one-dimensional continuum anisotropic Heisenberg ferromagnetic spin chain or an alpha helical protein has been investigated. Breathers and rogue waves are constructed via the Darboux transformation and generalized Darboux transformation, respectively. Results of the breathers and rogue waves are presented: (1) The first- and second-order Akhmediev breathers and Kuznetsov-Ma solitons are presented with different values of variable coefficients which are related to the energy transfer or higher-order excitations and interactions in the helical protein, or related to the spin excitations resulting from the lowest order continuum approximation and octupole-dipole interaction in a Heisenberg ferromagnetic spin chain, and the nonlinear periodic breathers resulting from the Akhmediev breathers are studied as well; (2) For the first- and second-order rogue waves, we find that they can be split into many similar components when the variable coefficients are polynomial functions of time; (3) Rogue waves can also be split when the variable coefficients are hyperbolic secant functions of time, but the profile of each component in such a case is different.
-
Neutralisation and binding of VHS virus by monovalent antibody fragments.
Cupit, P M; Lorenzen, N; Strachan, G; Kemp, G J; Secombes, C J; Cunningham, C
2001-12-04
We have previously reported the cloning and characterisation of the heavy and light chain variable domain genes encoding three monoclonal antibodies (Mabs) that bind viral haemorrhagic septicaemia virus (VHSV). Two of these antibodies, 3F1H10 and 3F1A2 both neutralised the virus though 3F1A2 appeared to recognise a broader range of virus isolates. The variable domains of these two antibodies differ by only four residues (Lorenzen et al., 2000a. Fish Shellfish Immunol. 10, 129-142). To further study the mechanism of neutralisation, Fab fragments as well as a series of recombinant bacterial single chain antibody (scAb) fragments were generated from the three anti-VHSV Mabs and their variable domain genes, respectively. Fabs and scAbs derived from the neutralising Mabs were both able to neutralise the VHSV type 1 isolate DK-F1. In addition, a series of scAb fragments were produced using the 3F1H10 variable heavy (VH) chain and variable light (Vkappa) chain domains but containing, either alone or in dual combination, each of the four different residues present in 3F1A2. The dissociation constants of Mabs 3F1H10 and 3F1A2 and their respective Fab and scAb fragments were measured by BIAcore analysis and found to correlate with the capacity of each molecule to neutralise DK-F1. These investigations, together with computer assisted molecular analysis of the theoretical influence of each mutation on antigen binding, led to the identification of a single mutation at position 35a in the VH domain as having the most marked impact on viral neutralisation.
-
Recombinant anti-podoplanin (NZ-1) immunotoxin for the treatment of malignant brain tumors
Chandramohan, Vidyalakshmi; Bao, Xuhui; Kaneko, Mika Kato; Kato, Yukinari; Keir, Stephen T.; Szafranski, Scott E.; Kuan, Chien-Tsun; Pastan, Ira H.; Bigner, Darell D.
2013-01-01
Current study demonstrates the glioma tumor antigen podoplanin to be present at very high levels (>90%) in both glioblastoma (D2159MG, D08-0308MG, and D08-0493MG) and medulloblastoma (D283MED, D425MED, and DAOY) xenografts and cell line. We constructed a novel recombinant single-chain antibody variable region fragment (scFv), NZ-1, specific for podoplanin from the NZ-1 hybridoma. NZ-1-scFv was then fused to Pseudomonas exotoxin A, carrying a C-terminal KDEL peptide (NZ-1-PE38KDEL). The immunotoxin was further stabilized by a disulfide (ds) bond between the heavy-chain and light-chain variable regions as the construct NZ-1-(scdsFv)-PE38KDEL. NZ-1-(scdsFv)-PE38KDEL exhibited significant reactivity to glioblastoma and medulloblastoma cells. The affinity of NZ-1-(scdsFv), NZ-1-(scdsFv)-PE38KDEL and NZ-1 antibody, for podoplanin peptide was 2.1×10−8 M, 8.0×10−8 M, and 3.9×10−10 M, respectively. In a protein stability assay, NZ-1-(scdsFv)-PE38KDEL retained 33-98% of its activity while that of NZ-1-PE38KDEL declined to 13% of its initial levels after incubation at 37°C for 3 days. In vitro cytotoxicity of the NZ-1-(scdsFv)-PE38KDEL was measured in cells isolated from glioblastoma xenografts, D2159MG, D08-0308MG, D08-0493MG, and in the medulloblastoma D283MED, D425MED, and DOAY xenografts and cell line. The NZ-1-(scdsFv)-PE38KDEL immunotoxin was highly cytotoxic, with an IC50 in the range of 1.6–29 ng/mL. Significantly, NZ-1-(scdsFv)-PE38KDEL demonstrated tumor-growth delay, averaging 24 days (P<0.001) and 21 days (P<0.001) in D2159MG and D283MED in vivo tumor models, respectively. Crucially, in the D425MED intracranial tumor model, NZ-1-(scdsFv)-PE38KDEL caused a 41% increase in survival (P≤0.001). In preclinical studies, NZ-1-(scdsFv)-PE38KDEL exhibited significant potential as a targeting agent for malignant brain tumors. PMID:23115013
-
Recombinant anti-podoplanin (NZ-1) immunotoxin for the treatment of malignant brain tumors.
Chandramohan, Vidyalakshmi; Bao, Xuhui; Kato Kaneko, Mika; Kato, Yukinari; Keir, Stephen T; Szafranski, Scott E; Kuan, Chien-Tsun; Pastan, Ira H; Bigner, Darell D
2013-05-15
Our study demonstrates the glioma tumor antigen podoplanin to be present at very high levels (>90%) in both glioblastoma (D2159MG, D08-0308MG and D08-0493MG) and medulloblastoma (D283MED, D425MED and DAOY) xenografts and cell line. We constructed a novel recombinant single-chain antibody variable region fragment (scFv), NZ-1, specific for podoplanin from the NZ-1 hybridoma. NZ-1-scFv was then fused to Pseudomonas exotoxin A, carrying a C-terminal KDEL peptide (NZ-1-PE38KDEL). The immunotoxin (IT) was further stabilized by a disulfide (ds) bond between the heavy-chain and light-chain variable regions as the construct NZ-1-(scdsFv)-PE38KDEL. NZ-1-(scdsFv)-PE38KDEL exhibited significant reactivity to glioblastoma and medulloblastoma cells. The affinity of NZ-1-(scdsFv), NZ-1-(scdsFv)-PE38KDEL and NZ-1 antibody for podoplanin peptide was 2.1 × 10(-8) M, 8.0 × 10(-8) M and 3.9 × 10(-10) M, respectively. In a protein stability assay, NZ-1-(scdsFv)-PE38KDEL retained 33-98% of its activity, whereas that of NZ-1-PE38KDEL declined to 13% of its initial levels after incubation at 37°C for 3 days. In vitro cytotoxicity of the NZ-1-(scdsFv)-PE38KDEL was measured in cells isolated from glioblastoma xenografts, D2159MG, D08-0308MG and D08-0493MG, and in the medulloblastoma D283MED, D425MED and DOAY xenografts and cell line. The NZ-1-(scdsFv)-PE38KDEL IT was highly cytotoxic, with an 50% inhibitory concentration in the range of 1.6-29 ng/ml. Significantly, NZ-1-(scdsFv)-PE38KDEL demonstrated tumor growth delay, averaging 24 days (p < 0.001) and 21 days (p < 0.001) in D2159MG and D283MED in vivo tumor models, respectively. Crucially, in the D425MED intracranial tumor model, NZ-1-(scdsFv)-PE38KDEL caused a 41% increase in survival (p ≤ 0.001). In preclinical studies, NZ-1-(scdsFv)-PE38KDEL exhibited significant potential as a targeting agent for malignant brain tumors. Copyright © 2012 UICC.
-
Reservoirs of Listeria Species in Three Environmental Ecosystems
Linke, Kristina; Rückerl, Irene; Brugger, Katharina; Karpiskova, Renata; Walland, Julia; Muri-Klinger, Sonja; Tichy, Alexander; Wagner, Martin
2014-01-01
Soil and water are suggested to represent pivotal niches for the transmission of Listeria monocytogenes to plant material, animals, and the food chain. In the present study, 467 soil and 68 water samples were collected in 12 distinct geological and ecological sites in Austria from 2007 to 2009. Listeria was present in 30% and 26% of the investigated soil and water samples, respectively. Generally, the most dominant species in soil and water samples were Listeria seeligeri, L. innocua, and L. ivanovii. The human- and animal-pathogenic L. monocytogenes was isolated exclusively from 6% soil samples in regions A (mountainous region) and B (meadow). Distinct ecological preferences were observed for L. seeligeri and L. ivanovii, which were more often isolated from wildlife reserve region C (Lake Neusiedl) and from sites in proximity to wild and domestic ruminants (region A). The higher L. monocytogenes detection and antibiotic resistance rates in regions A and B could be explained by the proximity to agricultural land and urban environment. L. monocytogenes multilocus sequence typing corroborated this evidence since sequence type 37 (ST37), ST91, ST101, and ST517 were repeatedly isolated from regions A and B over several months. A higher L. monocytogenes detection and strain variability was observed during flooding of the river Schwarza (region A) and Danube (region B) in September 2007, indicating dispersion via watercourses. PMID:25002422
-
Interactions of tectonic, igneous, and hydraulic processes in the North Tharsis Region of Mars
NASA Technical Reports Server (NTRS)
Davis, P. A.; Tanaka, Kenneth L.; Golombek, M. P.; Plescia, J. B.
1991-01-01
Recent work on the north Tharsis of Mars has revealed a complex geologic history involving volcanism, tectonism, flooding, and mass wasting. Our detailed photogeologic analysis of this region found many previously unreported volcanic vents, volcaniclastic flows, irregular cracks, and minor pit chains; additional evidence that volcanic tectonic processes dominated this region throughout Martian geologic time; and the local involvement of these processes with surface and near surface water. Also, photoclinometric profiles were obtained within the region of troughs, simple grabens, and pit chains, as well as average spacings of pits along pit chains. These data were used together with techniques to estimate depths of crustal mechanical discontinuities that may have controlled the development of these features. In turn, such discontinuities may be controlled by stratigraphy, presence of water or ice, or chemical cementation.
-
Krebber, A; Bornhauser, S; Burmester, J; Honegger, A; Willuda, J; Bosshard, H R; Plückthun, A
1997-02-14
A prerequisite for the use of recombinant antibody technologies starting from hybridomas or immune repertoires is the reliable cloning of functional immunoglobulin genes. For this purpose, a standard phage display system was optimized for robustness, vector stability, tight control of scFv-delta geneIII expression, primer usage for PCR amplification of variable region genes, scFv assembly strategy and subsequent directional cloning using a single rare cutting restriction enzyme. This integrated cloning, screening and selection system allowed us to rapidly obtain antigen binding scFvs derived from spleen-cell repertoires of mice immunized with ampicillin as well as from all hybridoma cell lines tested to date. As representative examples, cloning of monoclonal antibodies against a his tag, leucine zippers, the tumor marker EGP-2 and the insecticide DDT is presented. Several hybridomas whose genes could not be cloned in previous experimental setups, but were successfully obtained with the present system, expressed high amounts of aberrant heavy and light chain mRNAs, which were amplified by PCR and greatly exceeded the amount of binding antibody sequences. These contaminating variable region genes were successfully eliminated by employing the optimized phage display system, thus avoiding time consuming sequencing of non-binding scFv genes. To maximize soluble expression of functional scFvs subsequent to cloning, a compatible vector series to simplify modification, detection, multimerization and rapid purification of recombinant antibody fragments was constructed.
-
Unravelling connections between river flow and large-scale climate: experiences from Europe
NASA Astrophysics Data System (ADS)
Hannah, D. M.; Kingston, D. G.; Lavers, D.; Stagge, J. H.; Tallaksen, L. M.
2016-12-01
The United Nations has identified better knowledge of large-scale water cycle processes as essential for socio-economic development and global water-food-energy security. In this context, and given the ever-growing concerns about climate change/ variability and human impacts on hydrology, there is an urgent research need: (a) to quantify space-time variability in regional river flow, and (b) to improve hydroclimatological understanding of climate-flow connections as a basis for identifying current and future water-related issues. In this paper, we draw together studies undertaken at the pan-European scale: (1) to evaluate current methods for assessing space-time dynamics for different streamflow metrics (annual regimes, low flows and high flows) and for linking flow variability to atmospheric drivers (circulation indices, air-masses, gridded climate fields and vapour flux); and (2) to propose a plan for future research connecting streamflow and the atmospheric conditions in Europe and elsewhere. We believe this research makes a useful, unique contribution to the literature through a systematic inter-comparison of different streamflow metrics and atmospheric descriptors. In our findings, we highlight the need to consider appropriate atmospheric descriptors (dependent on the target flow metric and region of interest) and to develop analytical techniques that best characterise connections in the ocean-atmosphere-land surface process chain. We call for the need to consider not only atmospheric interactions, but also the role of the river basin-scale terrestrial hydrological processes in modifying the climate signal response of river flows.
-
Variable camshaft timing system
DOE Office of Scientific and Technical Information (OSTI.GOV)
Butterfield, R.P.; Smith, F.R.
1989-09-05
This patent describes an improvement in a variable camshaft timing system for an internal combustion engine having intake and exhaust valves and a camshaft for each of the intake and exhaust valves, an intake sprocket and an exhaust sprocket keyed to their respective camshaft, only one of the camshafts being directly driven by an engine crankshaft, and a timing chain engaging both sprockets. The improvement comprising a single bracket carrying at least one idler sprocket engaging the timing chain, the bracket being mounted for movement to alter the timing relationship between the intake and exhaust sprockets.
-
Roldán, S; Lluch, M D; Navarro Quesada, F J; Hevia, A
1995-01-01
Reference has been made in the literature of the variability in the clinical presentation of deficiency of complex III of the respiratory chain, identifying up to the moment, four groups, the first of which is characterized by hipotonia and wearness starting at variable ages. We report a new case of mitochondrial myopathy due to deficiency of this complex and included within this first group, and consider the importance of defining the clinical and histochemical characteristics of this polymorphous entity.
-
NASA Astrophysics Data System (ADS)
Fobair, Richard C., II
This research presents a model for forecasting the numbers of jobs created in the energy efficiency retrofit (EER) supply chain resulting from an investment in upgrading residential buildings in Florida. This investigation examined material supply chains stretching from mining to project installation for three product types: insulation, windows/doors, and heating, ventilating, and air conditioning (HVAC) systems. Outputs from the model are provided for the project, sales, manufacturing, and mining level. The model utilizes reverse-estimation to forecast the numbers of jobs that result from an investment. Reverse-estimation is a process that deconstructs a total investment into its constituent parts. In this research, an investment is deconstructed into profit, overhead, and hard costs for each level of the supply chain and over multiple iterations of inter-industry exchanges. The model processes an investment amount, the type of work and method of contracting into a prediction of the number of jobs created. The deconstruction process utilizes data from the U.S. Economic Census. At each supply chain level, the cost of labor is reconfigured into full-time equivalent (FTE) jobs (i.e. equivalent to 40 hours per week for 52 weeks) utilizing loaded labor rates and a typical employee mix. The model is sensitive to adjustable variables, such as percentage of work performed per type of product, allocation of worker time per skill level, annual hours for FTE calculations, wage rate, and benefits. This research provides several new insights into job creation. First, it provides definitions that can be used for future research on jobs in supply chains related to energy efficiency. Second, it provides a methodology for future investigators to calculate jobs in a supply chain resulting from an investment in energy efficiency upgrades to a building. The methodology used in this research is unique because it examines gross employment at the sub-industry level for specific commodities. Most research on employment examines the net employment change (job creation less job destruction) at levels for regions, industries, and the aggregate economy. Third, it provides a forecast of the numbers of jobs for an investment in energy efficiency over the entire supply chain for the selected industries and the job factors for major levels of the supply chain.
-
NASA Astrophysics Data System (ADS)
Wang, S.; Huang, G. H.; Baetz, B. W.; Ancell, B. C.
2017-05-01
The particle filtering techniques have been receiving increasing attention from the hydrologic community due to its ability to properly estimate model parameters and states of nonlinear and non-Gaussian systems. To facilitate a robust quantification of uncertainty in hydrologic predictions, it is necessary to explicitly examine the forward propagation and evolution of parameter uncertainties and their interactions that affect the predictive performance. This paper presents a unified probabilistic framework that merges the strengths of particle Markov chain Monte Carlo (PMCMC) and factorial polynomial chaos expansion (FPCE) algorithms to robustly quantify and reduce uncertainties in hydrologic predictions. A Gaussian anamorphosis technique is used to establish a seamless bridge between the data assimilation using the PMCMC and the uncertainty propagation using the FPCE through a straightforward transformation of posterior distributions of model parameters. The unified probabilistic framework is applied to the Xiangxi River watershed of the Three Gorges Reservoir (TGR) region in China to demonstrate its validity and applicability. Results reveal that the degree of spatial variability of soil moisture capacity is the most identifiable model parameter with the fastest convergence through the streamflow assimilation process. The potential interaction between the spatial variability in soil moisture conditions and the maximum soil moisture capacity has the most significant effect on the performance of streamflow predictions. In addition, parameter sensitivities and interactions vary in magnitude and direction over time due to temporal and spatial dynamics of hydrologic processes.
-
Production and characterization of recombinant scFv against digoxin by phage display technology.
Alirezapour, Behruz; Rajabibazl, Masoumeh; Rasaee, Mohhamad Javad; Omidfar, Kobra
2013-06-01
The cardiac glycoside digoxin is widely used for the treatment of congestive heart failure and cardiac arrhythmias. Digoxin is a highly toxic drug and consequently is routinely measured in sera of treated patients. In such cases, antibodies are required against digoxin for detection as well as detoxification purposes. To obtain recombinant single chain antibody against digoxin, RNA was extracted from spleen of BALB/c mice immunized with digoxin-BSA and converted to cDNA. The gene fragment corresponding to the variable regions of the repertoire of antibody genes were amplified by PCR. ScFv construct was generated by randomly joining individual heavy- and light-chain variable domains through gene splicing by overlapping extension PCR. Recombinant phage library expressing scFv polypeptides were produced. Phages with higher affinity toward digoxin were selected in the biopanning process. Sensitivity of produced recombinant MAb (AR85) was determined to be about 100 pg/well, while intact MAb (BBA) produced by hybridoma technology (data not shown) was reported to be around 100 pg/well too. The saturation value for recombinant scFv MAb was found to be 1000 ng/well while that for hybridoma MAb was reported to be 10 ng/well. The affinity constant of recombinant MAb (AR85) towards digoxin was also found to be around ka=3.8×10(7) M(-1) while that for hybridoma MAb (BBA) was reported to be ka=2.6×10(8) M(-1).
-
García-Seoane, Rita; Varela, Zulema; Carballeira, Alejo; Aboal, Jesús R; Fernández, J Ángel
2017-03-01
Temporal trends in Hg concentrations were investigated in primary flight feathers from 319 specimens of three birds of prey: P7 in the northern goshawk (Accipiter gentilis), P6 in the common buzzard (Buteo buteo) and P5 in the tawny owl (Strix aluco). The samples were stored in a regional environmental specimen bank and belonged to specimens which died between 2000 and 2013 in Galicia (NW Spain). We would expect to see a decline in Hg concentrations across the study period, as data of atmospheric emissions show a gradual reduction of this pollutant in Europe in the last two decades. The study did not reveal any temporal pattern in Hg concentrations of feathers in any of the three species for the study period, may be due to the persistence of Hg in the environment, but showed a low level of contamination by this metal in the study area. In addition, the results show high intra-specific, as well as, inter-annual and inter-specific variability in data, mainly attributed to the level of exposure of the raptors to this pollutant and to the biomagnification process of Hg through food chains. These findings indicate that the high variability can be a limiting factor in the use of raptors for biomonitoring temporal patterns of Hg, but nevertheless, the technique provides qualitative information about the amount of Hg that reach the top of the terrestrial food chains.
-
Assessment of Protein Side-Chain Conformation Prediction Methods in Different Residue Environments
Peterson, Lenna X.; Kang, Xuejiao; Kihara, Daisuke
2016-01-01
Computational prediction of side-chain conformation is an important component of protein structure prediction. Accurate side-chain prediction is crucial for practical applications of protein structure models that need atomic detailed resolution such as protein and ligand design. We evaluated the accuracy of eight side-chain prediction methods in reproducing the side-chain conformations of experimentally solved structures deposited to the Protein Data Bank. Prediction accuracy was evaluated for a total of four different structural environments (buried, surface, interface, and membrane-spanning) in three different protein types (monomeric, multimeric, and membrane). Overall, the highest accuracy was observed for buried residues in monomeric and multimeric proteins. Notably, side-chains at protein interfaces and membrane-spanning regions were better predicted than surface residues even though the methods did not all use multimeric and membrane proteins for training. Thus, we conclude that the current methods are as practically useful for modeling protein docking interfaces and membrane-spanning regions as for modeling monomers. PMID:24619909
-
Nursing home cost and ownership type: evidence of interaction effects.
Arling, G; Nordquist, R H; Capitman, J A
1987-06-01
Due to steadily increasing public expenditures for nursing home care, much research has focused on factors that influence nursing home costs, especially for Medicaid patients. Nursing home cost function studies have typically used a number of predictor variables in a multiple regression analysis to determine the effect of these variables on operating cost. Although several authors have suggested that nursing home ownership types have different goal orientations, not necessarily based on economic factors, little attention has been paid to this issue in empirical research. In this study, data from 150 Virginia nursing homes were used in multiple regression analysis to examine factors accounting for nursing home operating costs. The context of the study was the Virginia Medicaid reimbursement system, which has intermediate care and skilled nursing facility (ICF and SNF) facility-specific per diem rates, set according to facility cost histories. The analysis revealed interaction effects between ownership and other predictor variables (e.g., percentage Medicaid residents, case mix, and region), with predictor variables having different effects on cost depending on ownership type. Conclusions are drawn about the goal orientations and behavior of chain-operated, individual for-profit, and public and nonprofit facilities. The implications of these findings for long-term care reimbursement policies are discussed.
-
Nursing home cost and ownership type: evidence of interaction effects.
Arling, G; Nordquist, R H; Capitman, J A
1987-01-01
Due to steadily increasing public expenditures for nursing home care, much research has focused on factors that influence nursing home costs, especially for Medicaid patients. Nursing home cost function studies have typically used a number of predictor variables in a multiple regression analysis to determine the effect of these variables on operating cost. Although several authors have suggested that nursing home ownership types have different goal orientations, not necessarily based on economic factors, little attention has been paid to this issue in empirical research. In this study, data from 150 Virginia nursing homes were used in multiple regression analysis to examine factors accounting for nursing home operating costs. The context of the study was the Virginia Medicaid reimbursement system, which has intermediate care and skilled nursing facility (ICF and SNF) facility-specific per diem rates, set according to facility cost histories. The analysis revealed interaction effects between ownership and other predictor variables (e.g., percentage Medicaid residents, case mix, and region), with predictor variables having different effects on cost depending on ownership type. Conclusions are drawn about the goal orientations and behavior of chain-operated, individual for-profit, and public and nonprofit facilities. The implications of these findings for long-term care reimbursement policies are discussed. PMID:3301746
-
A comparative genomic hybridization study in a 46,XX male.
Rigola, M Angels; Carrera, Marta; Ribas, Isabel; Egozcue, Josep; Miró, Rosa; Fuster, Carme
2002-07-01
To identify Y chromosome material in an azoospermic male with an XX karyotype. Case report. Faculty of medicine and Centro de Patologia Celular (CPC) medical center. A 33-year-old man with infertility. G-banding, fluorescence in situ hybridization (FISH), polymerase chain reaction (PCR), and comparative genomic hybridization (CGH). FISH for X and Y chromosomes, PCR for the SRYgene and amelogenin gene in the Xp (AMGX) and (AMGY), and losses or gains with CGH. FISH analysis using X and Y chromosome-specific probes showed an X chromosome containing Y chromosome sequences on the top of the short arm; this Y chromosome region was not visible by conventional cytogenetic analysis. PCR amplification of DNA showed the presence of the sex-determining region of the Y chromosome (SRY) and the amelogenin gene in the pseudoautosomal boundary of the X chromosome (AMGX). CGH confirmed the presence of the chromosome region Yp11.2-pter and detected the presence of the two otherwise normal X chromosomes. The two Xpter (XPAR1) pseudoautosomal regions present in this XX male suggest the need to reevaluate XX males using CGH and PCR to characterize the clinical variability in XX males due to genes other than those located on the Y chromosome.
-
A full lipid biomarker based record from Lake Challa, Tanzania
NASA Astrophysics Data System (ADS)
Blaga, C. I.; de Leeuw, J. W.; Verschuren, D.; Sinninghe Damsté1, J. S.
2012-04-01
The climate of the regions surrounding the Indian Ocean - East Africa, Arabian and Indian peninsulas - is strongly dominated by the dynamics of the seasonal monsoon. To understand the long and short term driving forces behind the natural climatic variability in this region it is highly important to reconstruct climatic changes in the past and, thereby, predict future changes taking into account also anthropogenic activities. Most low latitude locations lack continuous, highly resolved continental records with good age control. From the few existing records acquired from tropical glacier ice, cave stalagmites and fossil diatoms a thorough understanding of the climatic variations reflected (rainfall and drought or temperature and its effect on precipitation) is scanty. Chemically stratified crater lakes accumulate high-quality climate-proxy records as shown in very recent studies done on the continuous and finely laminated sediment record of Lake Challa situated on the lower East slope of Mt. Kilimanjaro (Verschuren et al. 2009; Wolff et al. 2011). The unique location of this lake in equatorial East Africa implies that the climate variability is influenced by the Indian Ocean and not by the Atlantic due to the Congo Air Boundary (Thierney et al. 2011). The objective of this study is to fully explore the biomarker content of the Lake Challa sedimentary record already characterized by an excellent time resolution and chronology. Various normal chain lipids (n-alkanes, n-fatty acids, n-alcohols), sterols, long-chain diols, triterpenoids and glycolipids in sedimentary organic matter, were determined in their solvent-extractable (free) and saponification-released forms (bound). The changing composition of organic matter content from the investigated lake is used as a framework to trace palaeo-humidity, terrestrial input, algal input, temperature in sediment traps and underlying sediments of Lake Challa to further our palaeo-environmental knowledge based on GDGT's and alkanes (Sinninghe Damsté et al. 2009, 2011).
-
He, Xianzhi; Zhang, Lei; Liu, Pengchong; Liu, Li; Deng, Hui; Huang, Jinhai
2015-03-01
Staphylococcal enterotoxins (SEs) produced by Staphylococcus aureus have increasingly given rise to human health and food safety. Genetically engineered small molecular antibody is a useful tool in immuno-detection and treatment for clinical illness caused by SEs. In this study, we constructed the V(L)-V(H) tail-parallel genetically engineered antibody against SEs by using the repertoire of rearranged germ-line immunoglobulin variable region genes. Total RNA were extracted from six hybridoma cell lines that stably express anti-SEs antibodies. The variable region genes of light chain (V(L)) and heavy chain (V(H)) were cloned by reverse transcription PCR, and their classical murine antibody structure and functional V(D)J gene rearrangement were analyzed. To construct the eukaryotic V(H)-V(L) tail-parallel co-expression vectors based on the "5'-V(H)-ivs-IRES-V(L)-3'" mode, the ivs-IRES fragment and V(L) genes were spliced by two-step overlap extension PCR, and then, the recombined gene fragment and V(H) genes were inserted into the pcDNA3.1(+) expression vector sequentially. And then the constructed eukaryotic expression clones termed as p2C2HILO and p5C12HILO were transfected into baby hamster kidney 21 cell line, respectively. Two clonal cell lines stably expressing V(L)-V(H) tail-parallel antibodies against SEs were obtained, and the antibodies that expressed intracytoplasma were evaluated by enzyme-linked immunosorbent assay, immunofluorescence assay, and flow cytometry. SEs can stimulate the expression of some chemokines and chemokine receptors in porcine IPEC-J2 cells; mRNA transcription level of four chemokines and chemokine receptors can be blocked by the recombinant SE antibody prepared in this study. Our results showed that it is possible to get functional V(L)-V(H) tail-parallel genetically engineered antibodies in same vector using eukaryotic expression system.
