Science.gov

Sample records for acid substitution rates

  1. Estimating the Variability of Substitution Rates

    PubMed Central

    Bulmer, M.

    1989-01-01

    Suppose that amino acid or nucleotide data are available for a homologous gene in several species which diverged from a common ancestor at about the same time and that substitution rates between all pairs of species are calculated, correcting as necessary for multiple substitutions and for back and parallel substitutions. The variances and covariances of these corrected substitution rates are evaluated, and are used to construct a new test for uniformity (constancy of the molecular clock) and to find the best estimates of substitution rates in individual lineages with their standard errors. A substantial bias may arise if the effect of correcting the pairwise substitution rates is ignored. PMID:2599371

  2. Substitution Rates under Stabilizing Selection

    PubMed Central

    Hastings, Alan

    1987-01-01

    Allelic substitutions under stabilizing phenotypic selection on quantitative traits are studied in Monte Carlo simulations of 8 and 16 loci. The results are compared and contrasted to analytical models based on work of M. Kimura for two and "infinite" loci. Selection strengths of S = 4Nes approximately four (which correspond to reasonable strengths of selection for quantitative characters) can retard substitution rates tenfold relative to rates under neutrality. An important finding is a strong dependence of per locus substitution rates on the number of loci. PMID:3609727

  3. 40 CFR 721.10126 - Alkyl amino substituted triazine amino substituted benezenesulfonic acid reaction product with...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo substituted phenyl azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato...

  4. 40 CFR 721.10126 - Alkyl amino substituted triazine amino substituted benezenesulfonic acid reaction product with...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo substituted phenyl azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato...

  5. 40 CFR 721.10126 - Alkyl amino substituted triazine amino substituted benezenesulfonic acid reaction product with...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo substituted phenyl azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato...

  6. 40 CFR 721.10126 - Alkyl amino substituted triazine amino substituted benezenesulfonic acid reaction product with...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo substituted phenyl azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato...

  7. 40 CFR 721.10126 - Alkyl amino substituted triazine amino substituted benezenesulfonic acid reaction product with...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo substituted phenyl azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato azo... substituted triazine amino substituted benezenesulfonic acid reaction product with naphthalenesulfonato...

  8. Switch in codon bias and increased rates of amino acid substitution in the Drosophila saltans species group.

    PubMed Central

    Rodríguez-Trelles, F; Tarrío, R; Ayala, F J

    1999-01-01

    We investigated the nucleotide composition of five genes, Xdh, Adh, Sod, Per, and 28SrRNA, in nine species of Drosophila (subgenus Sophophora) and one of Scaptodrosophila. The six species of the Drosophila saltans group markedly differ from the others in GC content and codon use bias. The GC content in the third codon position, and to a lesser extent in the first position and the introns, is higher in the D. melanogaster and D. obscura groups than in the D. saltans group (in Scaptodrosophila it is intermediate but closer to the melanogaster and obscura species). Differences are greater for Xdh than for Adh, Sod, Per, and 28SrRNA, which are functionally more constrained. We infer that rapid evolution of GC content in the saltans lineage is largely due to a shift in mutation pressure, which may have been associated with diminished natural selection due to smaller effective population numbers rather than reduced recombination rates. The rate of GC content evolution impacts the rate of protein evolution and may distort phylogenetic inferences. Previous observations suggesting that GC content evolution is very limited in Drosophila may have been distorted due to the restricted number of genes and species (mostly D. melanogaster) investigated. PMID:10471717

  9. Capillary Electrophoresis of Substituted Benzoic Acids

    ERIC Educational Resources Information Center

    Mills, Nancy S.; Spence, John D.; Bushey, Michelle M.

    2005-01-01

    A series of substituted benzoic acids (SBAs) are prepared by students. The pKa shift, a result of the electron-withdrawing or electron-donating characteristics of the subsistent is examined in reference to the electrophoretic migration behavior of benzoic acid.

  10. 40 CFR 721.1680 - Substituted benzoic acid (generic).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Substituted benzoic acid (generic... Substances § 721.1680 Substituted benzoic acid (generic). (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as substituted benzoic acid (PMN...

  11. 40 CFR 721.1680 - Substituted benzoic acid (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Substituted benzoic acid (generic... Substances § 721.1680 Substituted benzoic acid (generic). (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as substituted benzoic acid (PMN...

  12. 40 CFR 721.1680 - Substituted benzoic acid (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Substituted benzoic acid (generic... Substances § 721.1680 Substituted benzoic acid (generic). (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as substituted benzoic acid (PMN...

  13. 40 CFR 721.1680 - Substituted benzoic acid (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Substituted benzoic acid (generic... Substances § 721.1680 Substituted benzoic acid (generic). (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as substituted benzoic acid (PMN...

  14. 40 CFR 721.1680 - Substituted benzoic acid (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Substituted benzoic acid (generic... Substances § 721.1680 Substituted benzoic acid (generic). (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified generically as substituted benzoic acid (PMN...

  15. 40 CFR 721.10679 - Carboxylic acid, substituted alkylstannylene ester, reaction products with inorganic acid tetra...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... alkylstannylene ester, reaction products with inorganic acid tetra alkyl ester (generic). 721.10679 Section 721... Carboxylic acid, substituted alkylstannylene ester, reaction products with inorganic acid tetra alkyl ester... identified generically as carboxylic acid, substituted alkylstannylene ester, reaction products...

  16. Transitions to asexuality result in excess amino acid substitutions.

    PubMed

    Paland, Susanne; Lynch, Michael

    2006-02-17

    Theory predicts that linkage between genetic loci reduces the efficiency of purifying selection. Because of the permanent linkage of all heritable genetic material, asexual lineages may be exceptionally prone to deleterious-mutation accumulation in both nuclear and organelle genes. Here, we show that the ratio of the rate of amino acid to silent substitution (Ka/Ks) in mitochondrial protein-coding genes is higher in obligately asexual lineages than in sexual lineages of the microcrustacean Daphnia pulex. Using a phylogeny-based approach to quantify the frequency of mutational-effect classes, we estimate that mitochondrial protein-coding genes in asexual lineages accumulate deleterious amino acid substitutions at four times the rate in sexual lineages. These results support the hypothesis that sexual reproduction plays a prominent role in reducing the mutational burden in populations.

  17. Overdispersion of the molecular clock: temporal variation of gene-specific substitution rates in Drosophila.

    PubMed

    Bedford, Trevor; Hartl, Daniel L

    2008-08-01

    Simple models of molecular evolution assume that sequences evolve by a Poisson process in which nucleotide or amino acid substitutions occur as rare independent events. In these models, the expected ratio of the variance to the mean of substitution counts equals 1, and substitution processes with a ratio greater than 1 are called overdispersed. Comparing the genomes of 10 closely related species of Drosophila, we extend earlier evidence for overdispersion in amino acid replacements as well as in four-fold synonymous substitutions. The observed deviation from the Poisson expectation can be described as a linear function of the rate at which substitutions occur on a phylogeny, which implies that deviations from the Poisson expectation arise from gene-specific temporal variation in substitution rates. Amino acid sequences show greater temporal variation in substitution rates than do four-fold synonymous sequences. Our findings provide a general phenomenological framework for understanding overdispersion in the molecular clock. Also, the presence of substantial variation in gene-specific substitution rates has broad implications for work in phylogeny reconstruction and evolutionary rate estimation.

  18. 40 CFR 721.1643 - Benzenesulfonic acid, amino substituted phenylazo-.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Benzenesulfonic acid, amino... Specific Chemical Substances § 721.1643 Benzenesulfonic acid, amino substituted phenylazo-. (a) Chemical... as a benzenesulfonic acid, amino substituted phenylazo- (PMN P-95-86) is subject to reporting...