-
Rapid isolation of IgNAR variable single-domain antibody fragments from a shark synthetic library.
Shao, Cui-Ying; Secombes, Chris J; Porter, Andrew J
2007-01-01
The immunoglobulin isotype IgNAR (Novel Antigen Receptor) was discovered in the serum of the nurse shark (Ginglymostoma cirratum) and wobbegong shark (Orectolobus maculates) as a homodimer of two protein chains, each composed of a single variable domain (V) domain and five constant domains. The IgNAR variable domain contains an intact antigen-binding site and functions as an independent domain able to react to antigen with both high specificity and affinity. Here we describe the successful construction of a synthetic phage-displayed library based upon a single anti-lysozyme clone HEL-5A7 scaffold, which was previously selected from an immune IgNAR variable domain library. The complementarity-determining region 3 (CDR3) loop of this clone was varied in both length and composition and the derived library was used to pan against two model proteins, lysozyme and leptin. A single anti-lysozyme clone (Ly-X20) and anti-leptin clone (Lep-12E1) were selected for further study. Both clones were shown to be functionally expressed in Escherichia coli, extremely thermostable and bind to corresponding antigens specifically. The results here demonstrate that a synthetic IgNAR variable domain library based on a single framework scaffold can be used as a route to generate antigen binders quickly, easily and without the need of immunization.
-
What is the Effect of Interannual Hydroclimatic Variability on Water Supply Reservoir Operations?
NASA Astrophysics Data System (ADS)
Galelli, S.; Turner, S. W. D.
2015-12-01
Rather than deriving from a single distribution and uniform persistence structure, hydroclimatic data exhibit significant trends and shifts in their mean, variance, and lagged correlation through time. Consequentially, observed and reconstructed streamflow records are often characterized by features of interannual variability, including long-term persistence and prolonged droughts. This study examines the effect of these features on the operating performance of water supply reservoirs. We develop a Stochastic Dynamic Programming (SDP) model that can incorporate a regime-shifting climate variable. We then compare the performance of operating policies—designed with and without climate variable—to quantify the contribution of interannual variability to standard policy sub-optimality. The approach uses a discrete-time Markov chain to partition the reservoir inflow time series into small number of 'hidden' climate states. Each state defines a distinct set of inflow transition probability matrices, which are used by the SDP model to condition the release decisions on the reservoir storage, current-period inflow and hidden climate state. The experimental analysis is carried out on 99 hypothetical water supply reservoirs fed from pristine catchments in Australia—all impacted by the Millennium drought. Results show that interannual hydroclimatic variability is a major cause of sub-optimal hedging decisions. The practical import is that conventional optimization methods may misguide operators, particularly in regions susceptible to multi-year droughts.
-
The AID enzyme induces class switch recombination in fibroblasts.
Okazaki, Il-mi; Kinoshita, Kazuo; Muramatsu, Masamichi; Yoshikawa, Kiyotsugu; Honjo, Tasuku
2002-03-21
The switch of the immunoglobulin isotype from IgM to IgG, IgE or IgA is mediated by class switch recombination (CSR). CSR changes the immunoglobulin heavy chain constant region (CH) gene from Cmu to one of the other CH genes. Somatic hypermutation introduces massive numbers of point mutations in the immunoglobulin variable (V) region gene, giving rise to immunoglobulin with higher affinity. Activation-induced cytidine deaminase (AID), a putative RNA-editing cytidine deaminase, is expressed strictly in activated B cells and is indispensable in both CSR and somatic hypermutation. But the exact function of AID is unknown. Here we show that ectopic expression of AID induces CSR in an artificial switch construct in fibroblasts at a level comparable to that in stimulated B cells. Sequences around recombination junctions in the artificial substrate have features similar to endogenous CSR junctions. Furthermore, AID-induced CSR in fibroblasts is dependent on transcription of the target S region, as shown in endogenous CSR in B cells. The results show that AID is the only B-cell-specific factor required for initiation of the CSR reaction in the activated locus.
-
The ARPAL operational high resolution Poor Man's Ensemble, description and validation
NASA Astrophysics Data System (ADS)
Corazza, Matteo; Sacchetti, Davide; Antonelli, Marta; Drofa, Oxana
2018-05-01
The Meteo Hydrological Functional Center for Civil Protection of the Environmental Protection Agency of the Liguria Region is responsible for issuing forecasts primarily aimed at the Civil Protection needs. Several deterministic high resolution models, run every 6 or 12 h, are regularly used in the Center to elaborate weather forecasts at short to medium range. The Region is frequently affected by severe flash floods over its very small basins, characterized by a steep orography close to the sea. These conditions led the Center in the past years to pay particular attention to the use and development of high resolution model chains for explicit simulation of convective phenomena. For years, the availability of several models has been used by the forecasters for subjective analyses of the potential evolution of the atmosphere and of its uncertainty. More recently, an Interactive Poor Man's Ensemble has been developed, aimed at providing statistical ensemble variables to help forecaster's evaluations. In this paper the structure of this system is described and results are validated using the regional dense ground observational network.
-
Craniofacial Characteristics of Thalassemia Major Patients
Karakas, Sacide; Tellioglu, Ayfer Metin; Bilgin, Mehmet; Omurlu, Imran Kurt; Caliskan, Sercin; Coskun, Salih
2016-01-01
Objective: Thalassemias major are the most common autosomal recessive disorders; they are characterized by anomalies in the synthesis of the beta chains of hemoglobin and are often associated with varying degrees of craniofacial anomalies. The purpose of this study was to evaluate the craniofacial dimensions of β-thalassemia patients and to identify differences by comparing them to those of a control group. Materials and Methods: The study comprised 43 thalassemia major patients and 26 age- and sex- matched healthy control subjects. Anthropometric measurements were performed in six different craniofacial regions (head, face, nose, mouth, eyes, and ears); a total of 23 craniofacial variables were measured. Results: Craniofacial measurements in the regions of the face, nose, lips and mouth, and ears in the thalassemia major patient group yielded statistically significant differences compared to those in the control group (p<0.05). However, no statistically significant differences were observed in the measurements of the head and eye regions. Conclusion: The study increased our understanding of the craniofacial anatomy of thalassemia major patients and enabled us to obtain quantitative results. PMID:28149147
-
New Herbig-Haro objects in star-forming regions
NASA Technical Reports Server (NTRS)
Reipurth, BO; Graham, J. A.
1988-01-01
A list of 25 new Herbig-Haro objects, HH 58 to HH 82, in the Orion molecular clouds and in southern molecular cloud complexes has been compiled. CCD images in the S II 6717, 6731 forbidden lines are presented for the objects, together with a few spectra and some IR observations. The individual objects and, when identified, their energy sources are discussed. HH 65 is located in the red lobe of the bipolar outflow associated with the highly variable reflection nebula Re 50. HH 67 is a 22-arcsec long sinusoidal jet. HH 68/69 consists of a long, linear chain of four HH knots. HH 72 emerges from a 120-solar luminosity IRAS source embedded in a Bok globule. HH 79 is the first HH object discovered in the Ophiuchus clouds. HH 80/81 in Sagittarius are among the brightest HH objects known, have complex velocities, high excitation conditions and emerge from a 6000-solar luminosity young B-star. HH 82 is associated with the bright variable star S Coronae Australis.
-
Can we detect oceanic biodiversity hotspots from space?
De Monte, Silvia; Soccodato, Alice; Alvain, Séverine; d'Ovidio, Francesco
2013-10-01
Understanding the variability of marine biodiversity is a central issue in microbiology. Current observational programs are based on in situ studies, but their implementation at the global scale is particularly challenging, owing to the ocean extent, its temporal variability and the heterogeneity of the data sources on which compilations are built. Here, we explore the possibility of identifying phytoplanktonic biodiversity hotspots from satellite. We define a Shannon entropy index based on patchiness in ocean color bio-optical anomalies. This index provides a high resolution (1 degree) global coverage. It shows a relation to temperature and mid-latitude maxima in accordance with those previously evidenced in microbiological biodiversity model and observational studies. Regional maxima are in remarkable agreement with several known biodiversity hotspots for plankton organisms and even for higher levels of the marine trophic chain, as well as with some in situ planktonic biodiversity estimates (from Atlantic Meridional Transect cruise). These results encourage to explore marine biodiversity with a coordinated effort of the molecular, ecological and remote sensing communities.
-
First report of genotype #65 of Toxoplasma gondii in pigs.
Samico-Fernandes, Erika Fernanda Torres; de Melo, Renata Pimentel Bandeira; de Cássia Peixoto Kim, Pomy; de Almeida, Jonatas Campos; de Barros, Luiz Daniel; Garcia, João Luis; da Silva, Jean Carlos Ramos; Mota, Rinaldo Aparecido
2015-10-01
The aim of the present study was to isolate and genotype Toxoplasma gondii from pigs slaughtered for human consumption in northeastern Brazil. Indirect immunofluorescence antibody test (IFAT) was used to screen positive pigs. Tissues samples of animals with antibody titers ≥64 were submitted to bioassay in mice. One isolate of T. gondii was obtained, and the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique, using 11 markers (SAG1, SAG2, altSAG2, SAG3, BTUB, GRA6, c228, c292, L358, PK1, and APICO), was applied to evaluate the genetic variability. DNA from reference strains was used as a positive control. By means of genetic analysis, genotype ToxoDB #65 was identified, which is considered an atypical strain. This is the first record of genotype #65 in pigs. Thus, further studies in this region are necessary to determine the genetic variability of T. gondii in pigs and possible impact on public health.
-
NASA Astrophysics Data System (ADS)
Jury, Mark R.
2016-11-01
Climate variability in the eastern Antilles island chain is analyzed via principal component analysis of high-resolution monthly rainfall in the period 1981-2013. The second mode reflecting higher rainfall in July-October season between Martinique and Grenada is the focus of this study. Higher rainfall corresponds with a weakened trade wind and boundary current along the southern edge of the Caribbean. This quells the coastal upwelling off Venezuela and builds the freshwater plume east of Trinidad. There is corresponding upper easterly wind flow that intensifies passing tropical waves. During a storm event over the Antilles on 4-5 October 2010, there was inflow from east of Guyana where low salinity and high sea temperatures enable surplus latent heat fluxes. A N-S convective rain band forms ˜500 km east of the cyclonic vortex. Many features at the weather timescale reflect the seasonal correlation and composite difference maps and El Nino Southern Oscillation (ENSO) modulation of oceanic inter-basin transfers.
-
Liberti, Assunta; Melillo, Daniela; Zucchetti, Ivana; Natale, Lenina; Dishaw, Larry J.; Litman, Gary W.; De Santis, Rosaria; Pinto, Maria Rosaria
2014-01-01
Variable region-containing chitin-binding proteins (VCBPs) are secreted, immune-type molecules that have been described in both amphioxus, a cephalochordate, and sea squirt, Ciona intestinalis, a urochordate. In adult Ciona, VCBP-A, -B and -C are expressed in hemocytes and the cells of the gastrointestinal tract. VCBP-C binds bacteria in the stomach lumen and functions as an opsonin in vitro. In the present paper the expression of VCBPs has been characterized during development using in situ hybridization, immunohistochemical staining and quantitative polymerase chain reaction (qPCR) technologies. The expression of VCBP-A and -C is detected first in discrete areas of larva endoderm and becomes progressively localized during differentiation in the stomach and intestine, marking the development of gut tracts. In “small adults” (1–2 cm juveniles) expression of VCBP-C persists and VCBP-A gradually diminishes, ultimately replaced by expression of VCBP-B. The expression of VCBP-A and -C in stage 7–8 juveniles, at which point animals have already started feeding, is influenced significantly by challenge with either Gram-positive or -negative bacteria. A potential role for VCBPs in gut-microbiota interactions and homeostasis is indicated. PMID:24788831
-
A dual host vector for Fab phage display and expression of native IgG in mammalian cells.
Tesar, Devin; Hötzel, Isidro
2013-10-01
A significant bottleneck in antibody discovery by phage display is the transfer of immunoglobulin variable regions from phage clones to vectors that express immunoglobulin G (IgG) in mammalian cells for screening. Here, we describe a novel phagemid vector for Fab phage display that allows expression of native IgG in mammalian cells without sub-cloning. The vector uses an optimized mammalian signal sequence that drives robust expression of Fab fragments fused to an M13 phage coat protein in Escherichia coli and IgG expression in mammalian cells. To allow the expression of Fab fragments fused to a phage coat protein in E.coli and full-length IgG in mammalian cells from the same vector without sub-cloning, the sequence encoding the phage coat protein was embedded in an optimized synthetic intron within the immunoglobulin heavy chain gene. This intron is removed from transcripts in mammalian cells by RNA splicing. Using this vector, we constructed a synthetic Fab phage display library with diversity in the heavy chain only and selected for clones binding different antigens. Co-transfection of mammalian cells with DNA from individual phage clones and a plasmid expressing the invariant light chain resulted in the expression of native IgG that was used to assay affinity, ligand blocking activity and specificity.
-
Shaffer, Justin F.; Kier, William M.
2012-01-01
SUMMARY The speed of muscle contraction is largely controlled at the sarcomere level by the ATPase activity of the motor protein myosin. Differences in amino acid sequence in catalytically important regions of myosin yield different myosin isoforms with varying ATPase activities and resulting differences in cross-bridge cycling rates and interfilamentary sliding velocities. Modulation of whole-muscle performance by changes in myosin isoform ATPase activity is regarded as a universal mechanism to tune contractile properties, especially in vertebrate muscles. Invertebrates such as squid, however, may exhibit an alternative mechanism to tune contractile properties that is based on differences in muscle ultrastructure, including variable myofilament and sarcomere lengths. To determine definitively whether contractile properties of squid muscles are regulated via different myosin isoforms (i.e. different ATPase activities), the nucleotide and amino acid sequences of the myosin heavy chain from the squid Doryteuthis pealeii were determined from the mantle, arm, tentacle, fin and funnel retractor musculature. We identified three myosin heavy chain isoforms in squid muscular tissues, with differences arising at surface loop 1 and the carboxy terminus. All three isoforms were detected in all five tissues studied. These results suggest that the muscular tissues of D. pealeii express identical myosin isoforms, and it is likely that differences in muscle ultrastructure, not myosin ATPase activity, represent the most important mechanism for tuning contractile speeds. PMID:22189767
-
Newton, Paul K.; Mason, Jeremy; Bethel, Kelly; Bazhenova, Lyudmila; Nieva, Jorge; Norton, Larry; Kuhn, Peter
2013-01-01
The classic view of metastatic cancer progression is that it is a unidirectional process initiated at the primary tumor site, progressing to variably distant metastatic sites in a fairly predictable, though not perfectly understood, fashion. A Markov chain Monte Carlo mathematical approach can determine a pathway diagram that classifies metastatic tumors as ‘spreaders’ or ‘sponges’ and orders the timescales of progression from site to site. In light of recent experimental evidence highlighting the potential significance of self-seeding of primary tumors, we use a Markov chain Monte Carlo (MCMC) approach, based on large autopsy data sets, to quantify the stochastic, systemic, and often multi-directional aspects of cancer progression. We quantify three types of multi-directional mechanisms of progression: (i) self-seeding of the primary tumor; (ii) re-seeding of the primary tumor from a metastatic site (primary re-seeding); and (iii) re-seeding of metastatic tumors (metastasis re-seeding). The model shows that the combined characteristics of the primary and the first metastatic site to which it spreads largely determine the future pathways and timescales of systemic disease. For lung cancer, the main ‘spreaders’ of systemic disease are the adrenal gland and kidney, whereas the main ‘sponges’ are regional lymph nodes, liver, and bone. Lung is a significant self-seeder, although it is a ‘sponge’ site with respect to progression characteristics. PMID:23447576
-
Ahmed, Belal; Jo, Hongil; Oh, Seung-Jin; Ok, Kang Min
2018-05-15
Four novel transition metal oxyfluorides, [Zn(pz) 3 ][MoO 2 F 4 ]·0.1H 2 O (1), [Zn(pz) 2 F 2 ][Zn(pz) 3 ] 2 [WO 2 F 4 ] 2 (2), [Cd(pz) 4 ][Cd(pz) 4 (H 2 O)][MoO 2 F 4 ] 2 ·0.625H 2 O (3), and [Zn(mpz) 3 ] 2 [MoO 2 F 4 ] 2 (4) (pz = pyrazole; mpz = 3-methyl pyrazole) have been synthesized. Compounds 1 and 4 contain helical chains. Compound 2 accommodates zigzag chains, and compound 3 has quasi-one-dimensional linear chains. The variable chain structures are found to be attributable to the different structure-directing anionic groups and hydrogen bonding interactions. Compound 4 crystallized in the noncentrosymmetric (NCS) polar space group, Pna2 1 , is nonphase-matchable (Type I), and reveals a moderate second-harmonic-generation (SHG) efficiency (10 × α-SiO 2 ). The observed SHG efficiency of compound 4 is due to the small net polarization occurring from the arrangement of ZnN 3 F 2 trigonal bipyramids. Spectroscopic and thermal characterizations along with calculations for the title materials are reported.
-
Making Supply Chains Resilient to Floods Using a Bayesian Network
NASA Astrophysics Data System (ADS)
Haraguchi, M.
2015-12-01
Natural hazards distress the global economy by disrupting the interconnected supply chain networks. Manufacturing companies have created cost-efficient supply chains by reducing inventories, streamlining logistics and limiting the number of suppliers. As a result, today's supply chains are profoundly susceptible to systemic risks. In Thailand, for example, the GDP growth rate declined by 76 % in 2011 due to prolonged flooding. Thailand incurred economic damage including the loss of USD 46.5 billion, approximately 70% of which was caused by major supply chain disruptions in the manufacturing sector. Similar problems occurred after the Great East Japan Earthquake and Tsunami in 2011, the Mississippi River floods and droughts during 2011 - 2013, and Hurricane Sandy in 2012. This study proposes a methodology for modeling supply chain disruptions using a Bayesian network analysis (BNA) to estimate expected values of countermeasures of floods, such as inventory management, supplier management and hard infrastructure management. We first performed a spatio-temporal correlation analysis between floods and extreme precipitation data for the last 100 years at a global scale. Then we used a BNA to create synthetic networks that include variables associated with the magnitude and duration of floods, major components of supply chains and market demands. We also included decision variables of countermeasures that would mitigate potential losses caused by supply chain disruptions. Finally, we conducted a cost-benefit analysis by estimating the expected values of these potential countermeasures while conducting a sensitivity analysis. The methodology was applied to supply chain disruptions caused by the 2011 Thailand floods. Our study demonstrates desirable typical data requirements for the analysis, such as anonymized supplier network data (i.e. critical dependencies, vulnerability information of suppliers) and sourcing data(i.e. locations of suppliers, and production rates and volume), and data from previous experiences (i.e. companies' risk mitigation strategy decisions).
-
Zhang, Ling; Nan, Zhuotong; Xu, Yi; Li, Shuo
2016-01-01
Land use change and climate variability are two key factors impacting watershed hydrology, which is strongly related to the availability of water resources and the sustainability of local ecosystems. This study assessed separate and combined hydrological impacts of land use change and climate variability in the headwater region of a typical arid inland river basin, known as the Heihe River Basin, northwest China, in the recent past (1995–2014) and near future (2015–2024), by combining two land use models (i.e., Markov chain model and Dyna-CLUE) with a hydrological model (i.e., SWAT). The potential impacts in the near future were explored using projected land use patterns and hypothetical climate scenarios established on the basis of analyzing long-term climatic observations. Land use changes in the recent past are dominated by the expansion of grassland and a decrease in farmland; meanwhile the climate develops with a wetting and warming trend. Land use changes in this period induce slight reductions in surface runoff, groundwater discharge and streamflow whereas climate changes produce pronounced increases in them. The joint hydrological impacts are similar to those solely induced by climate changes. Spatially, both the effects of land use change and climate variability vary with the sub-basin. The influences of land use changes are more identifiable in some sub-basins, compared with the basin-wide impacts. In the near future, climate changes tend to affect the hydrological regimes much more prominently than land use changes, leading to significant increases in all hydrological components. Nevertheless, the role of land use change should not be overlooked, especially if the climate becomes drier in the future, as in this case it may magnify the hydrological responses. PMID:27348224
-
Zhang, Ling; Nan, Zhuotong; Xu, Yi; Li, Shuo
2016-01-01
Land use change and climate variability are two key factors impacting watershed hydrology, which is strongly related to the availability of water resources and the sustainability of local ecosystems. This study assessed separate and combined hydrological impacts of land use change and climate variability in the headwater region of a typical arid inland river basin, known as the Heihe River Basin, northwest China, in the recent past (1995-2014) and near future (2015-2024), by combining two land use models (i.e., Markov chain model and Dyna-CLUE) with a hydrological model (i.e., SWAT). The potential impacts in the near future were explored using projected land use patterns and hypothetical climate scenarios established on the basis of analyzing long-term climatic observations. Land use changes in the recent past are dominated by the expansion of grassland and a decrease in farmland; meanwhile the climate develops with a wetting and warming trend. Land use changes in this period induce slight reductions in surface runoff, groundwater discharge and streamflow whereas climate changes produce pronounced increases in them. The joint hydrological impacts are similar to those solely induced by climate changes. Spatially, both the effects of land use change and climate variability vary with the sub-basin. The influences of land use changes are more identifiable in some sub-basins, compared with the basin-wide impacts. In the near future, climate changes tend to affect the hydrological regimes much more prominently than land use changes, leading to significant increases in all hydrological components. Nevertheless, the role of land use change should not be overlooked, especially if the climate becomes drier in the future, as in this case it may magnify the hydrological responses.
-
Precipitation dynamics and chemical properties in tropical mountain forests of Ecuador
NASA Astrophysics Data System (ADS)
Rollenbeck, R.; Fabian, P.; Bendix, J.
2006-01-01
Terrestrial ecosystems in southern Ecuador are strongly affected by interannual climate variations. This holds especially true for the episodic El Niño events, which cause above-normal precipitation in the coastal region of Ecuador and below normal values in the eastern provinces of the Amazon basin (Bendix, 1999). For the transitional zone between these two extremes, which consists mainly of the andean slopes and larger interandean basins the effect on interannual climate variability is not well known. The PREDICT project monitors regional climate in the provinces of Loja and Zamora-Chinchipe (4° S/79° W), where a strong gradients of precipitation are observed. Between the eastern slopes of the Cordillera Real and the dry valley of Catamayo, which are only 70km apart, rain totals drop from over 4000 mm to only 300 mm per year. These two extremes represent the both sides of the Andean mountain chain and are completely covered by the study area, which is 120 km in diameter. Methods used are a combination of point measurements (climate stations) and remote sensing devices (weather radar, satellite imagery), which enable a high-resolution real-time observation of rain distribution and underlying processes. By this, ideal conditions are given to monitor a potential shift of the transition zone between below-average and above-average rainfall situated in this region, if another ENSO-anomaly occurs. Furthermore variability of atmospheric nutrient inputs is analysed within the scope of the project, to assess further impacts on this ecosystem.
-
Zhao, Yaofeng; Cui, Huiting; Whittington, Camilla M; Wei, Zhiguo; Zhang, Xiaofeng; Zhang, Ziding; Yu, Li; Ren, Liming; Hu, Xiaoxiang; Zhang, Yaping; Hellman, Lars; Belov, Katherine; Li, Ning; Hammarström, Lennart
2009-09-01
The evolutionary origins of mammalian immunoglobulin H chain isotypes (IgM, IgD, IgG, IgE, and IgA) are still incompletely understood as these isotypes differ considerably in structure and number from their counterparts in nonmammalian tetrapods. We report in this study that the platypus (Ornithorhynchus anatinus) Ig H chain constant region gene locus contains eight Ig encoding genes, which are arranged in an mu-delta-omicron-gamma2-gamma1-alpha1-epsilon-alpha2 order, spanning a total of approximately 200 kb DNA, encoding six distinct isotypes. The omicron (omicron for Ornithorhynchus) gene encodes a novel Ig H chain isotype that consists of four constant region domains and a hinge, and is structurally different from any of the five known mammalian Ig classes. This gene is phylogenetically related to upsilon (epsilon) and gamma, and thus appears to be a structural intermediate between these two genes. The platypus delta gene encodes ten heavy chain constant region domains, lacks a hinge region and is similar to IgD in amphibians and fish, but strikingly different from that in eutherian mammals. The platypus Ig H chain isotype repertoire thus shows a unique combination of genes that share similarity both to those of nonmammalian tetrapods and eutherian animals and demonstrates how phylogenetically informative species can be used to reconstruct the evolutionary history of functionally important genes.
-
Plasticization effect of triacetin on structure and properties of starch ester film.
Zhu, Jie; Li, Xiaoxi; Huang, Chen; Chen, Ling; Li, Lin
2013-05-15
The aim of this work was to evaluate the plasticizing effect of triacetin on the structure and properties of starch ester film and further establish the structure-property relationships. The presence of triacetin resulted in multiple structure changes of the film. The mobility of macromolecular chain was increased to form scattered crystallite during the film formation process. The amorphous region was enlarged to contain more triacetin squeezed from crystalline region. The plasticization of triacetin and restriction of crystallite oppositely influenced the mobility of macromolecular chains in different regions. The thermal stability of triacetin changed along with its fluctuant interaction with macromolecules. Comparatively, the enhanced ether bond and the restriction from crystalline regions on the mobility of the amorphous chain consequently improved the thermal stability of the film matrix. The interaction between triacetin and starch ester was essential to film forming but unexpectedly lowered the triacetin stability. Copyright © 2013 Elsevier Ltd. All rights reserved.
-
Ben Ali, Sina-Elisabeth; Madi, Zita Erika; Hochegger, Rupert; Quist, David; Prewein, Bernhard; Haslberger, Alexander G; Brandes, Christian
2014-10-31
Genetic mutations must be avoided during the production and use of seeds. In the European Union (EU), Directive 2001/18/EC requires any DNA construct introduced via transformation to be stable. Establishing genetic stability is critical for the approval of genetically modified organisms (GMOs). In this study, genetic stability of two GMOs was examined using high resolution melting (HRM) analysis and real-time polymerase chain reaction (PCR) employing Scorpion primers for amplification. The genetic variability of the transgenic insert and that of the flanking regions in a single oilseed rape variety (GT73) and a stacked maize (MON88017×MON810) was studied. The GT73 and the 5' region of MON810 showed no instabilities in the examined regions. However; two out of 100 analyzed samples carried a heterozygous point mutation in the 3' region of MON810 in the stacked variety. These results were verified by direct sequencing of the amplified PCR products as well as by sequencing of cloned PCR fragments. The occurrence of the mutation suggests that the 5' region is more suitable than the 3' region for the quantification of MON810. The identification of the single nucleotide polymorphism (SNP) in a stacked event is in contrast to the results of earlier studies of the same MON810 region in a single event where no DNA polymorphism was found.