  19. 40 CFR 721.1643 - Benzenesulfonic acid, amino substituted phenylazo-.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Benzenesulfonic acid, amino... Specific Chemical Substances § 721.1643 Benzenesulfonic acid, amino substituted phenylazo-. (a) Chemical... as a benzenesulfonic acid, amino substituted phenylazo- (PMN P-95-86) is subject to reporting...

  20. 40 CFR 721.1643 - Benzenesulfonic acid, amino substituted phenylazo-.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Benzenesulfonic acid, amino... Specific Chemical Substances § 721.1643 Benzenesulfonic acid, amino substituted phenylazo-. (a) Chemical... as a benzenesulfonic acid, amino substituted phenylazo- (PMN P-95-86) is subject to reporting...

  1. 40 CFR 721.1643 - Benzenesulfonic acid, amino substituted phenylazo-.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Benzenesulfonic acid, amino... Specific Chemical Substances § 721.1643 Benzenesulfonic acid, amino substituted phenylazo-. (a) Chemical... as a benzenesulfonic acid, amino substituted phenylazo- (PMN P-95-86) is subject to reporting...

  2. 40 CFR 721.1643 - Benzenesulfonic acid, amino substituted phenylazo-.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Benzenesulfonic acid, amino... Specific Chemical Substances § 721.1643 Benzenesulfonic acid, amino substituted phenylazo-. (a) Chemical... as a benzenesulfonic acid, amino substituted phenylazo- (PMN P-95-86) is subject to reporting...

  3. FRAGS: estimation of coding sequence substitution rates from fragmentary data

    PubMed Central

    Swart, Estienne C; Hide, Winston A; Seoighe, Cathal

    2004-01-01

    Background Rates of substitution in protein-coding sequences can provide important insights into evolutionary processes that are of biomedical and theoretical interest. Increased availability of coding sequence data has enabled researchers to estimate more accurately the coding sequence divergence of pairs of organisms. However the use of different data sources, alignment protocols and methods to estimate substitution rates leads to widely varying estimates of key parameters that define the coding sequence divergence of orthologous genes. Although complete genome sequence data are not available for all organisms, fragmentary sequence data can provide accurate estimates of substitution rates provided that an appropriate and consistent methodology is used and that differences in the estimates obtainable from different data sources are taken into account. Results We have developed FRAGS, an application framework that uses existing, freely available software components to construct in-frame alignments and estimate coding substitution rates from fragmentary sequence data. Coding sequence substitution estimates for human and chimpanzee sequences, generated by FRAGS, reveal that methodological differences can give rise to significantly different estimates of important substitution parameters. The estimated substitution rates were also used to infer upper-bounds on the amount of sequencing error in the datasets that we have analysed. Conclusion We have developed a system that performs robust estimation of substitution rates for orthologous sequences from a pair of organisms. Our system can be used when fragmentary genomic or transcript data is available from one of the organisms and the other is a completely sequenced genome within the Ensembl database. As well as estimating substitution statistics our system enables the user to manage and query alignment and substitution data. PMID:15005802

  4. 40 CFR 721.1700 - Halonitrobenzoic acid, substituted (generic name).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Halonitrobenzoic acid, substituted (generic name). 721.1700 Section 721.1700 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1700 Halonitrobenzoic acid, substituted (generic name). (a)...

  5. 40 CFR 721.1700 - Halonitrobenzoic acid, substituted (generic name).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Halonitrobenzoic acid, substituted (generic name). 721.1700 Section 721.1700 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.1700 Halonitrobenzoic acid, substituted (generic name). (a)...

  6. 40 CFR 721.5278 - Substituted naphthalenesulfonic acid, alkali salt.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ..., alkali salt. 721.5278 Section 721.5278 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.5278 Substituted naphthalenesulfonic acid, alkali salt. (a) Chemical... as a substituted naphthalenesulfonic acid, alkali salt (PMN P-95-85) is subject to reporting...