-
Liu, Lihong; Wang, Weiming; Matz, Julie; Ye, Chaobaihui; Bracq, Lucie; Delon, Jerome; Kimata, Jason T.; Chen, Zhiwei
2016-01-01
ABSTRACT The variable regions (VHHs) of two heavy chain-only antibodies, JM2 and JM4, from llamas that have been immunized with a trimeric gp140 bound to a CD4 mimic have been recently isolated (here referred to as VHH JM2 and VHH JM4, respectively). JM2 binds the CD4-binding site of gp120 and neutralizes HIV-1 strains from subtypes B, C, and G. JM4 binds gp120 and neutralizes HIV-1 strains from subtypes A, B, C, A/E, and G in a CD4-dependent manner. In the present study, we constructed glycosylphosphatidylinositol (GPI)-anchored VHH JM2 and JM4 along with an E4 control and transduced them into human CD4+ cell lines and primary CD4 T cells. We report that by genetically linking the VHHs with a GPI attachment signal, VHHs are targeted to the lipid rafts of the plasma membranes. Expression of GPI-VHH JM4, but not GPI-VHH E4 and JM2, on the surface of transduced TZM.bl cells potently neutralizes multiple subtypes of HIV-1 isolates, including tier 2 or 3 strains, transmitted founders, quasispecies, and soluble single domain antibody (sdAb) JM4-resistant viruses. Moreover, transduction of CEMss-CCR5 cells with GPI-VHH JM4, but not with GPI-VHH E4, confers resistance to both cell-free and T cell-T cell transmission of HIV-1 and HIV-1 envelope-mediated fusion. Finally, GPI-VHH JM4-transduced human primary CD4 T cells efficiently resist both cell-free and T cell-T cell transmission of HIV-1. Thus, we conclude that VHH JM4, when targeted to the lipid rafts of the plasma membrane, efficiently neutralizes HIV-1 infection via both cell-free and T cell-T cell transmission. Our findings should have important implications for GPI-anchored antibody-based therapy against HIV-1. IMPORTANCE Lipid rafts are specialized dynamic microdomains of the plasma membrane and have been shown to be gateways for HIV-1 budding as well as entry into T cells and macrophages. In nature, many glycosylphosphatidylinositol (GPI)-anchored proteins localize in the lipid rafts. In the present study, we developed GPI-anchored variable regions (VHHs) of two heavy chain-only antibodies, JM2 and JM4, from immunized llamas. We show that by genetically linking the VHHs with a GPI attachment signal, VHHs are targeted to the lipid rafts of the plasma membranes. GPI-VHH JM4, but not GPI-VHH JM2, in transduced CD4+ cell lines and human primary CD4 T cells not only efficiently blocks diverse HIV-1 strains, including tier 2 or 3 strains, transmitted founders, quasispecies, and soluble sdAb JM4-resistant strains, but also efficiently interferes T cell-T cell transmissions of HIV-1 and HIV-1 envelope-mediated fusion. Our findings should have important implications in GPI-anchored antibody-based therapy against HIV-1. PMID:27654286
-
Uses of monoclonial antibody 8H9
Cheung, Nai-Kong V.
2015-06-23
This invention provides an antibody that binds the same antigen as that of monoclonal antibody 8H9, wherein the heavy chain CDR (Complementary Determining Region)1 comprises NYDIN, heavy chain CDR2 comprises WIFPGDGSTQY, heavy chain CDR3 comprises QTTATWFAY, and the light chain CDR1 comprises RASQSISDYLH, light chain CDR2 comprises YASQSIS, and light chain CDR3 comprises QNGHSFPLT. In another embodiment, there is provided a polypeptide that binds the same antigen as that of monoclonal antibody 8H9, wherein the polypeptide comprises NYDIN, WIFPGDGSTQY, QTTATWFAY, RASQSISDYLH, YASQSIS, and QNGHSFPLT.
-
Expression of immunoglobulin G in human podocytes, and its role in cell viability and adhesion.
Jing, Ziyang; Deng, Hui; Ma, Junfan; Guo, Yanhong; Liang, Yaoxian; Wu, Rui; A, Lata; Geng, Zihan; Qiu, Xiaoyan; Wang, Yue
2018-06-01
Podocyte injury occurs during the initiation and development of numerous forms of glomerular disease, and antibodies targeting podocytes have become a biomarker for diagnosis and monitoring treatment response. Accumulating evidence has suggested that immunoglobulin (Ig) is expressed in non‑B lineage cells, including epithelial cancer cells, myeloid cells and several types of normal cells. The main aim of the present study was to ascertain the expression of IgG in human podocytes and to determine its potential role in cellular bioactivity. The present study detected positive staining for IgG heavy chain (Igγ) and its subtype γ4, and the light chains κ and λ in the cytoplasm or on the membrane by immunofluorescence. In addition, positive bands were detected for Igγ, γ1, γ3, γ4, κ and λ in the lysates of a podocyte cell line by western blotting. Mass spectrometry confirmed IgG1 as an intact tetramer in the culture supernatant. Constant region transcripts of Igγ, γ1, γ3, γ4, κ and λ were identified by reverse transcription‑polymerase chain reaction, and DNA sequencing of these transcripts revealed 96‑99% similarity with Ig mRNAs in the National Center for Biotechnology Information database. Compared with the diverse gene rearrangements from B cell-derived Ig, podocyte‑derived Ig exhibited conservative V(D)J patterns in the variable regions of Igγ and κ chains. Furthermore, the present study investigated the mechanism underlying IgG production in these cells by examining the expression of recombination activating gene (RAG)1, RAG2 and activation‑induced cytidine deaminase. The expression levels of these proteins suggested that podocyte‑derived Ig and traditional Ig may be generated in a similar manner. Furthermore, small interfering RNA‑mediated downregulation of IgG expression reduced podocyte viability and adhesive capabilities. These findings suggested that IgG is expressed in podocytes and that this expression may be associated with podocyte function. Due to its potential biological and clinical significance, this phenomenon warrants further investigation.
-
Nanostructures and dynamics of macromolecules bound to attractive filler surfaces
NASA Astrophysics Data System (ADS)
Koga, Tad; Barkley, Deborah; Jiang, Naisheng; Endoh, Maya; Masui, Tomomi; Kishimoto, Hiroyuki; Nagao, Michihiro; Satija, Sushil; Taniguchi, Takashi
We report in-situ nanostructures and dynamics of polybutadiene (PB) chains bound to carbon black (CB) fillers (the so-called ``bound polymer layer (BPL)'') in a good solvent. The BPL on the CB fillers were extracted by solvent leaching of a CB-filled PB compound and subsequently dispersed in deuterated toluene to label the BPL for small-angle neutron scattering and neutron spin echo techniques. Intriguingly, the results demonstrate that the BPL is composed of two regions regardless of molecular weights of PB: the inner unswollen region of ~ 0.5 nm thick and outer swollen region where the polymer chains display a parabolic profile with a diffuse tail. This two-layer formation on the filler surface is similar to that reported for polymer chains adsorbed on planar substrates from melts. In addition, the results show that the dynamics of the swollen bound chains can be explained by the so-called ``breathing mode'' and is generalized with the thickness of the swollen BPL. Furthermore, we will discuss how the breathing collective dynamics is affected by the presence of polymer chains in a matrix solution. We acknowledge the financial support from NSF Grant No. CMMI-1332499.
-
Superheavy-element spectroscopy: Correlations along element 115 decay chains
NASA Astrophysics Data System (ADS)
Rudolph, D.; Forsberg, U.; Sarmiento, L. G.; Golubev, P.; Fahlander, C.
2016-05-01
Following a brief summary of the region of the heaviest atomic nuclei yet created in the laboratory, data on more than hundred α-decay chains associated with the production of element 115 are combined to investigate time and energy correlations along the observed decay chains. Several of these are analysed using a new method for statistical assessments of lifetimes in sets of decay chains.
-
Traces of business cycles in credit-rating migrations
Boreiko, Dmitri; Kaniovski, Serguei; Pflug, Georg
2017-01-01
Using migration data of a rating agency, this paper attempts to quantify the impact of macroeconomic conditions on credit-rating migrations. The migrations are modeled as a coupled Markov chain, where the macroeconomic factors are represented by unobserved tendency variables. In the simplest case, these binary random variables are static and credit-class-specific. A generalization treats tendency variables evolving as a time-homogeneous Markov chain. A more detailed analysis assumes a tendency variable for every combination of a credit class and an industry. The models are tested on a Standard and Poor’s (S&P’s) dataset. Parameters are estimated by the maximum likelihood method. According to the estimates, the investment-grade financial institutions evolve independently of the rest of the economy represented by the data. This might be an evidence of implicit too-big-to-fail bail-out guarantee policies of the regulatory authorities. PMID:28426758
-
Traces of business cycles in credit-rating migrations.
Boreiko, Dmitri; Kaniovski, Serguei; Kaniovski, Yuri; Pflug, Georg
2017-01-01
Using migration data of a rating agency, this paper attempts to quantify the impact of macroeconomic conditions on credit-rating migrations. The migrations are modeled as a coupled Markov chain, where the macroeconomic factors are represented by unobserved tendency variables. In the simplest case, these binary random variables are static and credit-class-specific. A generalization treats tendency variables evolving as a time-homogeneous Markov chain. A more detailed analysis assumes a tendency variable for every combination of a credit class and an industry. The models are tested on a Standard and Poor's (S&P's) dataset. Parameters are estimated by the maximum likelihood method. According to the estimates, the investment-grade financial institutions evolve independently of the rest of the economy represented by the data. This might be an evidence of implicit too-big-to-fail bail-out guarantee policies of the regulatory authorities.
-
Sridevi, N. V.; Shukra, A. M.; Neelakantam, B.; Anilkumar, J.; Madhanmohan, M.; Rajan, S.; Dev Chandran
2014-01-01
Recombinant antibody fragments like single chain variable fragments (scFvs) represent an attractive yet powerful alternative to immunoglobulins and hold great potential in the development of clinical diagnostic/therapeutic reagents. Structurally, scFvs are the smallest antibody fragments capable of retaining the antigen-binding capacity of whole antibodies and are composed of an immunoglobulin (Ig) variable light (VL) and variable heavy (VH) chain joined by a flexible polypeptide linker. In the present study, we constructed a scFv against bovine IgA from a hybridoma cell line IL-A71 that secretes a monoclonal antibody against bovine IgA using recombinant DNA technology. The scFv was expressed in Escherichia coli and purified using immobilized metal affinity chromatography (IMAC). The binding activity and specificity of the scFv was established by its non-reactivity toward other classes of immunoglobulins as determined by enzyme-linked immunosorbent assay (ELISA) and immunoblot analysis. Kinetic measurement of the scFv indicated that the recombinant antibody fragment had an affinity in picomolar range toward purified IgA. Furthermore, the scFv was used to develop a sensitive ELISA for the detection of foot and mouth disease virus (FMDV) carrier animals. PMID:24678404
-
Mutual Information and Information Gating in Synfire Chains
Xiao, Zhuocheng; Wang, Binxu; Sornborger, Andrew Tyler; ...
2018-02-01
Here, coherent neuronal activity is believed to underlie the transfer and processing of information in the brain. Coherent activity in the form of synchronous firing and oscillations has been measured in many brain regions and has been correlated with enhanced feature processing and other sensory and cognitive functions. In the theoretical context, synfire chains and the transfer of transient activity packets in feedforward networks have been appealed to in order to describe coherent spiking and information transfer. Recently, it has been demonstrated that the classical synfire chain architecture, with the addition of suitably timed gating currents, can support the gradedmore » transfer of mean firing rates in feedforward networks (called synfire-gated synfire chains—SGSCs). Here we study information propagation in SGSCs by examining mutual information as a function of layer number in a feedforward network. We explore the effects of gating and noise on information transfer in synfire chains and demonstrate that asymptotically, two main regions exist in parameter space where information may be propagated and its propagation is controlled by pulse-gating: a large region where binary codes may be propagated, and a smaller region near a cusp in parameter space that supports graded propagation across many layers.« less
-
Mutual Information and Information Gating in Synfire Chains
DOE Office of Scientific and Technical Information (OSTI.GOV)
Xiao, Zhuocheng; Wang, Binxu; Sornborger, Andrew Tyler
Here, coherent neuronal activity is believed to underlie the transfer and processing of information in the brain. Coherent activity in the form of synchronous firing and oscillations has been measured in many brain regions and has been correlated with enhanced feature processing and other sensory and cognitive functions. In the theoretical context, synfire chains and the transfer of transient activity packets in feedforward networks have been appealed to in order to describe coherent spiking and information transfer. Recently, it has been demonstrated that the classical synfire chain architecture, with the addition of suitably timed gating currents, can support the gradedmore » transfer of mean firing rates in feedforward networks (called synfire-gated synfire chains—SGSCs). Here we study information propagation in SGSCs by examining mutual information as a function of layer number in a feedforward network. We explore the effects of gating and noise on information transfer in synfire chains and demonstrate that asymptotically, two main regions exist in parameter space where information may be propagated and its propagation is controlled by pulse-gating: a large region where binary codes may be propagated, and a smaller region near a cusp in parameter space that supports graded propagation across many layers.« less
-
Dunnick, Wesley A; Shi, Jian; Holden, Victoria; Fontaine, Clinton; Collins, John T
2011-01-01
Germline transcription precedes class switch recombination (CSR). The promoter regions and I exons of these germline transcripts include binding sites for activation- and cytokine-induced transcription factors, and the promoter regions/I exons are essential for CSR. Therefore, it is a strong hypothesis that the promoter/I exons regions are responsible for much of cytokine-regulated, gene-specific CSR. We tested this hypothesis by swapping the germline promoter and I exons for the murine γ1 and γ2a H chain genes in a transgene of the entire H chain C-region locus. We found that the promoter/I exon for γ1 germline transcripts can direct robust IL-4-induced recombination to the γ2a gene. In contrast, the promoter/I exon for the γ2a germline transcripts works poorly in the context of the γ1 H chain gene, resulting in expression of γ1 H chains that is <1% the wild-type level. Nevertheless, the small amount of recombination to the chimeric γ1 gene is induced by IFN-γ. These results suggest that cytokine regulation of CSR, but not the magnitude of CSR, is regulated by the promoter/I exons.
-
2 + 1 Toda chain. I. Inverse scattering method
DOE Office of Scientific and Technical Information (OSTI.GOV)
Lipovskii, V.D.; Shirokov, A.V.
A formal scheme of the inverse scattering method is constructed for the2 + 1 Toda chain in the class of rapidly decreasing Cauchy data. Application of the inverse scattering method to the two-dimensional infinite Toda chain was made difficult by the circumstance that this system is a (2 + 1)-dimensional object, i.e., possesses time and two spatial variables, the role of one of these being played by the chain site number. Because of this, our information about the 2 + 1 Toda chain was limited to a rich set of particular solutions of soliton type obtained in the cycle ofmore » studies by the Darboux transformation method.« less
-
NASA Astrophysics Data System (ADS)
Yang, Jin-Wei; Gao, Yi-Tian; Wang, Qi-Min; Su, Chuan-Qi; Feng, Yu-Jie; Yu, Xin
2016-01-01
In this paper, a fourth-order variable-coefficient nonlinear Schrödinger equation is studied, which might describe a one-dimensional continuum anisotropic Heisenberg ferromagnetic spin chain with the octuple-dipole interaction or an alpha helical protein with higher-order excitations and interactions under continuum approximation. With the aid of auxiliary function, we derive the bilinear forms and corresponding constraints on the variable coefficients. Via the symbolic computation, we obtain the Lax pair, infinitely many conservation laws, one-, two- and three-soliton solutions. We discuss the influence of the variable coefficients on the solitons. With different choices of the variable coefficients, we obtain the parabolic, cubic, and periodic solitons, respectively. We analyse the head-on and overtaking interactions between/among the two and three solitons. Interactions between a bound state and a single soliton are displayed with different choices of variable coefficients. We also derive the quasi-periodic formulae for the three cases of the bound states.
-
Carvalho, S; Caldeira, R L; Simpson, A J; Vidigal, T H
2001-01-01
Freshwater snails belonging to the genus Biomphalaria are intermediate hosts of the trematode Schistosoma mansoni in the Neotropical region and Africa. In Brazil, one subspecies and ten species of Biomphalaria have been identified: B. glabrata, B. tenagophila, B. straminea, B. occidentalis, B. peregrina, B. kuhniana, B. schrammi, B. amazonica, B. oligoza, B. intermedia and B.t. guaibensis. However, only the first three species are found naturally infected with S. mansoni. The classical identification of these planorbids is based on comparison of morphological characteristics of the shell and male and female reproductive organs, which is greatly complicated by the extensive intra-specific variation. Several molecular techniques have been used in studies on the identification, genetic structure as well as phylogenetic relationships between these groups of organisms. Using the randomly amplified polymorphic DNAs (RAPD) analysis we demonstrated that B. glabrata exhibits a remarkable degree of intra-specific polymorphism. Thus, the genetics of the snail host may be more important to the epidemiology of schistosomiasis than those of the parasite itself. Using the simple sequence repeat anchored polymerase chain reaction (SSR-PCR) in intra-populational and intra-specific studies we have demonstrated that snails belonging to the B. straminea complex (B. straminea, B. kuhniana and B. intermedia) clearly presented higher heterogeneity. Using the low stringency polymerase chain reaction (LS-PCR) technique we were able to separate B. glabrata from B. tenagophila and B. tenagophila from B. occidentalis. To separate all Brazilian Biomphalaria species we used the restriction fragment length polymorphism (PCR-RFLP) of the internal transcribed spacer region (ITS) of the DNA gene. The method also proved to be efficient for the specific identification of DNA extracted from snail eggs. Recently we have sequenced the ITS2 region for phylogenetic studies of all Biomphalaria snails from Brazil.
-
Disseminated Mycobacterium intracellulare infection in a broad-snouted caiman Caiman latirostris.
Kik, Marja J L
2013-11-25
A 10 yr old broad-snouted caiman Caiman latirostris from a small Dutch animal park was presented with long-term variable periods of anorexia and weight loss. Blood chemistry showed slightly elevated uric acid levels and low ionised calcium concentration. Ultrasonographical thickening of the intestinal wall in the region of the duodenum was evident. Pathological changes were a thickening of the wall of 90% of the small intestines, enlarged spleen with multifocal white foci and an enlarged light-brown liver. Histopathological lesions consisted of disseminated granulomas in the intestinal wall, the liver and the spleen. Multinucleated giant cells and epitheloid macrophages were abundant. Ziehl-Neelsen staining showed numerous intralesional acid-fast bacteria. Polymerase chain reaction for Mycobacterium intracellulare was positive.
-
Increased Chain Length Promotes Pneumococcal Adherence and Colonization
Rodriguez, Jesse L.; Dalia, Ankur B.
2012-01-01
Streptococcus pneumoniae is a mucosal pathogen that grows in chains of variable lengths. Short-chain forms are less likely to activate complement, and as a consequence they evade opsonophagocytic clearance more effectively during invasive disease. When grown in human nasal airway surface fluid, pneumococci exhibited both short- and long-chain forms. Here, we determined whether longer chains provide an advantage during colonization when the organism is attached to the epithelial surface. Chain-forming mutants and the parental strain grown under conditions to promote chain formation showed increased adherence to human epithelial cells (A549 cells) in vitro. Additionally, adherence to A549 cells selected for longer chains within the wild-type strain. In vivo in a murine model of colonization, chain-forming mutants outcompeted the parental strain. Together, our results demonstrate that morphological heterogeneity in the pneumococcus may promote colonization of the upper respiratory tract by enhancing the ability of the organism to bind to the epithelial surface. PMID:22825449
-
Sidaway, Ben; Euloth, Tracey; Caron, Heather; Piskura, Matthew; Clancy, Jessica; Aide, Alyson
2012-07-01
The purpose of this study was to compare the reliability of three previously used techniques for the measurement of ankle dorsiflexion ROM, open-chained goniometry, closed-chained goniometry, and inclinometry, to a novel trigonometric technique. Twenty-one physiotherapy students used four techniques (open-chained goniometry, closed-chained goniometry, inclinometry, and trigonometry) to assess dorsiflexion range of motion in 24 healthy volunteers. All student raters underwent training to establish competence in the four techniques. Raters then measured dorsiflexion with a randomly assigned measuring technique four times over two sessions, one week apart. Data were analyzed using a technique by session analysis of variance, technique measurement variability being the primary index of reliability. Comparisons were also made between the measurements derived from the four techniques and those obtained from a computerized video analysis system. Analysis of the rater measurement variability around the technique means revealed significant differences between techniques with the least variation being found in the trigonometric technique. Significant differences were also found between the technique means but no differences between sessions were evident. The trigonometric technique produced mean ROMs closest in value to those derived from computer analysis. Application of the trigonometric technique resulted in the least variability in measurement across raters and consequently should be considered for use when changes in dorsiflexion ROM need to be reliably assessed. Copyright © 2012 Elsevier B.V. All rights reserved.
-
Li, Y; Spellerberg, M B; Stevenson, F K; Capra, J D; Potter, K N
1996-03-01
Essentially all cold agglutinins (CA) with red blood cell I/i specificity isolated from patients with CA disease stemming from lymphoproliferative disorders utilize the VH 4-34 (VH 4-21) gene segment. This near universality of the restricted use of a single gene segment is substantially greater than that demonstrated for other autoantibodies. The monoclonal antibody 9G4 exclusively binds VH 4-34 encoded antibodies and serves as a marker for the VH 4-34 gene segment. Previous studies form our laboratory localized the 9G4 reactive area to framework region 1 (FR1). In the present study, the relative roles of VH FR1, heavy (H) chain complementarity determining region 3 (CDRH 3) and the light (L) chain in I antigen binding were investigated. Mutants containing FR1 sequences from the other VH families, CDRH 3 exchanges, and combinatorial antibodies involving L chain interchanges were produced in the baculovirus system and tested in an I binding assay. The data indicate that FR1 of the VH 4-34 gene segment and the CDRH 3 are essential for the interaction between CA and the I antigen, with the CDRH 3 being fundamental in determining the fine specificity of antigen binding (I versus i). Mutants with substantially altered CDRH 1 and CDRH 2 regions bind I as long as the FR1 is VH 4-34 encoded and the CDRH 3 has a permissive sequence. Light chain swaps indicate that even though antigen binding is predominantly mediated by the H chain, the association with antigen can be abrogated by an incompatible L chain. The necessity for VH 4-34 FR1 explains the almost exclusive use of the VH 4-34 gene segment in cold agglutinins. We hypothesize that, as a general phenomenon, the H chain FR1 of many antibodies may be important in providing the contact required for the close association of antibody with antigen, while the CDRH 3 dictates the fine specificity and strenght of binding.
-
Multidimensional competences of supply chain managers: an empirical study
NASA Astrophysics Data System (ADS)
Shou, Yongyi; Wang, Weijiao
2017-01-01
Supply chain manager competences have attracted increasing attention from both practitioners and scholars in recent years. This paper conducted an explorative study to understand the dimensionality of supply chain manager competences. Online job advertisements for supply chain managers were collected as secondary data, since these advertisements reflect employers' real job requirements. We adopted the multidimensional scaling (MDS) technique to process and analyse the data. Five dimensions of supply chain manager competences are identified: generic skills, functional skills, supply chain management (SCM) qualifications and leadership, SCM expertise, and industry-specific and senior management skills. Statistic tests indicate that supply chain manager competence saliences vary in different industries and regions.
-
Kinks, loops, and protein folding, with protein A as an example
Krokhotin, Andrey; Liwo, Adam; Maisuradze, Gia G.; Niemi, Antti J.; Scheraga, Harold A.
2014-01-01
The dynamics and energetics of formation of loops in the 46-residue N-terminal fragment of the B-domain of staphylococcal protein A has been studied. Numerical simulations have been performed using coarse-grained molecular dynamics with the united-residue (UNRES) force field. The results have been analyzed in terms of a kink (heteroclinic standing wave solution) of a generalized discrete nonlinear Schrödinger (DNLS) equation. In the case of proteins, the DNLS equation arises from a Cα-trace-based energy function. Three individual kink profiles were identified in the experimental three-α-helix structure of protein A, in the range of the Glu16-Asn29, Leu20-Asn29, and Gln33-Asn44 residues, respectively; these correspond to two loops in the native structure. UNRES simulations were started from the full right-handed α-helix to obtain a clear picture of kink formation, which would otherwise be blurred by helix formation. All three kinks emerged during coarse-grained simulations. It was found that the formation of each is accompanied by a local free energy increase; this is expressed as the change of UNRES energy which has the physical sense of the potential of mean force of a polypeptide chain. The increase is about 7 kcal/mol. This value can thus be considered as the free energy barrier to kink formation in full α-helical segments of polypeptide chains. During the simulations, the kinks emerge, disappear, propagate, and annihilate each other many times. It was found that the formation of a kink is initiated by an abrupt change in the orientation of a pair of consecutive side chains in the loop region. This resembles the formation of a Bloch wall along a spin chain, where the Cα backbone corresponds to the chain, and the amino acid side chains are interpreted as the spin variables. This observation suggests that nearest-neighbor side chain–side chain interactions are responsible for initiation of loop formation. It was also found that the individual kinks are reflected as clear peaks in the principal modes of the analyzed trajectory of protein A, the shapes of which resemble the directional derivatives of the kinks along the chain. These observations suggest that the kinks of the DNLS equation determine the functionally important motions of proteins. PMID:24437917
-
NASA Astrophysics Data System (ADS)
Malekzadeh Moghani, Mahdy; Khomami, Bamin
2016-01-01
Macromolecules with ionizable groups are ubiquitous in biological and synthetic systems. Due to the complex interaction between chain and electrostatic decorrelation lengths, both equilibrium properties and micro-mechanical response of dilute solutions of polyelectrolytes (PEs) are more complex than their neutral counterparts. In this work, the bead-rod micromechanical description of a chain is used to perform hi-fidelity Brownian dynamics simulation of dilute PE solutions to ascertain the self-similar equilibrium behavior of PE chains with various linear charge densities, scaling of the Kuhn step length (lE) with salt concentration cs and the force-extension behavior of the PE chain. In accord with earlier theoretical predictions, our results indicate that for a chain with n Kuhn segments, lE ˜ cs-0.5 as linear charge density approaches 1/n. Moreover, the constant force ensemble simulation results accurately predict the initial non-linear force-extension region of PE chain recently measured via single chain experiments. Finally, inspired by Cohen's extraction of Warner's force law from the inverse Langevin force law, a novel numerical scheme is developed to extract a new elastic force law for real chains from our discrete set of force-extension data similar to Padè expansion, which accurately depicts the initial non-linear region where the total Kuhn length is less than the thermal screening length.
-
Malekzadeh Moghani, Mahdy; Khomami, Bamin
2016-01-14
Macromolecules with ionizable groups are ubiquitous in biological and synthetic systems. Due to the complex interaction between chain and electrostatic decorrelation lengths, both equilibrium properties and micro-mechanical response of dilute solutions of polyelectrolytes (PEs) are more complex than their neutral counterparts. In this work, the bead-rod micromechanical description of a chain is used to perform hi-fidelity Brownian dynamics simulation of dilute PE solutions to ascertain the self-similar equilibrium behavior of PE chains with various linear charge densities, scaling of the Kuhn step length (lE) with salt concentration cs and the force-extension behavior of the PE chain. In accord with earlier theoretical predictions, our results indicate that for a chain with n Kuhn segments, lE ∼ cs (-0.5) as linear charge density approaches 1/n. Moreover, the constant force ensemble simulation results accurately predict the initial non-linear force-extension region of PE chain recently measured via single chain experiments. Finally, inspired by Cohen's extraction of Warner's force law from the inverse Langevin force law, a novel numerical scheme is developed to extract a new elastic force law for real chains from our discrete set of force-extension data similar to Padè expansion, which accurately depicts the initial non-linear region where the total Kuhn length is less than the thermal screening length.
-
Li, Longhai; Feng, Cindy X; Qiu, Shi
2017-06-30
An important statistical task in disease mapping problems is to identify divergent regions with unusually high or low risk of disease. Leave-one-out cross-validatory (LOOCV) model assessment is the gold standard for estimating predictive p-values that can flag such divergent regions. However, actual LOOCV is time-consuming because one needs to rerun a Markov chain Monte Carlo analysis for each posterior distribution in which an observation is held out as a test case. This paper introduces a new method, called integrated importance sampling (iIS), for estimating LOOCV predictive p-values with only Markov chain samples drawn from the posterior based on a full data set. The key step in iIS is that we integrate away the latent variables associated the test observation with respect to their conditional distribution without reference to the actual observation. By following the general theory for importance sampling, the formula used by iIS can be proved to be equivalent to the LOOCV predictive p-value. We compare iIS and other three existing methods in the literature with two disease mapping datasets. Our empirical results show that the predictive p-values estimated with iIS are almost identical to the predictive p-values estimated with actual LOOCV and outperform those given by the existing three methods, namely, the posterior predictive checking, the ordinary importance sampling, and the ghosting method by Marshall and Spiegelhalter (2003). Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.
-
Pinto, Rute D; Randelli, Elisa; Buonocore, Francesco; Pereira, Pedro J B; dos Santos, Nuno M S
2013-03-01
In this work, the gene and cDNA of sea bass (Dicentrarchus labrax) β2-microglobulin (Dila-β2m) and several cDNAs of MHC class I heavy chain (Dila-UA) were characterized. While Dila-β2m is single-copy, numerous Dila-UA transcripts were identified per individual with variability at the peptide-binding domain (PBD), but also with unexpected diversity from the connective peptide (CP) through the 3' untranslated region (UTR). Phylogenetic analysis segregates Dila-β2m and Dila-UA into each subfamily cluster, placing them in the fish class and branching Dila-MHC-I with lineage U. The α1 domains resemble those of the recently proposed L1 trans-species lineage. Although no Dila-specific α1, α2 or α3 sub-lineages could be observed, two highly distinct sub-lineages were identified at the CP/TM/CYT regions. The three-dimensional homology model of sea bass MHC-I complex is consistent with other characterized vertebrate structures. Furthermore, basal tissue-specific expression profiles were determined for both molecules, and expression of β2m was evaluated after poly I:C stimulus. Results suggest these molecules are orthologues of other β2m and teleost classical MHC-I and their basic structure is evolutionarily conserved, providing relevant information for further studies on antigen presentation in this fish species. Copyright © 2012 Elsevier Ltd. All rights reserved.
-
Garcia-Rodriguez, Consuelo; Razai, Ali; Geren, Isin N; Lou, Jianlong; Conrad, Fraser; Wen, Wei-Hua; Farr-Jones, Shauna; Smith, Theresa J; Brown, Jennifer L; Skerry, Janet C; Smith, Leonard A; Marks, James D
2018-03-01
Human botulism is most commonly caused by botulinum neurotoxin (BoNT) serotypes A, B, and E. For this work, we sought to develop a human monoclonal antibody (mAb)-based antitoxin capable of binding and neutralizing multiple subtypes of BoNT/E. Libraries of yeast-displayed single chain Fv (scFv) antibodies were created from the heavy and light chain variable region genes of humans immunized with pentavalent-toxoid- and BoNT/E-binding scFv isolated by Fluorescence-Activated Cell Sorting (FACS). A total of 10 scFv were isolated that bound one or more BoNT/E subtypes with nanomolar-level equilibrium dissociation constants (K D ). By diversifying the V-regions of the lead mAbs and selecting for cross-reactivity, we generated three scFv that bound all four BoNT/E subtypes tested at three non-overlapping epitopes. The scFvs were converted to IgG that had K D values for the different BoNT/E subtypes ranging from 9.7 nM to 2.28 pM. An equimolar combination of the three mAbs was able to potently neutralize BoNT/E1, BoNT/E3, and BoNT/E4 in a mouse neutralization assay. The mAbs have potential utility as therapeutics and as diagnostics capable of recognizing multiple BoNT/E subtypes. A derivative of the three-antibody combination (NTM-1633) is in pre-clinical development with an investigational new drug (IND) application filing expected in 2018.
-
Savidor, Alon; Barzilay, Rotem; Elinger, Dalia; Yarden, Yosef; Lindzen, Moshit; Gabashvili, Alexandra; Adiv Tal, Ophir; Levin, Yishai
2017-06-01
Traditional "bottom-up" proteomic approaches use proteolytic digestion, LC-MS/MS, and database searching to elucidate peptide identities and their parent proteins. Protein sequences absent from the database cannot be identified, and even if present in the database, complete sequence coverage is rarely achieved even for the most abundant proteins in the sample. Thus, sequencing of unknown proteins such as antibodies or constituents of metaproteomes remains a challenging problem. To date, there is no available method for full-length protein sequencing, independent of a reference database, in high throughput. Here, we present Database-independent Protein Sequencing, a method for unambiguous, rapid, database-independent, full-length protein sequencing. The method is a novel combination of non-enzymatic, semi-random cleavage of the protein, LC-MS/MS analysis, peptide de novo sequencing, extraction of peptide tags, and their assembly into a consensus sequence using an algorithm named "Peptide Tag Assembler." As proof-of-concept, the method was applied to samples of three known proteins representing three size classes and to a previously un-sequenced, clinically relevant monoclonal antibody. Excluding leucine/isoleucine and glutamic acid/deamidated glutamine ambiguities, end-to-end full-length de novo sequencing was achieved with 99-100% accuracy for all benchmarking proteins and the antibody light chain. Accuracy of the sequenced antibody heavy chain, including the entire variable region, was also 100%, but there was a 23-residue gap in the constant region sequence. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.
-
Lee, Jiyeon; Kim, Hye-Jin; Roh, Jooho; Seo, Youngsil; Kim, Minjae; Jun, Hye-Ryeong; Pham, Chuong D.; Kwon, Myung-Hee
2013-01-01
Many murine monoclonal anti-DNA antibodies (Abs) derived from mice models for systemic lupus erythematosus have additional cell-penetration and/or nucleic acid-hydrolysis properties. Here, we examined the influence of deactivating each complementarity-determining region (CDR) within a multifunctional anti-nucleic acid antibody (Ab) that possesses these activities, the catalytic 3D8 single chain variable fragment (scFv). CDR-deactivated 3D8 scFv variants were generated by replacing all of the amino acids within each CDR with Gly/Ser residues. The structure of 3D8 scFv accommodated single complete CDR deactivations. Different functional activities of 3D8 scFv were affected differently depending on which CDR was deactivated. The only exception was CDR1, located within the light chain (LCDR1); deactivation of LCDR1 abolished all of the functional activities of 3D8 scFv. A hybrid Ab, HW6/3D8L1, in which the LCDR1 from an unrelated Ab (HW6) was replaced with the LCDR1 from 3D8, acquired all activities associated with the 3D8 scFv. These results suggest that the activity of a multifunctional 3D8 scFv Ab can be modulated by single complete CDR deactivation and that the LCDR1 plays a crucial role in maintaining Ab properties. This study presents a new approach for determining the role of individual CDRs in multifunctional Abs with important implications for the future of Ab engineering. PMID:24155236
-
Barnidge, David R; Lundström, Susanna L; Zhang, Bo; Dasari, Surendra; Murray, David L; Zubarev, Roman A
2015-12-04
In our previous work, we showed that electrospray ionization of intact polyclonal kappa and lambda light chains isolated from normal serum generates two distinct, Gaussian-shaped, molecular mass distributions representing the light-chain repertoire. During the analysis of a large (>100) patient sample set, we noticed a low-intensity molecular mass distribution with a mean of approximately 24 250 Da, roughly 800 Da higher than the mean of the typical kappa molecular-mass distribution mean of 23 450 Da. We also observed distinct clones in this region that did not appear to contain any typical post-translational modifications that would account for such a large mass shift. To determine the origin of the high molecular mass clones, we performed de novo bottom-up mass spectrometry on a purified IgM monoclonal light chain that had a calculated molecular mass of 24 275.03 Da. The entire sequence of the monoclonal light chain was determined using multienzyme digestion and de novo sequence-alignment software and was found to belong to the germline allele IGKV2-30. The alignment of kappa germline sequences revealed ten IGKV2 and one IGKV4 sequences that contained additional amino acids in their CDR1 region, creating the high-molecular-mass phenotype. We also performed an alignment of lambda germline sequences, which showed additional amino acids in the CDR2 region, and the FR3 region of functional germline sequences that result in a high-molecular-mass phenotype. The work presented here illustrates the ability of mass spectrometry to provide information on the diversity of light-chain molecular mass phenotypes in circulation, which reflects the germline sequences selected by the immunoglobulin-secreting B-cell population.
-
Rahbarizadeh, Fatemeh; Rasaee, Mohammad J; Forouzandeh, Mehdi; Allameh, Abdolamir; Sarrami, Ramin; Nasiry, Habib; Sadeghizadeh, Majid
2005-01-01
Camelidae are known to produce immunoglobulins (Igs) devoid of light chains and constant heavy-chain domains (CH1). Antigen-specific fragments of these heavy-chain IgGs (VHH) are of great interest in biotechnology applications. This paper describes the first example of successfully raised heavy-chain antibodies in Camelus dromedarius (single-humped camel) and Camelus bactrianus (two-humped camel) against a MUC1 related peptide that is found to be an important epitope expressed in cancerous tissue. Camels were immunized against a synthetic peptide corresponding to the tandem repeat region of MUC1 mucin and cancerous tissue preparation obtained from patients suffering from breast carcinoma. Three IgG subclasses with different binding properties to protein A and G were purified by affinity chromatography. Both conventional and heavy-chain IgG antibodies were produced in response to MUC1-related peptide. The elicited antibodies could react specifically with the tandem repeat region of MUC1 mucin in an enzyme linked immunosorbant assay (ELISA). Anti-peptide antibodies were purified after passing antiserum over two affinity chromatography columns. Using ELISA, immunocytochemistry and Western blotting, the interaction of purified antibodies with different antigens was evaluated. The antibodies were observed to be selectively bound to antigens namely: MUC1 peptide (tandem repeat region), human milk fat globule membrane (HMFG), deglycosylated human milk fat globule membrane (D-HMFG), homogenized cancerous breast tissue and a native MUC1 purified from ascitic fluid. Ka values of specific polyclonal antipeptide antibodies were estimated in C. dromedarius and C. bactrianus, as 7 x 10(10) M(-1) and 1.4 x 10(10) M(-1) respectively.
-
Satoh, Yuto; Yonemori, Saeka; Hirose, Mitsuhiro; Shogaki, Hiroko; Wakimoto, Hiroshi; Kitagawa, Yoshinori; Gotoh, Bin; Shirai, Tsuyoshi; Takahashi, Ken-Ichi; Itoh, Masae
2017-02-01
The fusion (F) protein of measles virus performs refolding from the thermodynamically metastable prefusion form to the highly stable postfusion form via an activated unstable intermediate stage, to induce membrane fusion. Some amino acids involved in the fusion regulation cluster in the heptad repeat B (HR-B) domain of the stalk region, among which substitution of residue 465 by various amino acids revealed that fusion activity correlates well with its side chain length from the Cα (P<0.01) and van der Waals volume (P<0.001), except for Phe, Tyr, Trp, Pro and His carrying ring structures. Directed towards the head region, longer side chains of the non-ring-type 465 residues penetrate more deeply into the head region and may disturb the hydrophobic interaction between the stalk and head regions and cause destabilization of the molecule by lowering the energy barrier for refolding, which conferred the F protein enhanced fusion activity. Contrarily, the side chain of ring-type 465 residues turned away from the head region, resulting in not only no contact with the head region but also extensive coverage of the HR-B surface, which may prevent the dissociation of the HR-B bundle for initiation of membrane fusion and suppress fusion activity. Located in the HR-B domain just at the junction between the head and stalk regions, amino acid 465 is endowed with a possible ability to either destabilize or stabilize the F protein depending on its molecular volume and the direction of the side chain, regulating fusion activity of measles virus F protein.
-
Nonequilibrium thermodynamic potentials for continuous-time Markov chains.
Verley, Gatien
2016-01-01
We connect the rare fluctuations of an equilibrium (EQ) process and the typical fluctuations of a nonequilibrium (NE) stationary process. In the framework of large deviation theory, this observation allows us to introduce NE thermodynamic potentials. For continuous-time Markov chains, we identify the relevant pairs of conjugated variables and propose two NE ensembles: one with fixed dynamics and fluctuating time-averaged variables, and another with fixed time-averaged variables, but a fluctuating dynamics. Accordingly, we show that NE processes are equivalent to conditioned EQ processes ensuring that NE potentials are Legendre dual. We find a variational principle satisfied by the NE potentials that reach their maximum in the NE stationary state and whose first derivatives produce the NE equations of state and second derivatives produce the NE Maxwell relations generalizing the Onsager reciprocity relations.
-
Assessing drivers of export orientation in the subsea oil and gas industry.
Aarstad, Jarle; Pettersen, Inger Beate; Jakobsen, Stig-Erik
2015-01-01
The purpose of this short study was to identify the drivers of export orientation of firms in the subsea oil and gas industry in Western Norway. As the oil fields in the North Sea are approaching a stage of maturity, gaining knowledge of these drivers is crucial. An online survey was conducted of firms operating in the subsea oil and gas industry in the region. Consistent with previous research, the data reveal that product innovation and a majority share of international ownership increase firms' export rates. The use of instrumental variables indicates that both product innovation and international ownership are causes of subsea petroleum exports. The study moreover finds that subcontractors have a lower rate of direct exports than system providers, but international ownership in particular boosts subcontractors' export rates, probably by decreasing their market dependency on regional system providers. A clear recommendation for managers and stakeholders is that they should encourage foreign investments throughout the value chain. The results of such a strategy appear to be especially positive for subcontractors.
-
Rickettsia felis in Ctenocephalides felis felis from Five Geographic Regions of Brazil
Horta, Mauricio C.; Ogrzewalska, Maria; Azevedo, Milka C.; Costa, Francisco B.; Ferreira, Fernando; Labruna, Marcelo B.
2014-01-01
This study evaluated rickettsial infection in 701 Ctenocephalides felis felis fleas that were collected from dogs and cats in 31 municipalities, encompassing all regions and major biomes of Brazil. A total of 268 (38.2%) fleas from 30 municipalities were polymerase chain reaction (PCR) positive for the rickettsial gltA gene. The PCR products from 44 fleas, consisting of at least 1 PCR-positive flea from each of 30 municipalities, generated DNA sequences identical to Rickettsia felis. Rickettsial prevalence was highly variable among 30 municipalities, with values ranging from 2.9% to 100%. Significantly higher infection rates by R. felis were associated with the Pampa biome (southern Brazil), and the temperate climate that prevails in southern Brazil. In contrast, lowest R. felis-infection rates were significantly associated with the Caatinga biome, and its semiarid climate. Further studies should evaluate the effect of temperature and moisture on the R. felis infection in Ctenocephalides fleas world widely. PMID:24778194
-
Prevalence of Torque teno virus in healthy donors of Paraná State, southern Brazil
Mazzola, Jocimara Costa; Saito, Patrícia Keiko; Yamakawa, Roger Haruki; Watanabe, Maria Angélica Ehara; da Silva Junior, Waldir Veríssimo; Matta, Alessandra Cristina Gobbi; Borelli, Sueli Donizete
2015-01-01
Objective To determine the prevalence of the Torque teno virus in healthy donors in the northern and northwestern regions of the state of Paraná, southern Brazil. Methods The Torque teno virus was detected by a nested polymerase chain reaction using a set of oligoprimers for the N22 region. Results The prevalence of the virus was 69% in 551 healthy blood donors in southern Brazil. There was no statistically significant difference between the presence of the virus and the variables gender, ethnicity and marital status. There was significant difference in the prevalence of the virus regarding the age of the donors (p-value = 0.024) with a higher incidence (74.7%) in 18- to 24-year-old donors. Conclusion A high prevalence of Torque teno virus was observed in the population studied. Further studies are needed to elucidate the routes of contamination and the clinical implications of the virus in the healthy population. PMID:26408369
-
Distribution, morphology, and origins of Martian pit crater chains
NASA Astrophysics Data System (ADS)
Wyrick, Danielle; Ferrill, David A.; Morris, Alan P.; Colton, Shannon L.; Sims, Darrell W.
2004-06-01
Pit craters are circular to elliptical depressions found in alignments (chains), which in many cases coalesce into linear troughs. They are common on the surface of Mars and similar to features observed on Earth and other terrestrial bodies. Pit craters lack an elevated rim, ejecta deposits, or lava flows that are associated with impact craters or calderas. It is generally agreed that the pits are formed by collapse into a subsurface cavity or explosive eruption. Hypotheses regarding the formation of pit crater chains require development of a substantial subsurface void to accommodate collapse of the overlying material. Suggested mechanisms of formation include: collapsed lava tubes, dike swarms, collapsed magma chamber, substrate dissolution (analogous to terrestrial karst), fissuring beneath loose material, and dilational faulting. The research described here is intended to constrain current interpretations of pit crater chain formation by analyzing their distribution and morphology. The western hemisphere of Mars was systematically mapped using Mars Orbiter Camera (MOC) images to generate ArcView™ Geographic Information System (GIS) coverages. All visible pit crater chains were mapped, including their orientations and associations with other structures. We found that pit chains commonly occur in areas that show regional extension or local fissuring. There is a strong correlation between pit chains and fault-bounded grabens. Frequently, there are transitions along strike from (1) visible faulting to (2) faults and pits to (3) pits alone. We performed a detailed quantitative analysis of pit crater morphology using MOC narrow angle images, Thermal Emission Imaging System (THEMIS) visual images, and Mars Orbiter Laser Altimeter (MOLA) data. This allowed us to determine a pattern of pit chain evolution and calculate pit depth, slope, and volume. Volumes of approximately 150 pits from five areas were calculated to determine volume size distribution and regional trends. The information collected in the study was then compared with non-Martian examples of pit chains and physical analog models. We evaluated the various mechanisms for pit chain development based on the data collected and conclude that dilational normal faulting and sub-vertical fissuring provide the simplest and most comprehensive mechanisms to explain the regional associations, detailed geometry, and progression of pit chain development.
-
Choice between Single and Multiple Reinforcers in Concurrent-Chains Schedules
ERIC Educational Resources Information Center
Mazur, James E.
2006-01-01
Pigeons responded on concurrent-chains schedules with equal variable-interval schedules as initial links. One terminal link delivered a single reinforcer after a fixed delay, and the other terminal link delivered either three or five reinforcers, each preceded by a fixed delay. Some conditions included a postreinforcer delay after the single…
-
Temporal Context in Concurrent Chains: I. Terminal-Link Duration
ERIC Educational Resources Information Center
Grace, Randolph C.
2004-01-01
Two experiments are reported in which the ratio of the average times spent in the terminal and initial links ("Tt/Ti") in concurrent chains was varied. In Experiment 1, pigeons responded in a three-component procedure in which terminal-link variable-interval schedules were in constant ratio, but their average duration increased across components…
-
USDA-ARS?s Scientific Manuscript database
Channel catfish, Ictalurus punctatus, T cell receptors (TCR) gamma and delta were identified by mining of expressed sequence tag databases and full length sequences were obtained by 5'-RACE and RT-PCR protocols. cDNAs for each of these TCR chains encode typical variable (V), (diversity; D), joining ...
-
Mobilising the Commodity Chain Concept in the Politics of Food and Farming
ERIC Educational Resources Information Center
Jackson, Peter; Ward, Neil; Russell, Polly
2006-01-01
Focusing on the concept of "commodity chains" within the food industry, this paper analyses the term's widespread and variable usage in both academic and policy-orientated work. Despite recent criticisms, the concept has retained its popular appeal alongside competing metaphors such as networks, circuits and assemblages. Examining the…
-
Transport of spin qubits with donor chains under realistic experimental conditions
NASA Astrophysics Data System (ADS)
Mohiyaddin, Fahd A.; Kalra, Rachpon; Laucht, Arne; Rahman, Rajib; Klimeck, Gerhard; Morello, Andrea
2016-07-01
The ability to transport quantum information across some distance can facilitate the design and operation of a quantum processor. One-dimensional spin chains provide a compact platform to realize scalable spin transport for a solid-state quantum computer. Here, we model odd-sized donor chains in silicon under a range of experimental nonidealities, including variability of donor position within the chain. We show that the tolerance against donor placement inaccuracies is greatly improved by operating the spin chain in a mode where the electrons are confined at the Si-SiO2 interface. We then estimate the required time scales and exchange couplings, and the level of noise that can be tolerated to achieve high-fidelity transport. We also propose a protocol to calibrate and initialize the chain, thereby providing a complete guideline for realizing a functional donor chain and utilizing it for spin transport.
-
Transport of spin qubits with donor chains under realistic experimental conditions
Mohiyaddin, Fahd A.; Kalra, Rachpon; Laucht, Arne; ...
2016-07-25
The ability to transport quantum information across some distance can facilitate the design and operation of a quantum processor. One-dimensional spin chains provide a compact platform to realize scalable spin transport for a solid-state quantum computer. Here, we model odd-sized donor chains in silicon under a range of experimental nonidealities, including variability of donor position within the chain. We show that the tolerance against donor placement inaccuracies is greatly improved by operating the spin chain in a mode where the electrons are confined at the Si-SiO 2 interface. We then estimate the required time scales and exchange couplings, and themore » level of noise that can be tolerated to achieve high-fidelity transport. As a result, we also propose a protocol to calibrate and initialize the chain, thereby providing a complete guideline for realizing a functional donor chain and utilizing it for spin transport.« less
-
Farris, M Heath; Scott, Andrew R; Texter, Pamela A; Bartlett, Marta; Coleman, Patricia; Masters, David
2018-04-11
Single nucleotide polymorphisms (SNPs) located within the human genome have been shown to have utility as markers of identity in the differentiation of DNA from individual contributors. Massively parallel DNA sequencing (MPS) technologies and human genome SNP databases allow for the design of suites of identity-linked target regions, amenable to sequencing in a multiplexed and massively parallel manner. Therefore, tools are needed for leveraging the genotypic information found within SNP databases for the discovery of genomic targets that can be evaluated on MPS platforms. The SNP island target identification algorithm (TIA) was developed as a user-tunable system to leverage SNP information within databases. Using data within the 1000 Genomes Project SNP database, human genome regions were identified that contain globally ubiquitous identity-linked SNPs and that were responsive to targeted resequencing on MPS platforms. Algorithmic filters were used to exclude target regions that did not conform to user-tunable SNP island target characteristics. To validate the accuracy of TIA for discovering these identity-linked SNP islands within the human genome, SNP island target regions were amplified from 70 contributor genomic DNA samples using the polymerase chain reaction. Multiplexed amplicons were sequenced using the Illumina MiSeq platform, and the resulting sequences were analyzed for SNP variations. 166 putative identity-linked SNPs were targeted in the identified genomic regions. Of the 309 SNPs that provided discerning power across individual SNP profiles, 74 previously undefined SNPs were identified during evaluation of targets from individual genomes. Overall, DNA samples of 70 individuals were uniquely identified using a subset of the suite of identity-linked SNP islands. TIA offers a tunable genome search tool for the discovery of targeted genomic regions that are scalable in the population frequency and numbers of SNPs contained within the SNP island regions. It also allows the definition of sequence length and sequence variability of the target region as well as the less variable flanking regions for tailoring to MPS platforms. As shown in this study, TIA can be used to discover identity-linked SNP islands within the human genome, useful for differentiating individuals by targeted resequencing on MPS technologies.
-
Modeling the disturbance of vegetation by fire in the boreal forest
NASA Astrophysics Data System (ADS)
Crevoisier, C.; Shevliakova, E.; Gloor, M.; Wirth, C.
2006-12-01
Boreal regions are important for the global carbon cycle because it is the largest forested area on earth and there are large belowground carbon pools (~1000 PgC). It is also a region where largest warming trends on the globe over the last decades have been observed and changes of the land ecosystems have already started. A major factor that determines the structure and carbon dynamics of the boreal forest is fire. As fire frequency depends strongly on climate, increased fire occurrence and related losses to the atmosphere are likely, and have already been reported. In order to predict with more confidence the occurrence and effect of fire on forest ecosystems in the boreal region, we have developed a fire model that takes advantage of the large on-ground, remote sensing and climate data from Canada, Alaska and Siberia. This prognostic model estimates the monthly burned area in a grid cell of 2 by 2.5 degrees, from four climate (air temperature, air relative humidity, precipitation and soil water content) and one human-related (road density) variables. Parameters are estimated using a Markov Chain Monte Carlo method applied to a dataset of observed burned area for Canada. The model is able to reproduce the seasonality of fire, the interannual variability, as well as the location of fire events, not only for Canada (on which data the model is based), but also for Siberia and Alaska, for which the results compare well with remote sensing observation, and are in the range of various current estimations of burned area. The fire model is being implemented in LM3V, the new vegetation model of GFDL earth system model, in order to make prediction of future fire behavior in boreal regions, and the related disturbance of the vegetation and carbon emissions.
-
Cerruti Mainardi, Paola
2006-01-01
The Cri du Chat syndrome (CdCS) is a genetic disease resulting from a deletion of variable size occurring on the short arm of chromosome 5 (5p-). The incidence ranges from 1:15,000 to 1:50,000 live-born infants. The main clinical features are a high-pitched monochromatic cry, microcephaly, broad nasal bridge, epicanthal folds, micrognathia, abnormal dermatoglyphics, and severe psychomotor and mental retardation. Malformations, although not very frequent, may be present: cardiac, neurological and renal abnormalities, preauricular tags, syndactyly, hypospadias, and cryptorchidism. Molecular cytogenetic analysis has allowed a cytogenetic and phenotypic map of 5p to be defined, even if results from the studies reported up to now are not completely in agreement. Genotype-phenotype correlation studies showed a clinical and cytogenetic variability. The identification of phenotypic subsets associated with a specific size and type of deletion is of diagnostic and prognostic relevance. Specific growth and psychomotor development charts have been established. Two genes, Semaphorin F (SEMAF) and δ-catenin (CTNND2), which have been mapped to the "critical regions", are potentially involved in cerebral development and their deletion may be associated with mental retardation in CdCS patients. Deletion of the telomerase reverse transcriptase (hTERT) gene, localised to 5p15.33, could contribute to the phenotypic changes in CdCS. The critical regions were recently refined by using array comparative genomic hybridisation. The cat-like cry critical region was further narrowed using quantitative polymerase chain reaction (PCR) and three candidate genes were characterised in this region. The diagnosis is based on typical clinical manifestations. Karyotype analysis and, in doubtful cases, FISH analysis will confirm the diagnosis. There is no specific therapy for CdCS but early rehabilitative and educational interventions improve the prognosis and considerable progress has been made in the social adjustment of CdCS patients. PMID:16953888
-
NASA Astrophysics Data System (ADS)
Araki, Mitsunori; Takano, Shuro; Sakai, Nami; Yamamoto, Satoshi; Oyama, Takahiro; Kuze, Nobuhiko; Tsukiyama, Koichi
2017-06-01
Carbon chains in the warm carbon chain chemistry (WCCC) region has been searched in the 42-44 GHz region by using Green Bank 100 m telescope. Long carbon chains C_{7}H, C_{6}H, CH_{3}CCCCH, and linear-C_{6}H_{2} and cyclic species C_{3}H and C_{3}H_{2}O have been detected in the low-mass star forming region L1527, performing the WCCC. C_{7}H was detected for the first time in molecular clouds. The column density of C_{7}H is derived to be 6.2 × 10^{10} cm^{-2} by using the detected J = 24.5-23.5 and 25.5-24.5 rotational lines. The ^{2}Π_{1/2} electronic state of C_{6}H, locating 21.6 K above the ^{2}Π_{3/2} electronic ground state, and the K_a = 0 line of the para species of linear-C_{6}H_{2} were also detected firstly in molecular clouds. The column densities of the ^{2}Π_{1/2} and ^{2}Π_{3/2} states of C_{6}H in L1527 were derived to be 1.6 × 10^{11} and 1.1 × 10^{12} cm^{-2}, respectively. The total column density of linear-C_{6}H_{2} is obtained to be 1.86 × 10^{11} cm^{-2}. While the abundance ratios of carbon chains in between L1527 and the starless dark cloud Taurus Molecular Cloud-1 Cyanopolyyne Peak (TMC-1 CP) have a trend of decrease by extension of carbon-chain length, column densities of CH_{3}CCCCH and C_{6}H are on the trend. However, the column densities of linear-C_{6}H_{2}, and C_{7}H are as abundant as those of TMC-1 CP in spite of long carbon chain, i.e., they are not on the trend. The abundances of linear-C_{6}H_{2} and C_{7}H show that L1527 is rich for long carbon chains as well as TMC-1 CP.