  7. 40 CFR 721.5278 - Substituted naphthalenesulfonic acid, alkali salt.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ..., alkali salt. 721.5278 Section 721.5278 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.5278 Substituted naphthalenesulfonic acid, alkali salt. (a) Chemical... as a substituted naphthalenesulfonic acid, alkali salt (PMN P-95-85) is subject to reporting...

  8. 40 CFR 721.1648 - Substituted benzenesulfonic acid salt (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Substituted benzenesulfonic acid salt (generic). 721.1648 Section 721.1648 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.1648 Substituted benzenesulfonic acid salt (generic). (a)...

  9. 40 CFR 721.5278 - Substituted naphthalenesulfonic acid, alkali salt.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ..., alkali salt. 721.5278 Section 721.5278 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.5278 Substituted naphthalenesulfonic acid, alkali salt. (a) Chemical... as a substituted naphthalenesulfonic acid, alkali salt (PMN P-95-85) is subject to reporting...

  10. 40 CFR 721.5278 - Substituted naphthalenesulfonic acid, alkali salt.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ..., alkali salt. 721.5278 Section 721.5278 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.5278 Substituted naphthalenesulfonic acid, alkali salt. (a) Chemical... as a substituted naphthalenesulfonic acid, alkali salt (PMN P-95-85) is subject to reporting...

  11. 40 CFR 721.5278 - Substituted naphthalenesulfonic acid, alkali salt.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ..., alkali salt. 721.5278 Section 721.5278 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY... Specific Chemical Substances § 721.5278 Substituted naphthalenesulfonic acid, alkali salt. (a) Chemical... as a substituted naphthalenesulfonic acid, alkali salt (PMN P-95-85) is subject to reporting...

  12. 40 CFR 721.1648 - Substituted benzenesulfonic acid salt (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Substituted benzenesulfonic acid salt (generic). 721.1648 Section 721.1648 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.1648 Substituted benzenesulfonic acid salt (generic). (a)...

  13. 40 CFR 721.1648 - Substituted benzenesulfonic acid salt (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Substituted benzenesulfonic acid salt (generic). 721.1648 Section 721.1648 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.1648 Substituted benzenesulfonic acid salt (generic). (a)...

  14. 40 CFR 721.1648 - Substituted benzenesulfonic acid salt (generic).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Substituted benzenesulfonic acid salt (generic). 721.1648 Section 721.1648 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.1648 Substituted benzenesulfonic acid salt (generic). (a)...

  15. 40 CFR 721.1648 - Substituted benzenesulfonic acid salt (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Substituted benzenesulfonic acid salt (generic). 721.1648 Section 721.1648 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Specific Chemical Substances § 721.1648 Substituted benzenesulfonic acid salt (generic). (a)...

  16. Redesigning Channel-Forming Peptides: Amino Acid Substitutions that Enhance Rates of Supramolecular Self-Assembly and Raise Ion Transport Activity

    PubMed Central

    Shank, Lalida P.; Broughman, James R.; Takeguchi, Wade; Cook, Gabriel; Robbins, Ashley S.; Hahn, Lindsey; Radke, Gary; Iwamoto, Takeo; Schultz, Bruce D.; Tomich, John M.

    2006-01-01

    Three series of 22-residue peptides derived from the transmembrane M2 segment of the glycine receptor α1-subunit (M2GlyR) have been designed, synthesized, and tested to determine the plasticity of a channel-forming sequence and to define whether channel pores with enhanced conductive properties could be created. Sixteen sequences were examined for aqueous solubility, solution-association tendency, secondary structure, and half-maximal concentration for supramolecular assembly, channel activity, and ion transport properties across epithelial monolayers. All peptides interact strongly with membranes: associating with, inserting across, and assembling to form homooligomeric bundles when in micromolar concentrations. Single and double amino acid replacements involving arginine and/or aromatic amino acids within the final five C-terminal residues of the peptide cause dramatic effects on the concentration dependence, yielding a range of K1/2 values from 36 ± 5 to 390 ± 220 μM for transport activity. New water/lipid interfacial boundaries were established for the transmembrane segment using charged or aromatic amino acids, thus limiting the peptides' ability to move perpendicularly to the plane of the bilayer. Formation of discrete water/lipid interfacial boundaries appears to be necessary for efficient supramolecular assembly and high anion transport activity. A peptide sequence is identified that may show efficacy in channel replacement therapy for channelopathies such as cystic fibrosis. PMID:16387776