-
Barros-Álvarez, Ximena; Kerchner, Keshia M; Koh, Cho Yeow; Turley, Stewart; Pardon, Els; Steyaert, Jan; Ranade, Ranae M; Gillespie, J Robert; Zhang, Zhongsheng; Verlinde, Christophe L M J; Fan, Erkang; Buckner, Frederick S; Hol, Wim G J
2017-07-01
The crystal structure of Leishmania donovani tyrosyl-tRNA synthetase (LdTyrRS) in complex with a nanobody and the tyrosyl adenylate analog TyrSA was determined at 2.75 Å resolution. Nanobodies are the variable domains of camelid heavy chain-only antibodies. The nanobody makes numerous crystal contacts and in addition reduces the flexibility of a loop of LdTyrRS. TyrSA is engaged in many interactions with active site residues occupying the tyrosine and adenine binding pockets. The LdTyrRS polypeptide chain consists of two pseudo-monomers, each consisting of two domains. Comparing the two independent chains in the asymmetric unit reveals that the two pseudo-monomers of LdTyrRS can bend with respect to each other essentially as rigid bodies. This flexibility might be useful in the positioning of tRNA for catalysis since both pseudo-monomers in the LdTyrRS chain are needed for charging tRNA Tyr . An "extra pocket" (EP) appears to be present near the adenine binding region of LdTyrRS. Since this pocket is absent in the two human homologous enzymes, the EP provides interesting opportunities for obtaining selective drugs for treating infections caused by L. donovani, a unicellular parasite causing visceral leishmaniasis, or kala azar, which claims 20,000 to 30,000 deaths per year. Sequence and structural comparisons indicate that the EP is a characteristic which also occurs in the active site of several other important pathogenic protozoa. Therefore, the structure of LdTyrRS could inspire the design of compounds useful for treating several different parasitic diseases. Copyright © 2017 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.
-
Zhu, Lijuan; Liao, Wenjun; Zhu, Huifen; Lei, Ping; Wang, Zhihua; Shao, Jingfang; Zhang, Yue; Shen, Guanxin
2006-01-01
The expression vector of SmIg scFv fragment was constructed in patient with B cell chronic lymphocyte leukemia (B-CLL) and expressed in E. coli to obtain scFv fragment, and the effect of the protein on the proliferation of stimulated peripheral blood mononuclear cells (PBMC) was investigated in vitro. Two pairs of primers were designed, and variable region genes of light chain and heavy chain were amplified by PCR respectively from the pGEM-T vectors previously constructed in our laboratory which containing light chain gene or Fd fragment of heavy chain gene. The PCR product was digested, purified and inserted into pHEN2 vector to construct the soluble expression vector pHEN2-scFv. After the induction by IPTG, the scFv protein was identified by SDS-PAGE electrophoresis and purified by Ni-NTA-Chromatography. MTT was used to determine the effect of purified protein on the proliferation of stimulated PBMC in vitro. Plasmid PCR and restriction enzyme digestion of pHEN2-scFv revealed the pHEN2-scFv vector was constructed successfully. Id-scFv protein was expressed in positive clone after induced by IPTG. SDS-PAGE analysis showed that the relative molecular weight of fusion protein was about 30 kD (1 kD= 0.9921 ku), which was consistent with the theoretically predicted value. Proliferation of PBMC could be induced by purified Id-scFv. It was suggested that the expression vector of SmIg scFv fragment was constructed successfully, and scFv protein was expressed and secreted from E. coli, which could induce proliferation of PBMC. This may lay an experimental foundation for further research of Id-HSP complex vaccine for B-CLL.
-
Predicting the Fluid-Phase Behavior of Aqueous Solutions of ELP (VPGVG) Sequences Using SAFT-VR.
Zhao, Binwu; Lindeboom, Tom; Benner, Steven; Jackson, George; Galindo, Amparo; Hall, Carol K
2017-10-24
The statistical associating fluid theory for potentials of variable range (SAFT-VR) is used to predict the fluid phase behavior of elastin-like polypeptide (ELP) sequences in aqueous solution with special focus on the loci of lower critical solution temperatures (LCSTs). A SAFT-VR model for these solutions is developed following a coarse-graining approach combining information from atomistic simulations and from previous SAFT models for previously reported relevant systems. Constant-pressure temperature-composition phase diagrams are determined for solutions of (VPGVG) n sequences + water with n = 1 to 300. The SAFT-VR equation of state lends itself to the straightforward calculation of phase boundaries so that complete fluid-phase equilibria can be calculated efficiently. A broad range of thermodynamic conditions of temperature and pressure are considered, and regions of vapor-liquid and liquid-liquid coexistence, including LCSTs, are found. The calculated phase boundaries at low concentrations match those measured experimentally. The temperature-composition phase diagrams of the aqueous ELP solutions at low pressure (0.1 MPa) are similar to those of types V and VI phase behavior in the classification of Scott and van Konynenburg. An analysis of the high-pressure phase behavior confirms, however, that a closed-loop liquid-liquid immiscibility region, separate from the gas-liquid envelope, is present for aqueous solutions of (VPGVG) 30 ; such a phase diagram is typical of type VI phase behavior. ELPs with shorter lengths exhibit both liquid-liquid and gas-liquid regions, both of which become less extensive as the chain length of the ELP is decreased. The strength of the hydrogen-bonding interaction is also found to affect the phase diagram of the (VPGVG) 30 system in that the liquid-liquid and gas-liquid regions expand as the hydrogen-bonding strength is decreased and shrink as it is increased. The LCSTs of the mixtures are seen to decrease as the ELP chain length is increased.
-
NASA Astrophysics Data System (ADS)
Lemos, A. T.; Ghisolfi, R. D. R.; Mazzini, P. L. F.
2018-06-01
The present study aimed to investigate the influence that four seamounts of the Vitória-Trindade Chain (VTC): the Vitória (VB), Jaseur (JB), Davis (DB) and Dogaressa (DoB) Banks, located on the western South Atlantic Ocean, potentially exert on the annual variability of the chlorophyll-a concentration [Chla] over their summits and surrounding regions. Nine years (January 2003 to December 2011) of monthly and weekly (8-days composite) satellite derived chlorophyll-a concentration, with 4 km spatial resolution were obtained for the study area using the Moderate Resolution Imaging Spectroradiometer (MODIS) aboard NASA's Aqua satellite. For comparison purposes, different time-series were analyzed for both the region over the seamounts and the surrounding waters. A Gaussian model was adjusted to each of the time series of monthly mean chlorophyll-a concentration, and the curve parameters were used in order to objectively characterize the blooms. The results showed that the entire study area (both above and beyond the seamounts) underwent seasonal blooms, with peak of chlorophyll-a occurring around the austral winter (June, July and August), when due to surface cooling the deepening of the surface mixed layer is observed, enriching the photic zone with nutrients. Nevertheless, the peak chlorophyll-a concentration over the shallow seamounts was twice higher than that over deep seamounts or in the adjacent deep ocean. Our results suggest that the presence of these seamounts and their morphological characteristics can significantly impact the primary productivity observed in this region. Thus, the VTC can be divided into areas of diffuse [Chla] (VB and JB), with lower zonal scattering and higher phytoplankton concentrations (DB), and areas distant from the continental shelf and the mesoscale processes that develop there, hence with lower [Chla] (DoB). The profound impact that these seamounts have on the oceanic ecosystem may turn them into becoming true oasis in the oligotrophic deep ocean, supporting higher trophic levels, as well as important fisheries in this region.
-
Capello, Daniela; Cerri, Michaela; Muti, Giuliana; Lucioni, Marco; Oreste, Pierluigi; Gloghini, Annunziata; Berra, Eva; Deambrogi, Clara; Franceschetti, Silvia; Rossi, Davide; Alabiso, Oscar; Morra, Enrica; Rambaldi, Alessandro; Carbone, Antonino; Paulli, Marco; Gaidano, Gianluca
2006-12-01
Post-transplant lymphoproliferative disorders (PTLD) derive from antigen-experienced B-cells and represent a major complication of solid organ transplantation. We characterized usage, mutation frequency and mutation pattern of immunoglobulin variable (IGV) gene rearrangements in 50 PTLD (polymorphic PTLD, n=10; diffuse large B-cell lymphoma, n=35; and Burkitt/Burkitt-like lymphoma, n=5). Among PTLD yielding clonal IGV amplimers, a functional IGV heavy chain (IGHV) rearrangement was found in 40/50 (80.0%) cases, whereas a potentially functional IGV light chain rearrangement was identified in 36/46 (78.3%) PTLD. By combining IGHV and IGV light chain rearrangements, 10/50 (20.0%) PTLD carried crippling mutations, precluding expression of a functional B-cell receptor (BCR). Immunohistochemistry showed detectable expression of IG light chains in only 18/43 (41.9%) PTLD. Failure to detect a functional IGV rearrangement associated with lack of IGV expression. Our data suggest that a large fraction of PTLD arise from germinal centre (GC)-experienced B-cells that display impaired BCR. Since a functional BCR is required for normal B-cell survival during GC transit, PTLD development may implicate rescue from apoptosis and expansion of B-cells that have failed the GC reaction. The high frequency of IGV loci inactivation appears to be a peculiar feature of PTLD among immunodeficiency-associated lymphoproliferations.
-
Size, shape, and diffusivity of a single Debye-Hückel polyelectrolyte chain in solution.
Soysa, W Chamath; Dünweg, B; Prakash, J Ravi
2015-08-14
Brownian dynamics simulations of a coarse-grained bead-spring chain model, with Debye-Hückel electrostatic interactions between the beads, are used to determine the root-mean-square end-to-end vector, the radius of gyration, and various shape functions (defined in terms of eigenvalues of the radius of gyration tensor) of a weakly charged polyelectrolyte chain in solution, in the limit of low polymer concentration. The long-time diffusivity is calculated from the mean square displacement of the centre of mass of the chain, with hydrodynamic interactions taken into account through the incorporation of the Rotne-Prager-Yamakawa tensor. Simulation results are interpreted in the light of the Odjik, Skolnick, Fixman, Khokhlov, and Khachaturian blob scaling theory (Everaers et al., Eur. Phys. J. E 8, 3 (2002)) which predicts that all solution properties are determined by just two scaling variables-the number of electrostatic blobs X and the reduced Debye screening length, Y. We identify three broad regimes, the ideal chain regime at small values of Y, the blob-pole regime at large values of Y, and the crossover regime at intermediate values of Y, within which the mean size, shape, and diffusivity exhibit characteristic behaviours. In particular, when simulation results are recast in terms of blob scaling variables, universal behaviour independent of the choice of bead-spring chain parameters, and the number of blobs X, is observed in the ideal chain regime and in much of the crossover regime, while the existence of logarithmic corrections to scaling in the blob-pole regime leads to non-universal behaviour.
-
Hora, Manuel; Carballo-Pacheco, Martin; Weber, Benedikt; Morris, Vanessa K.; Wittkopf, Antje; Buchner, Johannes; Strodel, Birgit; Reif, Bernd
2017-01-01
Antibody light chain amyloidosis is a rare disease caused by fibril formation of secreted immunoglobulin light chains (LCs). The huge variety of antibody sequences puts a serious challenge to drug discovery. The green tea polyphenol epigallocatechin-3-gallate (EGCG) is known to interfere with fibril formation in general. Here we present solution- and solid-state NMR studies as well as MD simulations to characterise the interaction of EGCG with LC variable domains. We identified two distinct EGCG binding sites, both of which include a proline as an important recognition element. The binding sites were confirmed by site-directed mutagenesis and solid-state NMR analysis. The EGCG-induced protein complexes are unstructured. We propose a general mechanistic model for EGCG binding to a conserved site in LCs. We find that EGCG reacts selectively with amyloidogenic mutants. This makes this compound a promising lead structure, that can handle the immense sequence variability of antibody LCs. PMID:28128355
-
TCR gamma chain diversity in the spleen of the duckbill platypus (Ornithorhynchus anatinus).
Parra, Zuly E; Arnold, Tamara; Nowak, Melissa A; Hellman, Lars; Miller, Robert D
2006-01-01
TCR gamma (TRG) chain diversity in splenic gammadelta T cells was determined for an egg-laying mammal (or monotreme), the duckbill platypus. Three distinct V subgroups were found in the expressed TRG chains and these three subgroups are members of a clade not found so far in eutherian mammals or birds. Each subgroup contains approximately five V gene segments, and their overall divergence is much less than is found in eutherians and birds, consistent with their recent evolution from an ancestral V gene segment. The platypus TRG locus also contains three C region genes and many of the residues involved in TCR function, such as interactions with CD3, were conserved in the monotreme C regions. All non-eutherian mammals (monotremes and marsupials) lacked the second cysteine residue necessary to form the intradomain disulfide bond in the C region, a loss apparently due to independent mutations in marsupials and monotremes. Monotreme TRGC regions also had among the most variation in the length of the connecting peptide region described for any species due to repeated motifs.
-
Off-Axis Seamount Lavas at 8°20' N Span the Entire Range of East Pacific Rise MORB Compositions
NASA Astrophysics Data System (ADS)
Anderson, M.; Wanless, V. D.; Perfit, M. R.; Gregg, P. M.; Fornari, D. J.; McCully, E.; Ridley, W. I.
2017-12-01
Lavas erupted at off-axis seamounts can provide a window into mantle heterogeneity and melting systematics that are not easily observed on-axis at fast-spreading mid-ocean ridges (MORs), where melts are efficiently mixed and homogenized within shallow axial magma chambers. To investigate off-axis magmatism, we systematically mapped the 8°20' N seamount chain in November of 2016 on R/V Atlantis using shipboard EM122 multibeam system and AUV Sentry. This 160-km long chain of off-axis seamounts and ridges is located perpendicular to the ridge axis, west of the East Pacific Rise (EPR) and north of the Siqueiros Fracture Zone. The high-resolution surface and AUV-based multibeam and AUV sidescan maps are combined with geochemical analyses of 300 basalt samples, collected using HOV Alvin and dredging, to evaluate magmatic plumbing and sources off-axis. Preliminary major and trace element concentrations reveal remarkable geochemical heterogeneity (including both normal and enriched basalt compositions) across the entire seamount chain and within individual seamounts. For example, (La/Sm)N contents span the entire range of known values for basalts from northern Pacific MORs and seamounts (0.45—2.76). MgO contents vary from 10.25 to 4.56 wt. % across the seamount chain and by as much as 3.61 wt. % from volcanic features sampled at an individual seamount (Beryl). Additionally, K2O/TiO2 ratios range from 4.9 to 61.3 across the seamount chain, and by as much as 54.4 at a single seamount (Beryl), indicating heterogeneous mantle sources or variable extents of melting occur at both regional and local scales. We combine the geochemical results and bathymetric maps with petrologic models to evaluate extents and depths of fractional crystallization and mantle melting in the off-axis environment.
-
Verification of Concurrent Programs. Part II. Temporal Proof Principles.
1981-09-01
not modify any of the shared program variables. In order to ensure the correct synchronization between the processes we use three semaphore variables...direct, simple, and intuitive rides for the establishment of these properties. rhey usually replace long but repetitively similar chains of primitive ...modify the variables on which Q actually depends. A typical case is that of semaphores . We have the following property: The Semaphore Variable Rule
-
Residue-Specific Side-Chain Polymorphisms via Particle Belief Propagation.
Ghoraie, Laleh Soltan; Burkowski, Forbes; Li, Shuai Cheng; Zhu, Mu
2014-01-01
Protein side chains populate diverse conformational ensembles in crystals. Despite much evidence that there is widespread conformational polymorphism in protein side chains, most of the X-ray crystallography data are modeled by single conformations in the Protein Data Bank. The ability to extract or to predict these conformational polymorphisms is of crucial importance, as it facilitates deeper understanding of protein dynamics and functionality. In this paper, we describe a computational strategy capable of predicting side-chain polymorphisms. Our approach extends a particular class of algorithms for side-chain prediction by modeling the side-chain dihedral angles more appropriately as continuous rather than discrete variables. Employing a new inferential technique known as particle belief propagation, we predict residue-specific distributions that encode information about side-chain polymorphisms. Our predicted polymorphisms are in relatively close agreement with results from a state-of-the-art approach based on X-ray crystallography data, which characterizes the conformational polymorphisms of side chains using electron density information, and has successfully discovered previously unmodeled conformations.
-
Tubiana, Luca; Polles, Guido; Orlandini, Enzo; Micheletti, Cristian
2018-06-07
The KymoKnot software package and web server identifies and locates physical knots or proper knots in a series of polymer conformations. It is mainly intended as an analysis tool for trajectories of linear or circular polymers, but it can be used on single instances too, e.g. protein structures in PDB format. A key element of the software package is the so-called minimally interfering chain closure algorithm that is used to detect physical knots in open chains and to locate the knotted region in both open and closed chains. The web server offers a user-friendly graphical interface that identifies the knot type and highlights the knotted region on each frame of the trajectory, which the user can visualize interactively from various viewpoints. The dynamical evolution of the knotted region along the chain contour is presented as a kymograph. All data can be downloaded in text format. The KymoKnot package is licensed under the BSD 3-Clause licence. The server is publicly available at http://kymoknot.sissa.it/kymoknot/interactive.php .
-
A cDNA Immunization Strategy to Generate Nanobodies against Membrane Proteins in Native Conformation
Eden, Thomas; Menzel, Stephan; Wesolowski, Janusz; Bergmann, Philine; Nissen, Marion; Dubberke, Gudrun; Seyfried, Fabienne; Albrecht, Birte; Haag, Friedrich; Koch-Nolte, Friedrich
2018-01-01
Nanobodies (Nbs) are soluble, versatile, single-domain binding modules derived from the VHH variable domain of heavy-chain antibodies naturally occurring in camelids. Nbs hold huge promise as novel therapeutic biologics. Membrane proteins are among the most interesting targets for therapeutic Nbs because they are accessible to systemically injected biologics. In order to be effective, therapeutic Nbs must recognize their target membrane protein in native conformation. However, raising Nbs against membrane proteins in native conformation can pose a formidable challenge since membrane proteins typically contain one or more hydrophobic transmembrane regions and, therefore, are difficult to purify in native conformation. Here, we describe a highly efficient genetic immunization strategy that circumvents these difficulties by driving expression of the target membrane protein in native conformation by cells of the immunized camelid. The strategy encompasses ballistic transfection of skin cells with cDNA expression plasmids encoding one or more orthologs of the membrane protein of interest and, optionally, other costimulatory proteins. The plasmid is coated onto 1 µm gold particles that are then injected into the shaved and depilated skin of the camelid. A gene gun delivers a helium pulse that accelerates the DNA-coated particles to a velocity sufficient to penetrate through multiple layers of cells in the skin. This results in the exposure of the extracellular domains of the membrane protein on the cell surface of transfected cells. Repeated immunization drives somatic hypermutation and affinity maturation of target-specific heavy-chain antibodies. The VHH/Nb coding region is PCR-amplified from B cells obtained from peripheral blood or a lymph node biopsy. Specific Nbs are selected by phage display or by screening of Nb-based heavy-chain antibodies expressed as secretory proteins in transfected HEK cells. Using this strategy, we have successfully generated agonistic and antagonistic Nbs against several cell surface ecto-enzymes and ligand-gated ion channels. PMID:29410663
-
Discharge data assimilation in a distributed hydrologic model for flood forecasting purposes
NASA Astrophysics Data System (ADS)
Ercolani, G.; Castelli, F.
2017-12-01
Flood early warning systems benefit from accurate river flow forecasts, and data assimilation may improve their reliability. However, the actual enhancement that can be obtained in the operational practice should be investigated in detail and quantified. In this work we assess the benefits that the simultaneous assimilation of discharge observations at multiple locations can bring to flow forecasting through a distributed hydrologic model. The distributed model, MOBIDIC, is part of the operational flood forecasting chain of Tuscany Region in Central Italy. The assimilation system adopts a mixed variational-Monte Carlo approach to update efficiently initial river flow, soil moisture, and a parameter related to runoff production. The evaluation of the system is based on numerous hindcast experiments of real events. The events are characterized by significant rainfall that resulted in both high and relatively low flow in the river network. The area of study is the main basin of Tuscany Region, i.e. Arno river basin, which extends over about 8300 km2 and whose mean annual precipitation is around 800 mm. Arno's mainstream, with its nearly 240 km length, passes through major Tuscan cities, as Florence and Pisa, that are vulnerable to floods (e.g. flood of November 1966). The assimilation tests follow the usage of the model in the forecasting chain, employing the operational resolution in both space and time (500 m and 15 minutes respectively) and releasing new flow forecasts every 6 hours. The assimilation strategy is evaluated in respect to open loop simulations, i.e. runs that do not exploit discharge observations through data assimilation. We compare hydrographs in their entirety, as well as classical performance indexes, as error on peak flow and Nash-Sutcliffe efficiency. The dependence of performances on lead time and location is assessed. Results indicate that the operational forecasting chain can benefit from the developed assimilation system, although with a significant variability due to the specific characteristics of any single event, and with downstream locations more sensitive to observations than upstream sites.
-
Characterizing Oregon's supply chains.
DOT National Transportation Integrated Search
2013-03-01
In many regions throughout the world, freight models are used to aid infrastructure investment and : policy decisions. Since freight is such an integral part of efficient supply chains, more realistic : transportation models can be of greater assista...
-
Surface vibrational structure at alkane liquid/vapor interfaces
NASA Astrophysics Data System (ADS)
Esenturk, Okan; Walker, Robert A.
2006-11-01
Broadband vibrational sum frequency spectroscopy (VSFS) has been used to examine the surface structure of alkane liquid/vapor interfaces. The alkanes range in length from n-nonane (C9H20) to n-heptadecane (C17H36), and all liquids except heptadecane are studied at temperatures well above their bulk (and surface) freezing temperatures. Intensities of vibrational bands in the CH stretching region acquired under different polarization conditions show systematic, chain length dependent changes. Data provide clear evidence of methyl group segregation at the liquid/vapor interface, but two different models of alkane chain structure can predict chain length dependent changes in band intensities. Each model leads to a different interpretation of the extent to which different chain segments contribute to the anisotropic interfacial region. One model postulates that changes in vibrational band intensities arise solely from a reduced surface coverage of methyl groups as alkane chain length increases. The additional methylene groups at the surface must be randomly distributed and make no net contribution to the observed VSF spectra. The second model considers a simple statistical distribution of methyl and methylene groups populating a three dimensional, interfacial lattice. This statistical picture implies that the VSF signal arises from a region extending several functional groups into the bulk liquid, and that the growing fraction of methylene groups in longer chain alkanes bears responsibility for the observed spectral changes. The data and resulting interpretations provide clear benchmarks for emerging theories of molecular structure and organization at liquid surfaces, especially for liquids lacking strong polar ordering.
-
Markov Chain Models for Stochastic Behavior in Resonance Overlap Regions
NASA Astrophysics Data System (ADS)
McCarthy, Morgan; Quillen, Alice
2018-01-01
We aim to predict lifetimes of particles in chaotic zoneswhere resonances overlap. A continuous-time Markov chain model isconstructed using mean motion resonance libration timescales toestimate transition times between resonances. The model is applied todiffusion in the co-rotation region of a planet. For particles begunat low eccentricity, the model is effective for early diffusion, butnot at later time when particles experience close encounters to the planet.
-
Iris B. Montague
2011-01-01
Many obstacles may deter hardwood manufacturers from obtaining chain-of-custody certification. Because the hardwood and softwood forest products industries have many differences between them, current certification systems may not fit the unique demographics of the hardwood industry. For this reason, it is important to understand chain-of-custody certification as it...
-
Zhang, Xiao; Xu, Chongxin; Zhang, Cunzheng; Liu, Yuan; Xie, Yajing; Liu, Xianjin
2014-04-01
ScFvs are composed of the variable regions of the heavy and light chains via a short linker that maintain the specific antigen binding abilities of antibodies. In this study, we constructed a naïve mouse phage displayed library to generate scFvs against Cry1Ab toxin. After affinity panning, positive phage-scFvs were isolated, sequenced and characterized by ELISA. The best binding ability scFv-G9 was expressed and purified. SDS-PAGE indicated that the relative molecular mass of scFv was estimated at 28 kDa. The purified scFv-G9 was used to develop a new DAS-ELISA for detecting Cry1Ab toxin, within minimum detection limit of 0.008 μg mL(-1), a working range 0.018-6.23 μg mL(-1), and the linear curve displayed an acceptable correlation coefficient of 0.98. The cross-reactivity showed that scFv-G9 had strongly binding ability to Cry1Ac toxin, but not to Cry1B, Cry1C and Cry1F toxin. The average recoveries of Cry1Ab toxin from spiked leaf and rice samples were in the range 92.1-94.8%, and 91.6-98.6%, respectively, with a coefficient of variation (C.V) less than 5.0%. These results showed promising applications of scfv-G9 for detecting Cry1Ab toxin with new DAS-ELISA. Copyright © 2014 Elsevier Ltd. All rights reserved.
-
Ordóñez, Adriana; Pérez, Juan; Tan, Lu; Dickens, Jennifer A; Motamedi-Shad, Neda; Irving, James A; Haq, Imran; Ekeowa, Ugo; Marciniak, Stefan J; Miranda, Elena; Lomas, David A
2015-06-01
Mutant Z α1-antitrypsin (E342K) accumulates as polymers within the endoplasmic reticulum (ER) of hepatocytes predisposing to liver disease, whereas low levels of circulating Z α1-antitrypsin lead to emphysema by loss of inhibition of neutrophil elastase. The ideal therapy should prevent polymer formation while preserving inhibitory activity. Here we used mAb technology to identify interactors with Z α1-antitrypsin that comply with both requirements. We report the generation of an mAb (4B12) that blocked α1-antitrypsin polymerization in vitro at a 1:1 molar ratio, causing a small increase of the stoichiometry of inhibition for neutrophil elastase. A single-chain variable fragment (scFv) intrabody was generated based on the sequence of mAb4B12. The expression of scFv4B12 within the ER (scFv4B12KDEL) and along the secretory pathway (scFv4B12) reduced the intracellular polymerization of Z α1-antitrypsin by 60%. The scFv4B12 intrabody also increased the secretion of Z α1-antitrypsin that retained inhibitory activity against neutrophil elastase. MAb4B12 recognized a discontinuous epitope probably located in the region of helices A/C/G/H/I and seems to act by altering protein dynamics rather than binding preferentially to the native state. This novel approach could reveal new target sites for small-molecule intervention that may block the transition to aberrant polymers without compromising the inhibitory activity of Z α1-antitrypsin. © FASEB.
-
Hu, Zu-Quan; Liu, Jin-Long; Li, He-Ping; Xing, Shu; Xue, Sheng; Zhang, Jing-Bo; Wang, Jian-Hua; Nölke, Greta; Liao, Yu-Cai
2012-01-01
Fusarium verticillioides is the primary causal agent of Fusarium ear and kernel rot in maize, producing fumonisin mycotoxins that are toxic to humans and domestic animals. Rapid detection and monitoring of fumonisin-producing fungi are pivotally important for the prevention of mycotoxins from entering into food/feed products. Chicken-derived single-chain variable fragments (scFvs) against cell wall-bound proteins from F. verticillioides were isolated from an immunocompetent phage display library. Comparative phage enzyme-linked immunosorbant assays (ELISAs) and sequencing analyses identified four different scFv antibodies with high sensitivity. Soluble antibody ELISAs identified two highly sensitive scFv antibodies, FvCA3 and FvCA4, with the latter being slightly more sensitive. Three-dimensional modeling revealed that the FvCA4 may hold a better overall structure with CDRH3, CDRL1 and CDRL3 centered in the core region of antibody surface compared with that of other scFvs. Immunofluorescence labeling revealed that the binding of FvCA4 antibody was localized to the cell walls of conidiospores and hyphae of F. verticillioides, confirming the specificity of this antibody for a surface target. This scFv antibody was able to detect the fungal mycelium as low as 10−2 μg/mL and contaminating mycelium at a quantity of 10−2 mg/g maize. This is the first report that scFv antibodies derived from phage display have a wide application for rapid and accurate detection and monitoring of fumonisin-producing pathogens in agricultural samples. PMID:22837678
-
NASA Astrophysics Data System (ADS)
von Trentini, F.; Willkofer, F.; Wood, R. R.; Schmid, F. J.; Ludwig, R.
2017-12-01
The ClimEx project (Climate change and hydrological extreme events - risks and perspectives for water management in Bavaria and Québec) focuses on the effects of climate change on hydro-meteorological extreme events and their implications for water management in Bavaria and Québec. Therefore, a hydro-meteorological model chain is applied. It employs high performance computing capacity of the Leibniz Supercomputing Centre facility SuperMUC to dynamically downscale 50 members of the Global Circulation Model CanESM2 over European and Eastern North American domains using the Canadian Regional Climate Model (RCM) CRCM5. Over Europe, the unique single model ensemble is conjointly analyzed with the latest information provided through the CORDEX-initiative, to better assess the influence of natural climate variability and climatic change in the dynamics of extreme events. Furthermore, these 50 members of a single RCM will enhance extreme value statistics (extreme return periods) by exploiting the available 1500 model years for the reference period from 1981 to 2010. Hence, the RCM output is applied to drive the process based, fully distributed, and deterministic hydrological model WaSiM in high temporal (3h) and spatial (500m) resolution. WaSiM and the large ensemble are further used to derive a variety of hydro-meteorological patterns leading to severe flood events. A tool for virtual perfect prediction shall provide a combination of optimal lead time and management strategy to mitigate certain flood events following these patterns.
-
NASA Astrophysics Data System (ADS)
Fraedrich, K.
2014-12-01
Processes along the continental rainfall-runoff chain cover a wide range of time and space scales which are presented here combining observations (ranging from minutes to decades) and minimalist concepts. (i) Rainfall, which can be simulated by a censored first-order autoregressive process (vertical moisture fluxes), exhibits 1/f-spectra if presented as binary events (tropics), while extrema world wide increase with duration according to Jennings' scaling law. (ii) Runoff volatility (Yangtze) shows data collapse which, linked to an intra-annual 1/f-spectrum, is represented by a single function not unlike physical systems at criticality and the short and long return times of extremes are Weibull-distributed. Atmospheric and soil moisture variabilities are also discussed. (iii) Soil moisture (in a bucket), whose variability is interpreted by a biased coinflip Ansatz for rainfall events, adds an equation of state to energy and water flux balances comprising Budyko's frame work for quasi-stationary watershed analysis. Eco-hydrologic state space presentations in terms of surface flux ratios of energy excess (loss by sensible heat over supply by net radiation) versus water excess (loss by discharge over gain by precipitation) allow attributions of state change to external (or climate) and internal (or anthropogenic) causes. Including the vegetation-greenness index (NDVI) as an active tracer extends the eco-hydrologic state space analysis to supplement the common geographical presentations. Two examples demonstrate the approach combining ERA and MODIS data sets: (a) global geobotanic classification by combining first and second moments of the dryness ratio (net radiation over precipitation) and (b) regional attributions (Tibetan Plateau) of vegetation changes.
-
Chen, Yu-Ping; Qiao, Yuan-Yuan; Zhao, Xiao-Hang; Chen, Hong-Song; Wang, Yan; Wang, Zhuozhi
2007-06-01
Bispecific antibodies have immense potential for use in clinical applications. In the present study, a bispecific diabody against human red blood cells (RBCs) and hepatitis B virus surface antigen (HBsAg) was used to detect HBsAg in blood specimens. The bispecific diabody was constructed by crossing over the variable region of the heavy chains and the light chains of anti-RBC and anti-HBsAg antibodies with a short linker, SRGGGS. In enzyme-linked immunosorbent assays, this bispecific diabody showed specific binding to both RBCs and HBsAg. When this bispecific diabody was mixed with human blood specimens in the presence of HBsAg, the dual binding sites of the diabody caused agglutination of human RBCs. This diabody-mediated agglutination assay was then used to test 712 clinical blood specimens and showed 97.7% sensitivity and 100% specificity when the results were compared with those of the conventional immunoassay, which was used as a reference. This autologous RBC agglutination assay provides a simple approach for rapid screening for HBsAg in blood specimens.
-
Chen, Yu-Ping; Qiao, Yuan-Yuan; Zhao, Xiao-Hang; Chen, Hong-Song; Wang, Yan; Wang, Zhuozhi
2007-01-01
Bispecific antibodies have immense potential for use in clinical applications. In the present study, a bispecific diabody against human red blood cells (RBCs) and hepatitis B virus surface antigen (HBsAg) was used to detect HBsAg in blood specimens. The bispecific diabody was constructed by crossing over the variable region of the heavy chains and the light chains of anti-RBC and anti-HBsAg antibodies with a short linker, SRGGGS. In enzyme-linked immunosorbent assays, this bispecific diabody showed specific binding to both RBCs and HBsAg. When this bispecific diabody was mixed with human blood specimens in the presence of HBsAg, the dual binding sites of the diabody caused agglutination of human RBCs. This diabody-mediated agglutination assay was then used to test 712 clinical blood specimens and showed 97.7% sensitivity and 100% specificity when the results were compared with those of the conventional immunoassay, which was used as a reference. This autologous RBC agglutination assay provides a simple approach for rapid screening for HBsAg in blood specimens. PMID:17442848
-
Gireeshkumar, T R; Deepulal, P M; Chandramohanakumar, N
2015-03-01
Surface sediments samples from the Cochin estuary were measured for elemental, stable isotopic and molecular biomarkers (aliphatic hydrocarbons and fatty acids) to study the sources and distribution of sedimentary organic matter. Concentrations of total organic carbon (TOC), total nitrogen (TN) and stable isotopic ratios of carbon (δ(13)C) ranged from 0.62 to 2.74 %, 0.09 to 0.25 % and -27.5 to 21.7 ‰, respectively. Sedimentary n-alkanes ranged from 6.03 to 43.23 μg g(-1) with an average of 16.79 μg g(-1), while total fatty acids varied from 22.55 to 440.69 μg g(-1). The TOC/TN ratios and δ(13)C suggest a mixture of marine- and terrestrial-derived organic matter in the surface sediments with increasing contributions from marine-derived organic matter towards the seaward side. Long-chain n-alkanes derived from higher plants predominated the inner part of the estuary, while short-chain n-alkanes derived from planktonic sources predominated the bar mouth region. The even carbon preference of the C12-C22 n-alkanes may refer to the direct biogenic contribution from bacteria, fungi and yeast species and to the potential direct petroleum inputs. The presence of odd mid-chain n-alkanes in the sediments indicates the organic matter inputs from submerged and floating macrophytes (water hyacinth). Various molecular indices such as carbon preference index, terrestrial to aquatic ratio, average chain length and the ratios of mid-chain n-alkanes support the aforementioned inferences. The high contribution of odd and branched chain fatty acids along with very low contribution of polyunsaturated fatty acids, suggest the effective utilisation of algae-derived organic matter by bacteria and the effective recycling of labile organic matter in whole settling and deposition processes. The distributional variability of n-alkanes and fatty acids reveals the preferential utilisation of marine-derived organic matter and the selective preservation of terrestrial-derived organic matter in surface sediments of the Cochin estuary.
-
Ji, Xiaonan; Shen, Yanli; Sun, Hao; Gao, Xiangdong
2016-08-01
Human hepatocellular carcinoma (HCC) has a high rate of tumor recurrence and metastasis, resulting in shortened survival time. The function of alpha-fetoprotein (AFP) as a regulatory factor in the growth of HCC cells has been well defined. The aim of this study was to investigate the use of a novel AFP-specific single-chain variable fragment that blocked AFP and inhibited HCC cell growth. The results indicated that the anti-AFP single-chain variable fragment (scFv) induced growth inhibition of AFP-expressing HCC cell lines in vitro through induction of G1 cell cycle arrest and apoptosis. The mechanism of apoptosis probably involved with blocking AFP internalization and regulation of the PTEN/PI3K/Akt signaling network. Moreover, the anti-AFP-scFv also effectively sensitized the HepG2 cells to paclitaxel (PTX) at a lower concentration. The combination effect of PTX and anti-AFP-scFv displayed a synergistic effect on HepG2 cells both in vitro and in vivo. Our results demonstrated that targeting AFP by specific antibodies has potential immunotherapeutic efficacy in human HCC.
-
Storage and sterilization techniques: the specific role of the cold chain.
Guinebault, A
1986-01-01
Focus in this discussion is on reasons for the cold chain, management of vaccine supplies (regional storage, peripheral centers, and the clinic), the facilities and their use, monitoring the cold chain, and training participants in the cold chain. To remain active, vaccines must be maintained continuously within a specific temperature range from the moment they are produced until they are injected. This is the meaning of the cold chain. If the cold chain is broken at any point, the vaccines must be destroyed for they will have lost their effectiveness. To function properly, a cold chain requires the combined presence of efficient, reliable equipment, and of qualified, vigilant personnel at all levels. The cold chain is composed of the following elements: a national storage center, near an international airport, with a 1-year supply of vaccines for the entire target population; regional storage centers with a 3-month supply of vaccines for the entire population of the region; peripheral immunization centers scattered throughout the region, managing a supply for about 1 month; clinics, which either perform vaccinations on the spot and/or supply mobile teams, depending on the strategy; and mobile teams, with portable cold boxes, with an autonomy of several days. The main problems occur at the local levels, and more specifically with respect to transportation and the fuel and power supplies, as well as cold packs. At the central level, the 1-year supply of vaccines generally is stored in cold rooms. Personnel in charge of central strorage also are responsible for transportation to and from these cold rooms. Once the space required for storing vaccines is determined, the facilities required at each level may be evaluated. The information essential to the choice must be considered in each case. The main criteria involved are outlined. There are many devices for monitoring the function of the cold chain: indicators, which accompany the vaccines from the central depot to the peripheral centers show any excesses in temperature and their duration; and devices such as thermometers show the present temperature, independently of the "history" of the vaccine. Some devices are available for checking individual elements of the cold chain from time to time. The World Health Organization (WHO) has developed a training strategy aimed at people on all levels: international consultants and decisionmakers involved in programming the Expanded Program on Immunization; technicians in charge of maintenance; and medical personnel.
-
Tachedjian, Mary; Wang, Lin-Fa
2010-01-01
Bats are the natural host reservoir for range of emerging and re-emerging viruses, many of which cause significant morbidity and mortality in other mammals, yet appear to result in no clinical consequences for bats. The ability of bats to coexist with a variety of viruses presents an interesting immunological problem that has not been examined in any detail but which could provide significant insights into the evolution of antiviral mechanisms in mammals. Towards a better understanding of the bat immune system, we analysed the expressed heavy chain variable (VH) regions of antibodies from the black flying fox, Pteropus alecto. The germline repertoire of the closely related Pteropid bat, Pteropus vampyrus, whose genome has been sequenced was also examined for comparative purposes. Representative VH genes were found in all three mammalian VH clans (I, II and III) in both the expressed P. alecto VH repertoire and the germline P. vampyrus VH repertoire. Evidence for the use of multiple heavy chain diversity (DH) and joining (JH) segments for the generation of diverse VDJ rearrangements was also present in the expressed antibody repertoire of P. alecto. The long period of co-evolutionary history of bats with viruses may have resulted in a variety of highly specific VH segments being hardwired into the genomes of bats and may have implications for their ability to successfully cope with a diversity of viral antigens. PMID:20162414
-
Kito, Naoko; Kita, Akihiro; Imokawa, Yuuki; Yamanaka, Kazuya; Maruyama, Chitose; Katano, Hajime
2014-01-01
ε-Poly-l-lysine (ε-PL), consisting of 25 to 35 l-lysine residues with linkages between the α-carboxyl groups and ε-amino groups, is produced by Streptomyces albulus NBRC14147. ε-PL synthetase (Pls) is a membrane protein with six transmembrane domains (TM1 to TM6) as well as both an adenylation domain and a thiolation domain, characteristic of the nonribosomal peptide synthetases. Pls directly generates ε-PL chain length diversity (25- to 35-mer), but the processes that control the chain length of ε-PL during the polymerization reaction are still not fully understood. Here, we report on the identification of Pls amino acid residues involved in the regulation of the ε-PL chain length. From approximately 12,000 variants generated by random mutagenesis, we found 8 Pls variants that produced shorter chains of ε-PL. These variants have one or more mutations in two linker regions connecting the TM1 and TM2 domains and the TM3 and TM4 domains. In the Pls catalytic mechanism, the growing chain of ε-PL is not tethered to the enzyme, implying that the enzyme must hold the growing chain until the polymerization reaction is complete. Our findings reveal that the linker regions are important contributors to grasp the growing chain of ε-PL. PMID:24907331
-
Tandem UIMs confer Lys48 ubiquitin chain substrate preference to deubiquitinase USP25
Kawaguchi, Kohei; Uo, Kazune; Tanaka, Toshiaki; Komada, Masayuki
2017-01-01
Ubiquitin-specific protease (USP) 25, belonging to the USP family of deubiquitinases, harbors two tandem ubiquitin-interacting motifs (UIMs), a ~20-amino-acid α-helical stretch that binds to ubiquitin. However, the role of the UIMs in USP25 remains unclear. Here we show that the tandem UIM region binds to Lys48-, but not Lys63-, linked ubiquitin chains, where the two UIMs played a critical and cooperative role. Purified USP25 exhibited higher ubiquitin isopeptidase activity to Lys48-, than to Lys63-, linked ubiquitin chains. Mutations that disrupted the ubiquitin-binding ability of the tandem UIMs resulted in a reduced ubiquitin isopeptidase activity of USP25, suggesting a role for the UIMs in exerting the full catalytic activity of USP25. Moreover, when mutations that convert the binding preference from Lys48- to Lys63-linked ubiquitin chains were introduced into the tandem UIM region, the USP25 mutants acquired elevated and reduced isopeptidase activity toward Lys63- and Lys48-linked ubiquitin chains, respectively. These results suggested that the binding preference of the tandem UIMs toward Lys48-linked ubiquitin chains contributes not only to the full catalytic activity but also to the ubiquitin chain substrate preference of USP25, possibly by selectively holding the Lys48-linked ubiquitin chain substrates in the proximity of the catalytic core. PMID:28327663
-
Ben Ali, Sina-Elisabeth; Madi, Zita Erika; Hochegger, Rupert; Quist, David; Prewein, Bernhard; Haslberger, Alexander G.; Brandes, Christian
2014-01-01
Genetic mutations must be avoided during the production and use of seeds. In the European Union (EU), Directive 2001/18/EC requires any DNA construct introduced via transformation to be stable. Establishing genetic stability is critical for the approval of genetically modified organisms (GMOs). In this study, genetic stability of two GMOs was examined using high resolution melting (HRM) analysis and real-time polymerase chain reaction (PCR) employing Scorpion primers for amplification. The genetic variability of the transgenic insert and that of the flanking regions in a single oilseed rape variety (GT73) and a stacked maize (MON88017 × MON810) was studied. The GT73 and the 5' region of MON810 showed no instabilities in the examined regions. However; two out of 100 analyzed samples carried a heterozygous point mutation in the 3' region of MON810 in the stacked variety. These results were verified by direct sequencing of the amplified PCR products as well as by sequencing of cloned PCR fragments. The occurrence of the mutation suggests that the 5' region is more suitable than the 3' region for the quantification of MON810. The identification of the single nucleotide polymorphism (SNP) in a stacked event is in contrast to the results of earlier studies of the same MON810 region in a single event where no DNA polymorphism was found. PMID:25365178
-
Pan, Jian-Jung; Ramamoorthy, Gurusankar; Poulter, C. Dale
2013-01-01
Long-chain E-polyprenyl diphosphate synthases (E-PDS) catalyze repetitive addition of isopentenyl diphosphate (IPP) to the growing prenyl chain of an allylic diphosphate. The polyprenyl diphosphate products are required for the biosynthesis of ubiquinones and menaquinones required for electron transport during oxidative phosphorylation to generate ATP. In vitro, the long-chain PDSs require addition of phospholipids or detergents to the assay buffer to enhance product release and maintain efficient turnover. During preliminary assays of product chain-length with anionic, zwitterionic, and non-ionic detergents, we discovered considerable variability. Examination of a series of non-ionic PEG detergents with several long-chain E-PDSs from different organisms revealed that in vitro incubations with nonaethylene glycol monododecyl ether or Triton X-100 typically gave chain lengths that corresponded to those of the isoprenoid moieties in respiratory quinones synthesized in vivo. In contrast incubations in buffer with n-butanol, CHAPS, DMSO, n-octyl-β-glucopyranoside, or β-cyclodextrin or in buffer without detergent typically proceeded more slowly and gave a broad range of chain lengths. PMID:23802587
-
Smit, Judith J; Monteferrario, Davide; Noordermeer, Sylvie M; van Dijk, Willem J; van der Reijden, Bert A; Sixma, Titia K
2012-01-01
Activation of the NF-κB pathway requires the formation of Met1-linked ‘linear' ubiquitin chains on NEMO, which is catalysed by the Linear Ubiquitin Chain Assembly Complex (LUBAC) E3 consisting of HOIP, HOIL-1L and Sharpin. Here, we show that both LUBAC catalytic activity and LUBAC specificity for linear ubiquitin chain formation are embedded within the RING-IBR-RING (RBR) ubiquitin ligase subunit HOIP. Linear ubiquitin chain formation by HOIP proceeds via a two-step mechanism involving both RING and HECT E3-type activities. RING1-IBR catalyses the transfer of ubiquitin from the E2 onto RING2, to transiently form a HECT-like covalent thioester intermediate. Next, the ubiquitin is transferred from HOIP onto the N-terminus of a target ubiquitin. This transfer is facilitated by a unique region in the C-terminus of HOIP that we termed ‘Linear ubiquitin chain Determining Domain' (LDD), which may coordinate the acceptor ubiquitin. Consistent with this mechanism, the RING2-LDD region was found to be important for NF-κB activation in cellular assays. These data show how HOIP combines a general RBR ubiquitin ligase mechanism with unique, LDD-dependent specificity for producing linear ubiquitin chains. PMID:22863777
-
Apgar, James R; Mader, Michelle; Agostinelli, Rita; Benard, Susan; Bialek, Peter; Johnson, Mark; Gao, Yijie; Krebs, Mark; Owens, Jane; Parris, Kevin; St Andre, Michael; Svenson, Kris; Morris, Carl; Tchistiakova, Lioudmila
2016-10-01
Antibodies are an important class of biotherapeutics that offer specificity to their antigen, long half-life, effector function interaction and good manufacturability. The immunogenicity of non-human-derived antibodies, which can be a major limitation to development, has been partially overcome by humanization through complementarity-determining region (CDR) grafting onto human acceptor frameworks. The retention of foreign content in the CDR regions, however, is still a potential immunogenic liability. Here, we describe the humanization of an anti-myostatin antibody utilizing a 2-step process of traditional CDR-grafting onto a human acceptor framework, followed by a structure-guided approach to further reduce the murine content of CDR-grafted antibodies. To accomplish this, we solved the co-crystal structures of myostatin with the chimeric (Protein Databank (PDB) id 5F3B) and CDR-grafted anti-myostatin antibody (PDB id 5F3H), allowing us to computationally predict the structurally important CDR residues as well as those making significant contacts with the antigen. Structure-based rational design enabled further germlining of the CDR-grafted antibody, reducing the murine content of the antibody without affecting antigen binding. The overall "humanness" was increased for both the light and heavy chain variable regions.
-
Apgar, James R.; Mader, Michelle; Agostinelli, Rita; Benard, Susan; Bialek, Peter; Johnson, Mark; Gao, Yijie; Krebs, Mark; Owens, Jane; Parris, Kevin; St. Andre, Michael; Svenson, Kris; Morris, Carl; Tchistiakova, Lioudmila
2016-01-01
ABSTRACT Antibodies are an important class of biotherapeutics that offer specificity to their antigen, long half-life, effector function interaction and good manufacturability. The immunogenicity of non-human-derived antibodies, which can be a major limitation to development, has been partially overcome by humanization through complementarity-determining region (CDR) grafting onto human acceptor frameworks. The retention of foreign content in the CDR regions, however, is still a potential immunogenic liability. Here, we describe the humanization of an anti-myostatin antibody utilizing a 2-step process of traditional CDR-grafting onto a human acceptor framework, followed by a structure-guided approach to further reduce the murine content of CDR-grafted antibodies. To accomplish this, we solved the co-crystal structures of myostatin with the chimeric (Protein Databank (PDB) id 5F3B) and CDR-grafted anti-myostatin antibody (PDB id 5F3H), allowing us to computationally predict the structurally important CDR residues as well as those making significant contacts with the antigen. Structure-based rational design enabled further germlining of the CDR-grafted antibody, reducing the murine content of the antibody without affecting antigen binding. The overall “humanness” was increased for both the light and heavy chain variable regions. PMID:27625211
-
Variation, Repetition, and Choice
ERIC Educational Resources Information Center
Abreu-Rodrigues, Josele; Lattal, Kennon A.; dos Santos, Cristiano V.; Matos, Ricardo A.
2005-01-01
Experiment 1 investigated the controlling properties of variability contingencies on choice between repeated and variable responding. Pigeons were exposed to concurrent-chains schedules with two alternatives. In the REPEAT alternative, reinforcers in the terminal link depended on a single sequence of four responses. In the VARY alternative, a…
-
General phase spaces: from discrete variables to rotor and continuum limits
NASA Astrophysics Data System (ADS)
Albert, Victor V.; Pascazio, Saverio; Devoret, Michel H.
2017-12-01
We provide a basic introduction to discrete-variable, rotor, and continuous-variable quantum phase spaces, explaining how the latter two can be understood as limiting cases of the first. We extend the limit-taking procedures used to travel between phase spaces to a general class of Hamiltonians (including many local stabilizer codes) and provide six examples: the Harper equation, the Baxter parafermionic spin chain, the Rabi model, the Kitaev toric code, the Haah cubic code (which we generalize to qudits), and the Kitaev honeycomb model. We obtain continuous-variable generalizations of all models, some of which are novel. The Baxter model is mapped to a chain of coupled oscillators and the Rabi model to the optomechanical radiation pressure Hamiltonian. The procedures also yield rotor versions of all models, five of which are novel many-body extensions of the almost Mathieu equation. The toric and cubic codes are mapped to lattice models of rotors, with the toric code case related to U(1) lattice gauge theory.
-
Manufacture of threads with variable pitch by using noncircular gears
NASA Astrophysics Data System (ADS)
Slătineanu, L.; Dodun, O.; Coteață, M.; Coman, I.; Nagîț, G.; Beșliu, I.
2016-08-01
There are mechanical equipments in which shafts threaded with variable pitch are included. Such a shaft could be met in the case of worm specific to the double enveloping worm gearing. Over the years, the researchers investigated some possibilities to geometrically define and manufacture the shaft zones characterized by a variable pitch. One of the methods able to facilitate the manufacture of threads with variable pitch is based on the use of noncircular gears in the threading kinematic chain for threading by cutting. In order to design the noncircular gears, the mathematical law of pitch variation has to be known. An analysis of pitch variation based on geometrical considerations was developed in the case of a double enveloping globoid worm. Subsequently, on the bases of a proper situation, a numerical model was determined. In this way, an approximately law of pitch variation was determined and it could be taken into consideration when designing the noncircular gears included in the kinematic chain of the cutting machine tool.
-
Tramontano, A; Bianchi, E; Venturini, S; Martin, F; Pessi, A; Sollazzo, M
1994-03-01
Conformationally constraining selectable peptides onto a suitable scaffold that enables their conformation to be predicted or readily determined by experimental techniques would considerably boost the drug discovery process by reducing the gap between the discovery of a peptide lead and the design of a peptidomimetic with a more desirable pharmacological profile. With this in mind, we designed the minibody, a 61-residue beta-protein aimed at retaining some desirable features of immunoglobulin variable domains, such as tolerance to sequence variability in selected regions of the protein and predictability of the main chain conformation of the same regions, based on the 'canonical structures' model. To test the ability of the minibody scaffold to support functional sites we also designed a metal binding version of the protein by suitably choosing the sequences of its loops. The minibody was produced both by chemical synthesis and expression in E. coli and characterized by size exclusion chromatography, UV CD (circular dichroism) spectroscopy and metal binding activity. All our data supported the model, but a more detailed structural characterization of the molecule was impaired by its low solubility. We were able to overcome this problem both by further mutagenesis of the framework and by addition of a solubilizing motif. The minibody is being used to select constrained human IL-6 peptidic ligands from a library displayed on the surface of the f1 bacteriophage.
-
Molecular Variability Among Isolates of Prunus Necrotic Ringspot Virus from Different Prunus spp.
Aparicio, F; Myrta, A; Di Terlizzi, B; Pallás, V
1999-11-01
ABSTRACT Viral sequences amplified by polymerase chain reaction from 25 isolates of Prunus necrotic ringspot virus (PNRSV), varying in the symptomatology they cause in six different Prunus spp., were analyzed for restriction fragment polymorphisms. Most of the isolates could be discriminated by using a combination of three different restriction enzymes. The nucleotide sequences of the RNA 4 of 15 of these isolates were determined. Sequence comparisons and phylogenetic analyses of the RNA 4 and coat proteins (CPs) revealed that all of the isolates clustered into three different groups, represented by three previously sequenced PNRSV isolates: PV32, PE5, and PV96. The PE5-type group was characterized by a 5' untranslated region that was clearly different from that of the other two groups. The PV32-type group was characterized by an extra hexanucleotide consisting of a duplication of the six immediately preceding nucleotides. Although most of the variability was observed in the first third of the CP, the amino acid residues in this region, which were previously thought to be functionally important in the replication cycle of the virus, were strictly conserved. No clear correlation with the type of symptom or host specificity could be observed. The validity of this grouping was confirmed when other isolates recently characterized by other authors were included in these analyses.
-
Zurawski, S M; Zurawski, G
1989-01-01
The function of two alpha-helical regions of mouse interleukin-2 were analyzed by saturation substitution analysis. The functional parts of the first alpha-helix (A) was defined as residues 31-39 by the observation that proline substitutions within this region inactivate the protein. Four residues within alpha-helix A, Leu31, Asp34, Leu35 and Leu38, were found to be crucial for biological activity. Structural modeling suggested that these four residues are clustered on one face of alpha-helix A. Residues 31 and 35 had to remain hydrophobic for the molecule to be functional. At residue 38 there was a preference for hydrophobic side chain residues, while at residue 34 some small side chain residues as well as acidic or amide side chain residues were functionally acceptable. Inactivating changes at residue 34 had no effect upon the ability of the protein to interact with the p55 receptor. Disruption of the fifth alpha-helix (E), which had little effect upon biological activity, resulted in an inability of the protein to interact with the p55 receptor. Mutagenesis of the alpha-helix E region demonstrated that alpha-helicity and the nature of the side chain residues in this region were unimportant for biological activity. The region immediately proximal to alpha-helix E was important only for the single intramolecular disulfide linkage. PMID:2583124
-
Cronin, T. M.; Dwyer, G.S.; Baker, P.A.; Rodriguez-Lazaro, J.; DeMartino, D.M.
2000-01-01
Magnesium/calcium (Mg/Ca) ratios were measured in the deep-sea ostracod (Crustacea) genus Krithe from Chain core 82-24-4PC from the western mid-Atlantic Ridge (3427 m) in order to estimate ocean circulation and bottom water temperature (BWT) variability over the past 200,000 years. Mg/Ca ratios have been used as a paleothermometer because the ratios are controlled primarily by ambient water temperatures at the time the organism secretes its adult carapace. Over the past two glacial–interglacial cycles, Mg/Ca values oscillated between about 7 mmol/mol and 12 mmol/mol, equivalent to a BWT range of 0 to >3.5°C. The lowest values were obtained on specimens from glacial marine isotope stages (MISs) 2, 4 and 6; the highest values were obtained from specimens from the early part of the Holocene interglacial (MIS 1), and also from MISs 5 and 7. These trends suggest that BWTs in the North Atlantic Ocean fluctuate over orbital time scales.Suborbital variability in Mg/Ca ratios and BWT was also observed for the past 100,000 years. Ratios rose from ∼8 mmol/mol to ∼10 mmol/mol (implying a BWT increase of ∼1 to 3°C) during 14 Mg/Ca excursions. The highest ratios were found in Krithe dated at approximately 32, 36–38, 43, 48, 73, 85 and 93 ka. Although the age model for the Chain 82-24-4PC and temporal resolution do not allow precise correlation, some of these deep-sea bottom temperature excursions appear to correspond to Heinrich events recorded in other regions of the North Atlantic and perhaps Dansgaard–Oeschger interstadial events recorded in Greenland ice cores. If confirmed, this would support the hypothesis that millennial-scale oscillations of climate in the North Atlantic are capable of affecting global climate via thermohaline circulation changes.
-
Verification of sex from harvested sea otters using DNA testing
Scribner, Kim T.; Green, Ben A.; Gorbics, Carol; Bodkin, James L.