  17. Gene Tree Discordance Causes Apparent Substitution Rate Variation.

    PubMed

    Mendes, Fábio K; Hahn, Matthew W

    2016-07-01

    Substitution rates are known to be variable among genes, chromosomes, species, and lineages due to multifarious biological processes. Here, we consider another source of substitution rate variation due to a technical bias associated with gene tree discordance. Discordance has been found to be rampant in genome-wide data sets, often due to incomplete lineage sorting (ILS). This apparent substitution rate variation is caused when substitutions that occur on discordant gene trees are analyzed in the context of a single, fixed species tree. Such substitutions have to be resolved by proposing multiple substitutions on the species tree, and we therefore refer to this phenomenon as Substitutions Produced by ILS (SPILS). We use simulations to demonstrate that SPILS has a larger effect with increasing levels of ILS, and on trees with larger numbers of taxa. Specific branches of the species trees are consistently, but erroneously, inferred to be longer or shorter, and we show that these branches can be predicted based on discordant tree topologies. Moreover, we observe that fixing a species tree topology when performing tests of positive selection increases the false positive rate, particularly for genes whose discordant topologies are most affected by SPILS. Finally, we use data from multiple Drosophila species to show that SPILS can be detected in nature. Although the effects of SPILS are modest per gene, it has the potential to affect substitution rate variation whenever high levels of ILS are present, particularly in rapid radiations. The problems outlined here have implications for character mapping of any type of trait, and for any biological process that causes discordance. We discuss possible solutions to these problems, and areas in which they are likely to have caused faulty inferences of convergence and accelerated evolution.

  18. Nonsynonymous substitution rate heterogeneity in the peptide-binding region among different HLA-DRB1 lineages in humans.

    PubMed

    Yasukochi, Yoshiki; Satta, Yoko

    2014-05-02

    An extraordinary diversity of amino acid sequences in the peptide-binding region (PBR) of human leukocyte antigen [HLA; human major histocompatibility complex (MHC)] molecules has been maintained by balancing selection. The process of accumulation of amino acid diversity in the PBR for six HLA genes (HLA-A, B, C, DRB1, DQB1, and DPB1) shows that the number of amino acid substitutions in the PBR among alleles does not linearly correlate with the divergence time of alleles at the six HLA loci. At these loci, some pairs of alleles show significantly less nonsynonymous substitutions at the PBR than expected from the divergence time. The same phenomenon was observed not only in the HLA but also in the rat MHC. To identify the cause for this, DRB1 sequences, a representative case of a typical nonlinear pattern of substitutions, were examined. When the amino acid substitutions in the PBR were placed with maximum parsimony on a maximum likelihood tree based on the non-PBR substitutions, heterogeneous rates of nonsynonymous substitutions in the PBR were observed on several branches. A computer simulation supported the hypothesis that allelic pairs with low PBR substitution rates were responsible for the stagnation of accumulation of PBR nonsynonymous substitutions. From these observations, we conclude that the nonsynonymous substitution rate at the PBR sites is not constant among the allelic lineages. The deceleration of the rate may be caused by the coexistence of certain pathogens for a substantially long time during HLA evolution.

  19. Substitution rate and natural selection in parvovirus B19

    PubMed Central

    Stamenković, Gorana G.; Ćirković, Valentina S.; Šiljić, Marina M.; Blagojević, Jelena V.; Knežević, Aleksandra M.; Joksić, Ivana D.; Stanojević, Maja P.

    2016-01-01

    The aim of this study was to estimate substitution rate and imprints of natural selection on parvovirus B19 genotype 1. Studied datasets included 137 near complete coding B19 genomes (positions 665 to 4851) for phylogenetic and substitution rate analysis and 146 and 214 partial genomes for selection analyses in open reading frames ORF1 and ORF2, respectively, collected 1973–2012 and including 9 newly sequenced isolates from Serbia. Phylogenetic clustering assigned majority of studied isolates to G1A. Nucleotide substitution rate for total coding DNA was 1.03 (0.6–1.27) x 10−4 substitutions/site/year, with higher values for analyzed genome partitions. In spite of the highest evolutionary rate, VP2 codons were found to be under purifying selection with rare episodic positive selection, whereas codons under diversifying selection were found in the unique part of VP1, known to contain B19 immune epitopes important in persistent infection. Analyses of overlapping gene regions identified nucleotide positions under opposite selective pressure in different ORFs, suggesting complex evolutionary mechanisms of nucleotide changes in B19 viral genomes. PMID:27775080

  20. 40 CFR 721.10448 - Acetic acid, hydroxy- methoxy-, methyl ester, reaction products with substituted alkylamine...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... ester, reaction products with substituted alkylamine (generic). 721.10448 Section 721.10448 Protection... Acetic acid, hydroxy- methoxy-, methyl ester, reaction products with substituted alkylamine (generic). (a... generically as acetic acid, hydroxymethoxy-, methyl ester, reaction products with substituted alkylamine...

  1. 40 CFR 721.10448 - Acetic acid, hydroxy- methoxy-, methylester, reaction products with substituted alkylamine...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...-, methylester, reaction products with substituted alkylamine (generic). 721.10448 Section 721.10448 Protection... Acetic acid, hydroxy- methoxy-, methylester, reaction products with substituted alkylamine (generic). (a... generically as acetic acid, hydroxymethoxy-, methyl ester, reaction products with substituted alkylamine...

  2. FLU, an amino acid substitution model for influenza proteins

    PubMed Central

    2010-01-01

    Background The amino acid substitution model is the core component of many protein analysis systems such as sequence similarity search, sequence alignment, and phylogenetic inference. Although several general amino acid substitution models have been estimated from large and diverse protein databases, they remain inappropriate for analyzing specific species, e.g., viruses. Emerging epidemics of influenza viruses raise the need for comprehensive studies of these dangerous viruses. We propose an influenza-specific amino acid substitution model to enhance the understanding of the evolution of influenza viruses. Results A maximum likelihood approach was applied to estimate an amino acid substitution model (FLU) from ~113, 000 influenza protein sequences, consisting of ~20 million residues. FLU outperforms 14 widely used models in constructing maximum likelihood phylogenetic trees for the majority of influenza protein alignments. On average, FLU gains ~42 log likelihood points with an alignment of 300 sites. Moreover, topologies of trees constructed using FLU and other models are frequently different. FLU does indeed have an impact on likelihood improvement as well as tree topologies. It was implemented in PhyML and can be downloaded from ftp://ftp.sanger.ac.uk/pub/1000genomes/lsq/FLU or included in PhyML 3.0 server at http://www.atgc-montpellier.fr/phyml/. Conclusions FLU should be useful for any influenza protein analysis system which requires an accurate description of amino acid substitutions. PMID:20384985

  3. Gas phase acidity of substituted benzenes

    NASA Astrophysics Data System (ADS)

    Bouchoux, Guy

    2011-04-01

    Deprotonation thermochemistry of benzene derivatives C 6H 5X (X = H, F, Cl, OH, NH 2, CN, CHO, NO 2, CH 3, C 2H 5, CHCH 2, CCH) has been examined at the G3B3 level of theory. For X = F, Cl, CN, CHO and NO 2, the most favorable deprotonation site is the ortho position of the phenyl ring. This regio-specificity is directly related to the field/inductive effect of the substituent. G3B3 gas phase acidities, Δ acidH° and Δ acidG°, compare within less than 4 kJ mol -1 with experimental data. A noticeable exception is nitrobenzene for which tabulated acidity appear to be underestimated by ca. 120 kJ mol -1.