2005-01-01
We used molecular genetic methods to determine the sex of 138 sea otters (Enhydra lutris) harvested from 3 regions of Alaska from 1994 to 1997, to assess the accuracy of post‐harvest field‐sexing. We also tested each of a series of factors associated with errors in field‐sexing of sea otters, including male or female bias, age‐class bias, regional bias, and bias associated with hunt characteristics. Blind control results indicated that sex was determined with 100% accuracy using polymerase chain reaction (PCR) amplification using primers that co‐amplify the zinc finger‐Y‐X gene, located on both the mammalian Y‐ and X‐chromosomes, and Testes Determining Factor (TDF), located on the mammalian Y‐chromosome. DNA‐based sexing revealed that 12.3% of the harvested sea otters were incorrectly sexed in the field, with most errors (13 of 17) occurring as males incorrectly reported as females. Thus, female harvest was overestimated. Using logistic regression analysis, we detected no statistical association of incorrect determination of sex in the field with age class, hunt region, or hunt type. The error in field‐sexing appears to be random, at least with respect to the variables evaluated in this study.
-
2009-12-02
an overarching regional information technology architecture to synchronize the vessel arrival and departure schedules with marine terminal, short... wakeup call for the rest of the nation’s ports and regions. The impacts of these disruptions are felt throughout the supply chain - in the national...and equipment repositioning. Supply chain unreliability wreaks havoc with planning, scheduling , purchasing, sales, and distribution. Suppliers are
-
Time-space mapping of Easter Chain volcanism
NASA Astrophysics Data System (ADS)
O'Connor, John M.; Stoffers, Peter; McWilliams, Michael O.
1995-12-01
New 40Ar/ 39Ar and published K sbnd Ar ages show that the locus of volcanism along the Easter Volcanic Chain (EVC) has shifted systematically from the Nazca Ridge, at about 26 m.y., to the recently active Sala y Gomez Island/Easter Island region. This indicates a plume rather than a hotline (i.e., mantle roll) origin for the EVC. The time-space distribution of ages, combined with published ages for the Galapagos and Juan Fernandez volcanic chains, is used to reconstruct Nazca plate velocities over the past 26 m.y. A plume now located in the region of Sala y Gomez Island is most compatible with these data. West of the plume, the EVC records neither Nazca nor Pacific plate motions. This section of the EVC may be related to westward channeling of plume material to the Pacific-Nazca spreading boundary region.
-
DOE Office of Scientific and Technical Information (OSTI.GOV)
Mohiyaddin, Fahd A.; Kalra, Rachpon; Laucht, Arne
The ability to transport quantum information across some distance can facilitate the design and operation of a quantum processor. One-dimensional spin chains provide a compact platform to realize scalable spin transport for a solid-state quantum computer. Here, we model odd-sized donor chains in silicon under a range of experimental nonidealities, including variability of donor position within the chain. We show that the tolerance against donor placement inaccuracies is greatly improved by operating the spin chain in a mode where the electrons are confined at the Si-SiO 2 interface. We then estimate the required time scales and exchange couplings, and themore » level of noise that can be tolerated to achieve high-fidelity transport. As a result, we also propose a protocol to calibrate and initialize the chain, thereby providing a complete guideline for realizing a functional donor chain and utilizing it for spin transport.« less
-
Grußmayer, Kristin S; Steiner, Florian; Lupton, John M; Herten, Dirk-Peter; Vogelsang, Jan
2015-12-01
Blinking of the photoluminescence (PL) emitted from individual conjugated polymer chains is one of the central observations made by single-molecule spectroscopy (SMS). Important information, for example regarding excitation energy transfer, can be extracted by evaluating dynamic quenching. However, the nature of trap states, which are responsible for PL quenching, often remains obscured. We present a detailed investigation of the photon statistics of single poly(3-hexylthiophene) (P3HT) chains obtained by SMS. The photon statistics provide a measure of the number and brightness of independently emitting areas on a single chain. These observables can be followed during blinking. A decrease in PL intensity is shown to be correlated with either 1) a decrease in the average brightness of the emitting sites; or 2) a decrease in the number of emitting regions. We attribute these phenomena to the formation of 1) shallow charge traps, which can weakly affect all emitting areas of a single chain at once; and 2) deep traps, which have a strong effect on small regions within the single chains. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
-
Crossover transition in flowing granular chains
NASA Astrophysics Data System (ADS)
Ulrich, Xialing; Fried, Eliot; Shen, Amy Q.
2009-09-01
We report on the dynamical and statistical behavior of flowing collections of granular chains confined two-dimensionally (2D) within a rotating tumbler. Experiments are conducted with systems of chains of fixed length, but various lengths are considered. The dynamics are punctuated by cascades of chains along a free-surface cascades, which drive the development of mixed porous/laminar packing arrangements in bulk. We investigate the conformation of the system, as characterized by the porosity of the flow region occupied by the chains and the mean-square end-to-end distance of the chains during flow. Both of these measures show crossover transitions from a 2D self-avoiding walk to a 2D random walk when the chain length becomes long enough to allow self-contact.
-
Expansion of the Preimmune Antibody Repertoire by Junctional Diversity in Bos taurus
Liljavirta, Jenni; Niku, Mikael; Pessa-Morikawa, Tiina; Ekman, Anna; Iivanainen, Antti
2014-01-01
Cattle have a limited range of immunoglobulin genes which are further diversified by antigen independent somatic hypermutation in fetuses. Junctional diversity generated during somatic recombination contributes to antibody diversity but its relative significance has not been comprehensively studied. We have investigated the importance of terminal deoxynucleotidyl transferase (TdT) -mediated junctional diversity to the bovine immunoglobulin repertoire. We also searched for new bovine heavy chain diversity (IGHD) genes as the information of the germline sequences is essential to define the junctional boundaries between gene segments. New heavy chain variable genes (IGHV) were explored to address the gene usage in the fetal recombinations. Our bioinformatics search revealed five new IGHD genes, which included the longest IGHD reported so far, 154 bp. By genomic sequencing we found 26 new IGHV sequences that represent potentially new IGHV genes or allelic variants. Sequence analysis of immunoglobulin heavy chain cDNA libraries of fetal bone marrow, ileum and spleen showed 0 to 36 nontemplated N-nucleotide additions between variable, diversity and joining genes. A maximum of 8 N nucleotides were also identified in the light chains. The junctional base profile was biased towards A and T nucleotide additions (64% in heavy chain VD, 52% in heavy chain DJ and 61% in light chain VJ junctions) in contrast to the high G/C content which is usually observed in mice. Sequence analysis also revealed extensive exonuclease activity, providing additional diversity. B-lymphocyte specific TdT expression was detected in bovine fetal bone marrow by reverse transcription-qPCR and immunofluorescence. These results suggest that TdT-mediated junctional diversity and exonuclease activity contribute significantly to the size of the cattle preimmune antibody repertoire already in the fetal period. PMID:24926997
-
A new ChainMail approach for real-time soft tissue simulation.
Zhang, Jinao; Zhong, Yongmin; Smith, Julian; Gu, Chengfan
2016-07-03
This paper presents a new ChainMail method for real-time soft tissue simulation. This method enables the use of different material properties for chain elements to accommodate various materials. Based on the ChainMail bounding region, a new time-saving scheme is developed to improve computational efficiency for isotropic materials. The proposed method also conserves volume and strain energy. Experimental results demonstrate that the proposed ChainMail method can not only accommodate isotropic, anisotropic and heterogeneous materials but also model incompressibility and relaxation behaviors of soft tissues. Further, the proposed method can achieve real-time computational performance.
-
Jantus Lewintre, Eloisa; Reinoso Martín, Cristina; Montaner, David; Marín, Miguel; José Terol, María; Farrás, Rosa; Benet, Isabel; Calvete, Juan J; Dopazo, Joaquín; García-Conde, Javier
2009-01-01
B cell chronic lymphocytic leukemia (CLL) is a lymphoproliferative disorder with a variable clinical course. Patients with unmutated IgV(H) gene show a shorter progression-free and overall survival than patients with immunoglobulin heavy chain variable regions (IgV(H)) gene mutated. In addition, BCL6 mutations identify a subgroup of patients with high risk of progression. Gene expression was analysed in 36 early-stage patients using high-density microarrays. Around 150 genes differentially expressed were found according to IgV(H) mutations, whereas no difference was found according to BCL6 mutations. Functional profiling methods allowed us to distinguish KEGG and gene ontology terms showing coordinated gene expression changes across subgroups of CLL. We validated a set of differentially expressed genes according to IgV(H) status, scoring them as putative prognostic markers in CLL. Among them, CRY1, LPL, CD82 and DUSP22 are the ones with at least equal or superior performance to ZAP70 which is actually the most used surrogate marker of IgV(H) status.
-
Nondestructive Quantitative Sampling for Freshwater Mussels in Variable Substrate Streams
John B. Richardson; Winston Paul Smith
1994-01-01
Unionidmussels were sampled in the Big South Fork of the Cumberland River, Tennessee and Kentucky, from July to October 1988 with a chain grid of10 l-m2 quadrats. The chain grid was used to define 100-m2 areas along the stream bed by repeatedly moving the10-m2 rectangle upstream. Within each100-m
-
ERIC Educational Resources Information Center
Dogoe, Maud; Banda, Devender R.
2009-01-01
We reviewed twelve studies that used the constant time delay (CTD) procedure to teach chained tasks to individuals with developmental disabilities from years 1996-2006. Variables analyzed include types of tasks that have been taught with the procedure, how effective CTD has been in teaching participants, and whether researchers have investigated…
-
Independence of Terminal-Link Entry Rate and Immediacy in Concurrent Chains
ERIC Educational Resources Information Center
Berg, Mark E.; Grace, Randolph C.
2004-01-01
In Phase 1, 4 pigeons were trained on a three-component multiple concurrent-chains procedure in which components differed only in terms of relative terminal-link entry rate. The terminal links were variable-interval schedules and were varied across four conditions to produce immediacy ratios of 4:1, 1:4, 2:1, and 1:2. Relative terminal-link entry…
-
Genetic diversity of the human immunoglobulin heavy chain VH region.
Li, Honghua; Cui, Xiangfeng; Pramanik, Sreemanta; Chimge, Nyam-Osor
2002-12-01
The human immunoglobulin heavy chain VH region is one of the most complex regions in the human genome. The high level of diversity of this region has been shown by a number of studies. However, because of the limitations of the conventional experimental methods, it has been difficult to learn the extent of the diversity and the underlying mechanisms. This review describes a number of new genetic approaches developed in the authors' laboratory. By using these approaches, significant progress has been made in assigning different VH sequences to their respective loci, in learning the diversity of gene segment number and composition among the VH haplotypes, and in learning VH gene segment organization in individual haplotypes. Information obtained toward this direction could help in understanding the mechanisms underlying VH region diversity and the biological impact of the VH region diversity.
-
Air pollution and health impact emboided in supply chains in China
NASA Astrophysics Data System (ADS)
Zhang, Q.
2016-12-01
Close economic linkage and consequent air pollutant emissions embodied in trade among Chinese regions have been widely discussed. Yet the related health impacts across regions remain unaddressed. Here, we integrated four state-of-the-art models to for the first time estimate PM2.5 related premature deaths along the supply chains across seven Chinese regions, and we quantified cross impacts among receptors, producers, assemblers, consumers, and sectors. We find that, due to the atmospheric transport, in 2010, 33% of national premature deaths were caused by emissions released in other regions, and the trans-boundary effect is more significant from north to south and from east to west. From a supply chain perspective, 38% of national premature deaths were associated with production for in a region to supply other regions' consumption. For instance, 20-35% of premature deaths related to the highly developed east coastal regions' consumption were caused by emissions in the central and western regions. Sectorally, similar to the widely concerned heavy industries, direct emissions from agricultural and residential activities together contributed near half of national total premature deaths, posing a great challenge for recent pollution reduction action, which are mainly focusing on industrial restructuring. Our results emphasize the importance of regarding pollution related premature deaths in China as a national systemic problem, instead of targeting the pollution producers (region or sector) in isolation. Multilateral and multi-sector cooperation is in urgent need to improve the national atmospheric environment.
-
Riley, P.; Tikoff, B.; Hildreth, Wes
2012-01-01
The Long Valley region of eastern California (United States) is the site of abundant late Tertiary–present magmatism, including three geochemically distinct stages of magmatism since ca. 3 Ma: Mammoth Mountain, the Mono-Inyo volcanic chain, and Long Valley Caldera. We propose two tectonic models, one explaining the Mammoth Mountain–Mono-Inyo magmatism and the other explaining the presence of Long Valley Caldera. First, the ongoing Mammoth Mountain–Mono-Inyo volcanic chain magmatism is explained by a ridge-transform-ridge system, with the Mono-Inyo volcanic chain acting as one ridge segment and the South Moat fault acting as a transform fault. Implicit in this first model is that this region of eastern California is beginning to act as an incipient plate boundary. Second, the older Long Valley Caldera system is hypothesized to occur in a region of enhanced extension resulting from regional fault block rotation, specifically involving activation of the sinistral faults of the Mina deflection. The tectonic models are consistent with observed spatial and temporal differences in the geochemistry of the regional magmas, and the westward progression of magmatism since ca. 12 Ma.
-
NASA Astrophysics Data System (ADS)
van Westen, Thijs; Vlugt, Thijs J. H.; Gross, Joachim
2014-01-01
An analytical equation of state (EoS) is derived to describe the isotropic (I) and nematic (N) phase of linear- and partially flexible tangent hard-sphere chain fluids and their mixtures. The EoS is based on an extension of Onsager's second virial theory that was developed in our previous work [T. van Westen, B. Oyarzún, T. J. H. Vlugt, and J. Gross, J. Chem. Phys. 139, 034505 (2013)]. Higher virial coefficients are calculated using a Vega-Lago rescaling procedure, which is hereby generalized to mixtures. The EoS is used to study (1) the effect of length bidispersity on the I-N and N-N phase behavior of binary linear tangent hard-sphere chain fluid mixtures, (2) the effect of partial molecular flexibility on the binary phase diagram, and (3) the solubility of hard-sphere solutes in I- and N tangent hard-sphere chain fluids. By changing the length bidispersity, two types of phase diagrams were found. The first type is characterized by an I-N region at low pressure and a N-N demixed region at higher pressure that starts from an I-N-N triphase equilibrium. The second type does not show the I-N-N equilibrium. Instead, the N-N region starts from a lower critical point at a pressure above the I-N region. The results for the I-N region are in excellent agreement with the results from molecular simulations. It is shown that the N-N demixing is driven both by orientational and configurational/excluded volume entropy. By making the chains partially flexible, it is shown that the driving force resulting from the configurational entropy is reduced (due to a less anisotropic pair-excluded volume), resulting in a shift of the N-N demixed region to higher pressure. Compared to linear chains, no topological differences in the phase diagram were found. We show that the solubility of hard-sphere solutes decreases across the I-N phase transition. Furthermore, it is shown that by using a liquid crystal mixture as the solvent, the solubility difference can by maximized by tuning the composition. Theoretical results for the Henry's law constant of the hard-sphere solute are in good agreement with the results from molecular simulation.
-
How well-connected is the surface of the global ocean?
Froyland, Gary; Stuart, Robyn M; van Sebille, Erik
2014-09-01
The Ekman dynamics of the ocean surface circulation is known to contain attracting regions such as the great oceanic gyres and the associated garbage patches. Less well-known are the extents of the basins of attractions of these regions and how strongly attracting they are. Understanding the shape and extent of the basins of attraction sheds light on the question of the strength of connectivity of different regions of the ocean, which helps in understanding the flow of buoyant material like plastic litter. Using short flow time trajectory data from a global ocean model, we create a Markov chain model of the surface ocean dynamics. The surface ocean is not a conservative dynamical system as water in the ocean follows three-dimensional pathways, with upwelling and downwelling in certain regions. Using our Markov chain model, we easily compute net surface upwelling and downwelling, and verify that it matches observed patterns of upwelling and downwelling in the real ocean. We analyze the Markov chain to determine multiple attracting regions. Finally, using an eigenvector approach, we (i) identify the five major ocean garbage patches, (ii) partition the ocean into basins of attraction for each of the garbage patches, and (iii) partition the ocean into regions that demonstrate transient dynamics modulo the attracting garbage patches.
-
Norris, Peter M; da Silva, Arlindo M
2016-07-01
A method is presented to constrain a statistical model of sub-gridcolumn moisture variability using high-resolution satellite cloud data. The method can be used for large-scale model parameter estimation or cloud data assimilation. The gridcolumn model includes assumed probability density function (PDF) intra-layer horizontal variability and a copula-based inter-layer correlation model. The observables used in the current study are Moderate Resolution Imaging Spectroradiometer (MODIS) cloud-top pressure, brightness temperature and cloud optical thickness, but the method should be extensible to direct cloudy radiance assimilation for a small number of channels. The algorithm is a form of Bayesian inference with a Markov chain Monte Carlo (MCMC) approach to characterizing the posterior distribution. This approach is especially useful in cases where the background state is clear but cloudy observations exist. In traditional linearized data assimilation methods, a subsaturated background cannot produce clouds via any infinitesimal equilibrium perturbation, but the Monte Carlo approach is not gradient-based and allows jumps into regions of non-zero cloud probability. The current study uses a skewed-triangle distribution for layer moisture. The article also includes a discussion of the Metropolis and multiple-try Metropolis versions of MCMC.
-
NASA Technical Reports Server (NTRS)
Norris, Peter M.; Da Silva, Arlindo M.
2016-01-01
A method is presented to constrain a statistical model of sub-gridcolumn moisture variability using high-resolution satellite cloud data. The method can be used for large-scale model parameter estimation or cloud data assimilation. The gridcolumn model includes assumed probability density function (PDF) intra-layer horizontal variability and a copula-based inter-layer correlation model. The observables used in the current study are Moderate Resolution Imaging Spectroradiometer (MODIS) cloud-top pressure, brightness temperature and cloud optical thickness, but the method should be extensible to direct cloudy radiance assimilation for a small number of channels. The algorithm is a form of Bayesian inference with a Markov chain Monte Carlo (MCMC) approach to characterizing the posterior distribution. This approach is especially useful in cases where the background state is clear but cloudy observations exist. In traditional linearized data assimilation methods, a subsaturated background cannot produce clouds via any infinitesimal equilibrium perturbation, but the Monte Carlo approach is not gradient-based and allows jumps into regions of non-zero cloud probability. The current study uses a skewed-triangle distribution for layer moisture. The article also includes a discussion of the Metropolis and multiple-try Metropolis versions of MCMC.
-
Norris, Peter M.; da Silva, Arlindo M.
2018-01-01
A method is presented to constrain a statistical model of sub-gridcolumn moisture variability using high-resolution satellite cloud data. The method can be used for large-scale model parameter estimation or cloud data assimilation. The gridcolumn model includes assumed probability density function (PDF) intra-layer horizontal variability and a copula-based inter-layer correlation model. The observables used in the current study are Moderate Resolution Imaging Spectroradiometer (MODIS) cloud-top pressure, brightness temperature and cloud optical thickness, but the method should be extensible to direct cloudy radiance assimilation for a small number of channels. The algorithm is a form of Bayesian inference with a Markov chain Monte Carlo (MCMC) approach to characterizing the posterior distribution. This approach is especially useful in cases where the background state is clear but cloudy observations exist. In traditional linearized data assimilation methods, a subsaturated background cannot produce clouds via any infinitesimal equilibrium perturbation, but the Monte Carlo approach is not gradient-based and allows jumps into regions of non-zero cloud probability. The current study uses a skewed-triangle distribution for layer moisture. The article also includes a discussion of the Metropolis and multiple-try Metropolis versions of MCMC. PMID:29618847
-
Knirel, Y A; Kocharova, N A; Hynes, S O; Widmalm, G; Andersen, L P; Jansson, P E; Moran, A P
1999-11-01
In contrast to other Helicobacter pylori strains, which have serologically detectable Lewis(x)+ (Le(x)) and Lewis(y)++ (++Le(y)) antigenic determinants in the O-specific polysaccharide chains of the lipopolysaccharides, H. pylori AF1 and 007 were non-typable with anti-Le(x) and anti-Le(y) antibodies. The carbohydrate portions of the lipopolysaccharides were liberated by mild acid hydrolysis and subsequently studied by sugar and methylation analyses, 1H-NMR spectroscopy and electrospray ionization-mass spectrometry. Compared with each other, and with lipopolysaccharides of strains studied previously, the lipopolysaccharides of both AF1 and 007 showed similarities, but also differences, in the structures of the core region and O-specific polysaccharide chains. The O-specific polysaccharide chains of both strains consisted of a short or long polyfucosylated poly-N-acetyl-beta-lactosamine chains, which were distinguished from those of other strains by a high degree of fucosylation producing a polymeric Le(x)chain terminating with Le(x) or Le(y) units:[sequence: see text] where n = 0 or 1 in strain AF1 and 0 in strain 007, m = 0-2, 6-7 in strain AF1 and m = 0-2, 6-7 or approximately 40 in strain 007, the medium-size species being predominant. Therefore, compared with other strains, the lack of reactivity of lipopolysaccharide of H. pylori AF1 and 007 with anti-Le(x) and anti-Le(y) may reflect the presence of a polymeric Le(x) chain and has important implications for serological and pathogenesis studies. As the substitution pattern of a D-glycero-D-manno-heptose residue in the outer core varied in the two strains, and an extended DD-heptan chain was present in some lipopolysaccharide species but not in others, this region was less conservative than the inner core region. The inner core L-glycero-D-manno-heptose region of both strains carried a 2-aminoethyl phosphate group, rather than a phosphate group, as reported previously for other H. pylori strains.
-
Yakum, Martin Ndinakie; Ateudjieu, Jerome; Walter, Ebile Akoh; Watcho, Pierre
2015-04-14
The cold chain must be monitored continuously in order to guarantee vaccines' quality. From field reports and previous studies, cold chain monitoring for expanded program on immunization (EPI) is still not satisfactory in Cameroon. This study was conducted to evaluate the availability and functioning of cold chain equipment as well as knowledge. It was a cross-sectional study involving a multistage sampling. 3urban and 5rural districts were selected randomly from the 19 health districts of the North West region. In each district all the health facilities taking part in the EPI were targeted. Data were collected using a questionnaire administered face to face to health personnel and with an observational grid to assess availability, functioning, and monitoring of cold chain equipment and power supply. The data were analyzed using the epi-info software. A total of 70 health facilities were contacted and 65(88.6%) of them included in the study. Fifty-three (81.5%) out of 65 health facilities had at least one functional vaccine refrigerator. The national guideline of EPI was not present in 21(33.9%) health facilities. Temperature chart was complete/correctly filled in 25(50.0%) of the 50(96.2%) facilities having it. About 14 (26.9%) of the health facilities record at least one abnormal temperature during the last 2 months following data collection. Seventeen (28.3%) personnel did not know the correct vaccine storage temperature. The availability of vaccine storage equipment for EPI is acceptable in the North West Region of Cameroon but the capacity of those in charge to properly monitor it in all health facilities is still limited. To ensure that vaccines administered in the North West Region are stored at the recommended temperature, all District Health Services should train and regularly supervise the health personnel in charge of cold chain monitoring.
-
Viability assessment of regional biomass pre-processing center based bioethanol value chains
NASA Astrophysics Data System (ADS)
Carolan, Joseph E.
Petroleum accounts for 94% of all liquid fuels and 36% of the total of all energy consumed in the United States. Petroleum dependence is problematic because global petroleum reserves are estimated to last only for 40 to 60 years at current consumption rates; global supplies are often located in politically unstable or unfriendly regions; and fossil fuels have negative environmental footprints. Domestic policies have aimed at promoting alternative, renewable liquid fuels, specifically bio-fuels derived from organic matter. Cellulosic bio-ethanol is one promising alternative fuel that has featured prominently in federal bio-fuel mandates under the Energy Independence and Security Act, 2007. However, the cellulosic bio-ethanol industry faces several technical, physical and industrial organization challenges. This dissertation examines the concept of a network of regional biomass pre-treatment centers (RBPC) that form an extended biomass supply chain feeding into a simplified biorefinery as a way to overcome these challenges. The analyses conducted address the structural and transactional issues facing bio-ethanol value chain establishment; the technical and financial feasibility of a stand alone pre-treatment center (RBPC); the impact of distributed pre-treatment on biomass transport costs; a comparative systems cost evaluation of the performance of the RBPC chain versus a fully integrated biorefinery (gIBRh), followed by application of the analytical framework to three case study regions.
-
NASA Astrophysics Data System (ADS)
Mishra, Piyush; Hewett, Daniel M.; Zwier, Timothy S.
2018-05-01
The single-conformation ultraviolet and infrared spectroscopy of three short-chain para-dialkylbenzenes (para-diethylbenzene, para-dipropylbenzene, and para-dibutylbenzene) is reported for the jet-cooled, isolated molecules. The present study builds off previous work on single-chain n-alkylbenzenes, where an anharmonic local mode Hamiltonian method was developed to account for stretch-bend Fermi resonance in the alkyl CH stretch region [D. P. Tabor et al., J. Chem. Phys. 144, 224310 (2016)]. The jet-cooled molecules are interrogated using laser-induced fluorescence (LIF) excitation, fluorescence dip infrared spectroscopy, and dispersed fluorescence. The LIF spectra in the S1 ← S0 origin region show a dramatic increase in the number of resolved transitions with increasing length of the alkyl chains, reflecting an explosion in the number of unique low-energy conformations formed when two independent alkyl chains are present. Since the barriers to isomerization of the alkyl chain are similar in size, this results in an "egg carton" shaped potential energy surface. A combination of electronic frequency shift and alkyl CH stretch infrared spectra is used to generate a consistent set of conformational assignments. Using these experimental techniques in conjunction with computational methods, subsets of origin transitions in the LIF excitation spectrum can be classified into different conformational families. Two conformations are resolved in para-diethylbenzene, seven in para-dipropylbenzene, and about nineteen in para-dibutylbenzene. These chains are largely independent of each other as there are no new single-chain conformations induced by the presence of a second chain. A cursory LIF excitation scan of para-dioctylbenzene shows a broad congested spectrum at frequencies consistent with interactions of alkyl chains with the phenyl π cloud.
-
Mishra, Piyush; Hewett, Daniel M; Zwier, Timothy S
2018-05-14
The single-conformation ultraviolet and infrared spectroscopy of three short-chain para-dialkylbenzenes (para-diethylbenzene, para-dipropylbenzene, and para-dibutylbenzene) is reported for the jet-cooled, isolated molecules. The present study builds off previous work on single-chain n-alkylbenzenes, where an anharmonic local mode Hamiltonian method was developed to account for stretch-bend Fermi resonance in the alkyl CH stretch region [D. P. Tabor et al., J. Chem. Phys. 144, 224310 (2016)]. The jet-cooled molecules are interrogated using laser-induced fluorescence (LIF) excitation, fluorescence dip infrared spectroscopy, and dispersed fluorescence. The LIF spectra in the S 1 ← S 0 origin region show a dramatic increase in the number of resolved transitions with increasing length of the alkyl chains, reflecting an explosion in the number of unique low-energy conformations formed when two independent alkyl chains are present. Since the barriers to isomerization of the alkyl chain are similar in size, this results in an "egg carton" shaped potential energy surface. A combination of electronic frequency shift and alkyl CH stretch infrared spectra is used to generate a consistent set of conformational assignments. Using these experimental techniques in conjunction with computational methods, subsets of origin transitions in the LIF excitation spectrum can be classified into different conformational families. Two conformations are resolved in para-diethylbenzene, seven in para-dipropylbenzene, and about nineteen in para-dibutylbenzene. These chains are largely independent of each other as there are no new single-chain conformations induced by the presence of a second chain. A cursory LIF excitation scan of para-dioctylbenzene shows a broad congested spectrum at frequencies consistent with interactions of alkyl chains with the phenyl π cloud.
-
Cysteine Racemization on IgG Heavy and Light Chains
Zhang, Qingchun; Flynn, Gregory C.
2013-01-01
Under basic pH conditions, the heavy chain 220-light chain 214 (H220-L214) disulfide bond, found in the flexible hinge region of an IgG1, can convert to a thioether. Similar conditions also result in racemization of the H220 cysteine. Here, we report that racemization occurs on both H220 and L214 on an IgG1 with a λ light chain (IgG1λ) but almost entirely on H220 of an IgGl with a κ light chain (IgG1κ) under similar conditions. Likewise, racemization was detected at significant levels on H220 and L214 on endogenous human IgG1λ but only at the H220 position on IgG1κ. Low but measurable levels of d-cysteines were found on IgG2 cysteines in the hinge region, both with monoclonal antibodies incubated under basic pH conditions and on antibodies isolated from human serum. A simplified reaction mechanism involving reversible β-elimination on the cysteine is presented that accounts for both base-catalyzed racemization and thioether formation at the hinge disulfide. PMID:24142697
-
Temporal variation in pelagic food chain length in response to environmental change
Ruiz-Cooley, Rocio I.; Gerrodette, Tim; Fiedler, Paul C.; Chivers, Susan J.; Danil, Kerri; Ballance, Lisa T.
2017-01-01
Climate variability alters nitrogen cycling, primary productivity, and dissolved oxygen concentration in marine ecosystems. We examined the role of this variability (as measured by six variables) on food chain length (FCL) in the California Current (CC) by reconstructing a time series of amino acid–specific δ15N values derived from common dolphins, an apex pelagic predator, and using two FCL proxies. Strong declines in FCL were observed after the 1997–1999 El Niño Southern Oscillation (ENSO) event. Bayesian models revealed longer FCLs under intermediate conditions for surface temperature, chlorophyll concentration, multivariate ENSO index, and total plankton volume but not for hypoxic depth and nitrate concentration. Our results challenge the prevalent paradigm that suggested long-term stability in the food web structure in the CC and, instead, reveal that pelagic food webs respond strongly to disturbances associated with ENSO events, local oceanography, and ongoing changes in climate. PMID:29057322
-
Hamano, Yoshimitsu; Kito, Naoko; Kita, Akihiro; Imokawa, Yuuki; Yamanaka, Kazuya; Maruyama, Chitose; Katano, Hajime
2014-08-01
ε-Poly-l-lysine (ε-PL), consisting of 25 to 35 l-lysine residues with linkages between the α-carboxyl groups and ε-amino groups, is produced by Streptomyces albulus NBRC14147. ε-PL synthetase (Pls) is a membrane protein with six transmembrane domains (TM1 to TM6) as well as both an adenylation domain and a thiolation domain, characteristic of the nonribosomal peptide synthetases. Pls directly generates ε-PL chain length diversity (25- to 35-mer), but the processes that control the chain length of ε-PL during the polymerization reaction are still not fully understood. Here, we report on the identification of Pls amino acid residues involved in the regulation of the ε-PL chain length. From approximately 12,000 variants generated by random mutagenesis, we found 8 Pls variants that produced shorter chains of ε-PL. These variants have one or more mutations in two linker regions connecting the TM1 and TM2 domains and the TM3 and TM4 domains. In the Pls catalytic mechanism, the growing chain of ε-PL is not tethered to the enzyme, implying that the enzyme must hold the growing chain until the polymerization reaction is complete. Our findings reveal that the linker regions are important contributors to grasp the growing chain of ε-PL. Copyright © 2014, American Society for Microbiology. All Rights Reserved.
-
Variability and repertoire size of T-cell receptor V alpha gene segments.
Becker, D M; Pattern, P; Chien, Y; Yokota, T; Eshhar, Z; Giedlin, M; Gascoigne, N R; Goodnow, C; Wolf, R; Arai, K
The immune system of higher organisms is composed largely of two distinct cell types, B lymphocytes and T lymphocytes, each of which is independently capable of recognizing an enormous number of distinct entities through their antigen receptors; surface immunoglobulin in the case of the former, and the T-cell receptor (TCR) in the case of the latter. In both cell types, the genes encoding the antigen receptors consist of multiple gene segments which recombine during maturation to produce many possible peptides. One striking difference between B- and T-cell recognition that has not yet been resolved by the structural data is the fact that T cells generally require a major histocompatibility determinant together with an antigen whereas, in most cases, antibodies recognize antigen alone. Recently, we and others have found that a series of TCR V beta gene sequences show conservation of many of the same residues that are conserved between heavy- and light-chain immunoglobulin V regions, and these V beta sequences are predicted to have an immunoglobulin-like secondary structure. To extend these studies, we have isolated and sequenced eight additional alpha-chain complementary cDNA clones and compared them with published sequences. Analyses of these sequences, reported here, indicate that V alpha regions have many of the characteristics of V beta gene segments but differ in that they almost always occur as cross-hybridizing gene families. We conclude that there may be very different selective pressures operating on V alpha and V beta sequences and that the V alpha repertoire may be considerably larger than that of V beta.
-
Rapid polymerase chain reaction-based screening assay for bacterial biothreat agents.
Yang, Samuel; Rothman, Richard E; Hardick, Justin; Kuroki, Marcos; Hardick, Andrew; Doshi, Vishal; Ramachandran, Padmini; Gaydos, Charlotte A
2008-04-01
To design and evaluate a rapid polymerase chain reaction (PCR)-based assay for detecting Eubacteria and performing early screening for selected Class A biothreat bacterial pathogens. The authors designed a two-step PCR-based algorithm consisting of an initial broad-based universal detection step, followed by specific pathogen identification targeted for identification of the Class A bacterial biothreat agents. A region in the bacterial 16S rRNA gene containing a highly variable sequence flanked by clusters of conserved sequences was chosen as the target for the PCR assay design. A previously described highly conserved region located within the 16S rRNA amplicon was selected as the universal probe (UniProbe, Integrated DNA Technology, Coralville, IA). Pathogen-specific TaqMan probes were designed for Bacillus anthracis, Yersinia pestis, and Francisella tularensis. Performance of the assay was assessed using genomic DNA extracted from the aforementioned biothreat-related organisms (inactivated or surrogate) and other common bacteria. The UniProbe detected the presence of all tested Eubacteria (31/31) with high analytical sensitivity. The biothreat-specific probes accurately identified organisms down to the closely related species and genus level, but were unable to discriminate between very close surrogates, such as Yersinia philomiragia and Bacillus cereus. A simple, two-step PCR-based assay proved capable of both universal bacterial detection and identification of select Class A bacterial biothreat and biothreat-related pathogens. Although this assay requires confirmatory testing for definitive species identification, the method has great potential for use in ED-based settings for rapid diagnosis in cases of suspected Category A bacterial biothreat agents.
-
Metastates in Mean-Field Models with Random External Fields Generated by Markov Chains
NASA Astrophysics Data System (ADS)
Formentin, M.; Külske, C.; Reichenbachs, A.
2012-01-01
We extend the construction by Külske and Iacobelli of metastates in finite-state mean-field models in independent disorder to situations where the local disorder terms are a sample of an external ergodic Markov chain in equilibrium. We show that for non-degenerate Markov chains, the structure of the theorems is analogous to the case of i.i.d. variables when the limiting weights in the metastate are expressed with the aid of a CLT for the occupation time measure of the chain. As a new phenomenon we also show in a Potts example that for a degenerate non-reversible chain this CLT approximation is not enough, and that the metastate can have less symmetry than the symmetry of the interaction and a Gaussian approximation of disorder fluctuations would suggest.
-
Southeast regional fatal study : a causal chain analysis in North Carolina
DOT National Transportation Integrated Search
2002-01-01
This project completed the North Carolina portion of the pooled fund Southeast Fatal Analysis Project. In addition : to completing this portion of the causal chain analysis for NC Crashes, the project developed a ranked : comprehensive list of candid...
-
ERIC Educational Resources Information Center
Kyonka, Elizabeth G. E.; Grace, Randolph C.
2008-01-01
Pigeons responded in a concurrent-chains procedure in which terminal-link reinforcer variables were changed unpredictably across sessions. In Experiment 1, the terminal-link schedules were fixed-interval (FI) 8 s and FI 16 s, and the reinforcer magnitudes were 2 s and 4 s. In Experiment 2 the probability of reinforcement (100% or 50%) was varied…
-
Indonesia as seen from STS-66 shuttle Atlantis
1994-11-14
This is a striking, oblique view to the south of the Indonesian islands of Java (right), Bali and Lombok (upper left). The linear array of dark regions across the photo is a chain of volcanoes which make up the back bone of this part of the Indonesian Islands. This chain has been quite active over the past six months. Plumes of steam can be seen rising from the summits of Arjuno (west-central Java) and Merapi (central Java, near the right side of this photo). The region appears hazy due to an extended drought over Indonesia and Australia. Because of drought conditions, huge fires continue to burn over other regions of Indonesia, New Guinea and norther Australia, producing a regional smoke pall.
-
Methane Leakage from Oil & Gas Operations. What have we learned from recent studies in the U.S.?
NASA Astrophysics Data System (ADS)
Zavala-Araiza, Daniel; Hamburg, Steven
2016-04-01
Methane, the principal component of natural gas, is a powerful greenhouse gas. Methane losses from the natural gas supply chain erode the climate benefits of fuel switching to natural gas from other fossil fuels, reducing or eliminating them for several decades or longer. Global data on methane emissions from the oil and gas sector is uncertain and as a consequence, measuring and characterizing methane emissions is critical to the design of effective mitigation strategies. In this work, we synthesize lessons learned from dozens of U.S. studies that characterized methane emissions along each stage of the natural gas supply chain. These results are relevant to the design of methane measurement campaigns outside the U.S. A recurring theme in the research conducted in the U.S. is that public emissions inventories (e.g., The U.S. Environmental Protection Agency's National Greenhouse gas Inventory) tend to underestimate emissions for two key reasons: (1) use of non-representative emission factors and (2) inaccurate activity data (incomplete counts of facilities and equipment). Similarly, the accuracy of emission factors and the effectiveness of mitigation strategies are heavily affected by the existence of low-probability, unpredictable high emitters-which have been observed all along the supply chain- and are spatiotemporally variable. We conducted a coordinated campaign to measure methane emissions in a major gas producing region of the U.S. (Barnett Shale region of Texas) using a diversity of approaches. As part of this study we identified methods for effective quantification of regional fossil methane emissions using atmospheric data (through replicate mass balance flights and source apportionment using methane to ethane ratios) as well as how to build an accurate inventory that includes a statistical estimator that more rigorously captures the magnitude and frequency of high emitters. We found agreement between large-scale atmospheric sampling estimates and source-based estimates (custom inventory). With measured oil and gas methane being roughly twice what estimates based on the U.S. Environmental Protection Agency's National Greenhouse gas Inventory would suggest. Ten percent of oil and gas facilities in the region -the high emitters or fat tail of the distribution- account for 90% of the emissions. We observed significant regional heterogeneity (e.g., local practices, technologies used, physical properties of the reservoirs) during the production, processing, transportation, and use of natural gas, describing this heterogeneity is critical to constructing accurate methane emission inventories. The lessons learned in the U.S. provide robust methodological guidelines that can be used to extend our understanding of the climatic implications of global oil and gas methane emissions with regards to, accurate quantification, reporting, and mitigation of methane emissions.
-
Srinivasan, A R; Yathindra, N
1977-01-01
A novel description of the conformational characteristics of all the individual nucleotides and the phosphodiesters in tRNAs is presented in the form of a circular plot. This representation furnishes information of the base sequence with the folding patterns of the polynucleotide chain as one traverses along the circumference and with the individual nucleotide and phosphodiester linkage torsions along the radii. The circular plot obtained for yeast tRNAPhe strikingly distinguishes the helical and the loop regions. The variation of the different nucleotide torsions along the entire chain length and their effect on the secondary helical and tertiary loop regions become readily apparent. PMID:339206
-
Effect of vehicular size on chain-reaction crash
NASA Astrophysics Data System (ADS)
Nagatani, Takashi
2015-11-01
We present the dynamic model of the chain-reaction crash to take account of the vehicular size. Drivers brake according to taillights of the forward vehicle. We investigate the effect of the vehicular size on the chain-reaction crash (multiple-vehicle collision) in the traffic flow controlled by taillights. In the multiple-vehicle collision, the first crash induces more collisions. We investigate how the first collision induces the chain-reaction crash numerically. We derive, analytically, the transition points and the region maps for the chain-reaction crash in the traffic flow of vehicles with finite sizes. We clarify the effect of the vehicular size on the multiple-vehicle collision.
-
Determinants of choice for pigeons and humans on concurrent-chains schedules of reinforcement.
Belke, T W; Pierce, W D; Powell, R A
1989-09-01
Concurrent-chains schedules of reinforcement were arranged for humans and pigeons. Responses of humans were reinforced with tokens exchangeable for money, and key pecks of 4 birds were reinforced with food. Variable-interval 30-s and 40-s schedules operated in the terminal links of the chains. Condition 1 exposed subjects to variable-interval 90-s and variable-interval 30-s initial links, respectively. Conditions 2 and 3 arranged equal initial-link schedules of 40 s or 120 s. Experimental conditions tested the descriptive adequacy of five equations: reinforcement density, delay reduction, modified delay reduction, matching and maximization. Results based on choice proportions and switch rates during the initial links showed that pigeons behaved in accord with delay-reduction models, whereas humans maximized overall rate of reinforcement. As discussed by Logue and associates in self-control research, different types of reinforcement may affect sensitivity to delay differentially. Pigeons' responses were reinforced with food, a reinforcer that is consumable upon presentation. Humans' responses were reinforced with money, a reinforcer exchanged for consumable reinforcers after it was earned. Reinforcers that are immediately consumed may generate high sensitivity to delay and behavior described as delay reduction. Reinforces with longer times to consumption may generate low sensitivity to delay and behavior that maximizes overall payoff.
-
Resche-Rigon, Matthieu; White, Ian R
2018-06-01
In multilevel settings such as individual participant data meta-analysis, a variable is 'systematically missing' if it is wholly missing in some clusters and 'sporadically missing' if it is partly missing in some clusters. Previously proposed methods to impute incomplete multilevel data handle either systematically or sporadically missing data, but frequently both patterns are observed. We describe a new multiple imputation by chained equations (MICE) algorithm for multilevel data with arbitrary patterns of systematically and sporadically missing variables. The algorithm is described for multilevel normal data but can easily be extended for other variable types. We first propose two methods for imputing a single incomplete variable: an extension of an existing method and a new two-stage method which conveniently allows for heteroscedastic data. We then discuss the difficulties of imputing missing values in several variables in multilevel data using MICE, and show that even the simplest joint multilevel model implies conditional models which involve cluster means and heteroscedasticity. However, a simulation study finds that the proposed methods can be successfully combined in a multilevel MICE procedure, even when cluster means are not included in the imputation models.
-
Kuffner, Ilsa B.; Brock, John C.; Grober-Dunsmore, Rikki; Bonito, Victor E.; Hickey, T. Donald; Wright, C. Wayne
2007-01-01
The realization that coral reef ecosystem management must occur across multiple spatial scales and habitat types has led scientists and resource managers to seek variables that are easily measured over large areas and correlate well with reef resources. Here we investigate the utility of new technology in airborne laser surveying (NASA Experimental Advanced Airborne Research Lidar (EAARL)) in assessing topographical complexity (rugosity) to predict reef fish community structure on shallow (n = 10–13 per reef). Rugosity at each station was assessed in situ by divers using the traditional chain-transect method (10-m scale), and remotely using the EAARL submarine topography data at multiple spatial scales (2, 5, and 10 m). The rugosity and biological datasets were analyzed together to elucidate the predictive power of EAARL rugosity in describing the variance in reef fish community variables and to assess the correlation between chain-transect and EAARL rugosity. EAARL rugosity was not well correlated with chain-transect rugosity, or with species richness of fishes (although statistically significant, the amount of variance explained by the model was very low). Variance in reef fish community attributes was better explained in reef-by-reef variability than by physical variables. However, once the reef-by-reef variability was taken into account in a two-way analysis of variance, the importance of rugosity could be seen on individual reefs. Fish species richness and abundance were statistically higher at high rugosity stations compared to medium and low rugosity stations, as predicted by prior ecological research. The EAARL shows promise as an important mapping tool for reef resource managers as they strive to inventory and protect coral reef resources.
-
Lee, K-E; Lee, E-J; Park, H-S
2016-08-30
Recent advances in computational epigenetics have provided new opportunities to evaluate n-gram probabilistic language models. In this paper, we describe a systematic genome-wide approach for predicting functional roles in inactive chromatin regions by using a sequence-based Markovian chromatin map of the human genome. We demonstrate that Markov chains of sequences can be used as a precursor to predict functional roles in heterochromatin regions and provide an example comparing two publicly available chromatin annotations of large-scale epigenomics projects: ENCODE project consortium and Roadmap Epigenomics consortium.
-
Chain-reaction crash in traffic flow controlled by taillights
NASA Astrophysics Data System (ADS)
Nagatani, Takashi
2015-02-01
We study the chain-reaction crash (multiple-vehicle collision) in low-visibility condition on a road. In the traffic situation, drivers brake according to taillights of the forward vehicle. The first crash may induce more collisions. We investigate whether or not the first collision induces the chain-reaction crash, numerically and analytically. The dynamic transitions occur from no collisions through a single collision, double collisions and triple collisions, to multiple collisions with decreasing the headway. Also, we find that the dynamic transition occurs from the finite chain reaction to the infinite chain reaction when the headway is less than the critical value. We derive, analytically, the transition points and the region maps for the chain-reaction crash in traffic flow controlled by taillights.
-
Vázquez, C; Reyes, R; Alcaraz, F; Balsa, J A; Botella-Carretero, J I
2006-01-01
Lipodystrophy is a frequent disorder among patients with human immunodeficiency virus (HIV) infection, characterized by a loss of adipose tissue from the extremities, gluteal region and face, with excess fat in the neck and abdominal region. Metabolic abnormalities such as hyperlipidaemia and diabetes mellitus frequently coexist, posing these patients to an increased cardiovascular risk. Drug therapy may improve some of these metabolic disturbances, but to date there are no treatments for lipodystrophy with proven benefit. A 42-year-old man with HIV lipodystrophy was started on a standard low caloric diet with <30% of total fat and <10% of saturated fat, together with rosiglitazone 8 mg daily. After five months of treatment, given that lipodystrophic features and dyslipidaemia were still present in our patient, we tried to further improve therapeutic results by eucaloric substitution of medium chain triglycerides for dietary long chain fatty acids. Three months later, a dramatic change in body composition was shown with an increase in lean mass and a decrease in fat mass, together with an improvement in lipid profile. Eucaloric substitution of medium chain triglycerides for dietary long chain fatty acids may produce therapeutic benefits in HIV lipodystrophy.
-
NASA Astrophysics Data System (ADS)
Maruyama, Kenji; Hiroi (Sato), Satoshi; Endo, Hirohisa; Hoshino, Hideoki; Odagaki, Takashi; Hensel, Friedrich
2017-08-01
The reverse Monte Carlo (RMC) and Voronoi-Delaunay (VD) void analyses were applied to study the modification of chain geometries near the semiconductor (SC) to metal (M) transition in expanded liquid Se along the isochore of d = 3.4 g/cm3. Fluctuations of dihedral angles with increasing temperature and pressure cause modification of the helical (H) chain to the planar zigzag (Z) chain conformations. The distribution of voids size (rV ) supported by chain segments and distances to the 4th 6th neighbor atoms on the chain segments provide information on the stacking of planar zigzag chains compensated by empty space (L-voids, rV 3.6 Å) which leads to the formation of metallic domains. Near SC-M transition region the number fraction NZ/NH for Z and H chain segments increases.
-
2008-01-01
Bioinformatics, 19, ii246–ii255. 52. Lawrence, M. C. & Colman, P. M. (1993). Shape complementarity at protein / protein interfaces . J. Mol. Biol. 234, 946...envelope spike, which is the sole protein expressed on the surface of the Ebola virus and is involved in receptor binding, tropism, and viral entry.6–9 It...variable light chain/heavy chain (VL/VH) interface of 13F6-1-2, ∼1025 Å2 surface area is buried on VL Fig. 1. Nucleotide and translated amino acid
-
2007-10-01
twists. Bioinformatics, 19, ii246–ii255. 52. Lawrence, M. C. & Colman, P. M. (1993). Shape complementarity at protein / protein interfaces . J. Mol. Biol...envelope spike, which is the sole protein expressed on the surface of the Ebola virus and is involved in receptor binding, tropism, and viral entry.6–9 It...26 At the variable light chain/heavy chain (VL/VH) interface of 13F6-1-2, ∼1025 Å2 surface area is buried on VL Fig. 1. Nucleotide and translated amino
-
DOT National Transportation Integrated Search
2009-05-01
This 2009 study, funded by the Southwest Region University Transportation Center, investigates : competing ethanol supply chains terminating in the State of Texas. Midwest corn ethanol and : Brazilian sugarcane ethanol constitute two sources of the b...
-
NASA Astrophysics Data System (ADS)
Ho, M. W.; Lall, U.; Cook, E. R.
2015-12-01
Advances in paleoclimatology in the past few decades have provided opportunities to expand the temporal perspective of the hydrological and climatological variability across the world. The North American region is particularly fortunate in this respect where a relatively dense network of high resolution paleoclimate proxy records have been assembled. One such network is the annually-resolved Living Blended Drought Atlas (LBDA): a paleoclimate reconstruction of the Palmer Drought Severity Index (PDSI) that covers North America on a 0.5° × 0.5° grid based on tree-ring chronologies. However, the use of the LBDA to assess North American streamflow variability requires a model by which streamflow may be reconstructed. Paleoclimate reconstructions have typically used models that first seek to quantify the relationship between the paleoclimate variable and the environmental variable of interest before extrapolating the relationship back in time. In contrast, the pre-instrumental streamflow is here considered as "missing" data. A method of imputing the "missing" streamflow data, prior to the instrumental record, is applied through multiple imputation using chained equations for streamflow in the Missouri River Basin. In this method, the distribution of the instrumental streamflow and LBDA is used to estimate sets of plausible values for the "missing" streamflow data resulting in a ~600 year-long streamflow reconstruction. Past research into external climate forcings, oceanic-atmospheric variability and its teleconnections, and assessments of rare multi-centennial instrumental records demonstrate that large temporal oscillations in hydrological conditions are unlikely to be captured in most instrumental records. The reconstruction of multi-centennial records of streamflow will enable comprehensive assessments of current and future water resource infrastructure and operations under the existing scope of natural climate variability.
-
NASA Astrophysics Data System (ADS)
Arnold, J.; Gutmann, E. D.; Clark, M. P.; Nijssen, B.; Vano, J. A.; Addor, N.; Wood, A.; Newman, A. J.; Mizukami, N.; Brekke, L. D.; Rasmussen, R.; Mendoza, P. A.
2016-12-01
Climate change narratives for water-resource applications must represent the change signals contextualized by hydroclimatic process variability and uncertainty at multiple scales. Building narratives of plausible change includes assessing uncertainties across GCM structure, internal climate variability, climate downscaling methods, and hydrologic models. Work with this linked modeling chain has dealt mostly with GCM sampling directed separately to either model fidelity (does the model correctly reproduce the physical processes in the world?) or sensitivity (of different model responses to CO2 forcings) or diversity (of model type, structure, and complexity). This leaves unaddressed any interactions among those measures and with other components in the modeling chain used to identify water-resource vulnerabilities to specific climate threats. However, time-sensitive, real-world vulnerability studies typically cannot accommodate a full uncertainty ensemble across the whole modeling chain, so a gap has opened between current scientific knowledge and most routine applications for climate-changed hydrology. To close that gap, the US Army Corps of Engineers, the Bureau of Reclamation, and the National Center for Atmospheric Research are working on techniques to subsample uncertainties objectively across modeling chain components and to integrate results into quantitative hydrologic storylines of climate-changed futures. Importantly, these quantitative storylines are not drawn from a small sample of models or components. Rather, they stem from the more comprehensive characterization of the full uncertainty space for each component. Equally important from the perspective of water-resource practitioners, these quantitative hydrologic storylines are anchored in actual design and operations decisions potentially affected by climate change. This talk will describe part of our work characterizing variability and uncertainty across modeling chain components and their interactions using newly developed observational data, models and model outputs, and post-processing tools for making the resulting quantitative storylines most useful in practical hydrology applications.
-
Henry, Kevin A
2018-01-01
Immunogenetic analyses of expressed antibody repertoires are becoming increasingly common experimental investigations and are critical to furthering our understanding of autoimmunity, infectious disease, and cancer. Next-generation DNA sequencing (NGS) technologies have now made it possible to interrogate antibody repertoires to unprecedented depths, typically by sequencing of cDNAs encoding immunoglobulin variable domains. In this chapter, we describe simple, fast, and reliable methods for producing and sequencing multiplex PCR amplicons derived from the variable regions (V H , V H H or V L ) of rearranged immunoglobulin heavy and light chain genes using the Illumina MiSeq platform. We include complete protocols and primer sets for amplicon sequencing of V H /V H H/V L repertoires directly from human, mouse, and llama lymphocytes as well as from phage-displayed V H /V H H/V L libraries; these can be easily be adapted to other types of amplicons with little modification. The resulting amplicons are diverse and representative, even using as few as 10 3 input B cells, and their generation is relatively inexpensive, requiring no special equipment and only a limited set of primers. In the absence of heavy-light chain pairing, single-domain antibodies are uniquely amenable to NGS analyses. We present a number of applications of NGS technology useful in discovery of single-domain antibodies from phage display libraries, including: (i) assessment of library functionality; (ii) confirmation of desired library randomization; (iii) estimation of library diversity; and (iv) monitoring the progress of panning experiments. While the case studies presented here are of phage-displayed single-domain antibody libraries, the principles extend to other types of in vitro display libraries.
-
Main, Sarah; Handy, Rachel; Wilton, Jane; Smith, Stephen; Williams, Liz; Fou, Leila Du; Andrews, John; Conroy, Louise A; May, Richard; Anderson, Ian; Vaughan, Tristan J
2006-12-01
The CC chemokine, eotaxin1 (CCL11) is an important regulator of eosinophil function. A marked accumulation of eosinophils in tissues has been correlated with the up-regulation of eotaxin1 expression in several diseases. The potential therapeutic value of neutralizing the effects of eotaxin1 in inflammatory conditions (including asthma) is under investigation. A human single-chain fragment variable antibody that neutralizes human eotaxin1 (CAT-212) was produced using antibody phage display and converted to whole antibody IgG4 format (CAT-213). A novel approach to lead optimization in which the length of the variable heavy chain complementarity-determining region 3 was reduced by one amino acid resulted in an increase in potency of >1000-fold compared with the parent anti-eotaxin1 antibody. The optimized antibody binds eotaxin1 with high affinity (80.4 pM) and specificity. CAT-213 and CAT-212 do not bind or neutralize a range of other human proteins including human monocyte chemoattractant protein-1, a structurally similar chemokine. CAT-213 neutralizes the ability of eotaxin1 to cause an increase in intracellular calcium signaling (with an IC(50) value of 2.86 nM), migration of CCR3-expressing L1.2 cells (with an IC(50) value of 0.48 nM), and inhibition of the eotaxin1-evoked shape change of human eosinophils in vitro (with an IC(50) of 0.71 nM). Local administration of CAT-213 to mice (1-100 microg kg(-1)) attenuates dermal eosinophilia induced by human eotaxin1, achieving >90% inhibition of eosinophil influx. CAT-213 may therefore be of therapeutic value in inhibiting diseases in which eotaxin1 and eosinophils play a major role, for example, severe asthma.
-
Human immunoglobulin allotypes
Lefranc, Marie-Paule
2009-01-01
More than twenty recombinant monoclonal antibodies are approved as therapeutics. Almost all of these are based on the whole IgG isotype format, but vary in the origin of the variable regions between mouse (chimeric), humanized mouse and fully human sequences; all of those with whole IgG format employ human constant region sequences. Currently, the opposing merits of the four IgG subclasses are considered with respect to the in vivo biological activities considered to be appropriate to the disease indication being treated. Human heavy chain genes also exhibit extensive structural polymorphism(s) and, being closely linked, are inherited as a haplotype. Polymorphisms (allotypes) within the IgG isotype were originally discovered and described using serological reagents derived from humans; demonstrating that allotypic variants can be immunogenic and provoke antibody responses as a result of allo-immunization. The serologically defined allotypes differ widely within and between population groups; therefore, a mAb of a given allotype will, inevitably, be delivered to a cohort of patients homozygous for the alternative allotype. This publication reviews the serologically defined human IgG allotypes and considers the potential for allotype differences to contribute to or potentiate immunogenicity. PMID:20073133