  4. Systematic Exploration of an Efficient Amino Acid Substitution Matrix: MIQS.

    PubMed

    Tomii, Kentaro; Yamada, Kazunori

    2016-01-01

    Amino acid sequence comparisons to find similarities between proteins are fundamental sequence information analyses for inferring protein structure and function. In this study, we improve amino acid substitution matrices to identify distantly related proteins. We systematically sampled and benchmarked substitution matrices generated from the principal component analysis (PCA) subspace based on a set of typical existing matrices. Based on the benchmark results, we identified a region of highly sensitive matrices in the PCA subspace using kernel density estimation (KDE). Using the PCA subspace, we were able to deduce a novel sensitive matrix, called MIQS, which shows better detection performance for detecting distantly related proteins than those of existing matrices. This approach to derive an efficient amino acid substitution matrix might influence many fields of protein sequence analysis. MIQS is available at http://csas.cbrc.jp/Ssearch/ . PMID:27115635

  5. Genotoxic effect of substituted phenoxyacetic acids.

    PubMed

    Venkov, P; Topashka-Ancheva, M; Georgieva, M; Alexieva, V; Karanov, E

    2000-11-01

    The potential toxic and mutagenic action of 2,4-dichlorophenoxyacetic acid has been studied in different test systems, and the obtained results range from increased chromosomal damage to no effect at all. We reexamined the effect of this herbicide by simultaneous using three tests based on yeast, transformed hematopoietic, and mouse bone marrow cells. The results obtained demonstrated that 2,4-dichlorophenoxyacetic acid has cytotoxic and mutagenic effects. The positive response of yeast and transformed hematopoietic cells was verified in kinetics and dose-response experiments. The analysis of metaphase chromosomes indicated a statistically proved induction of breaks, deletions, and exchanges after the intraperitoneal administration of 2,4-dichlorophenoxyacetic acid in mice. The study of phenoxyacetic acid and its differently chlorinated derivatives showed that cytotoxicity and mutagenicity are induced by chlorine atoms at position 2 and/or 4 in the benzene ring. The mutagenic effect was abolished by introduction of a third chlorine atom at position 5. Thus 2,4,5-trichlorophenoxyacetic acid was found to have very weak, if any mutagenic effect; however, the herbicide preserved its toxic effect.

  6. In Vitro Investigation of Self-Assembled Nanoparticles Based on Hyaluronic Acid-Deoxycholic Acid Conjugates for Controlled Release Doxorubicin: Effect of Degree of Substitution of Deoxycholic Acid

    PubMed Central

    Wei, Wen-Hao; Dong, Xue-Meng; Liu, Chen-Guang

    2015-01-01

    Self-assembled nanoparticles based on a hyaluronic acid-deoxycholic acid (HD) chemical conjugate with different degree of substitution (DS) of deoxycholic acid (DOCA) were prepared. The degree of substitution (DS) was determined by titration method. The nanoparticles were loaded with doxorubicin (DOX) as the model drug. The human cervical cancer (HeLa) cell line was utilized for in vitro studies and cell cytotoxicity of DOX incorporated in the HD nanoparticles was accessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. In addition, cellular uptake of fluorescently labeled nanoparticles was also investigated. An increase in the degree of deoxycholic acid substitution reduced the size of the nanoparticles and also enhanced their drug encapsulation efficiency (EE), which increased with the increase of DS. A higher degree of deoxycholic acid substitution also lead to a lower release rate and an initial burst release of doxorubicin from the nanoparticles. In summary, the degree of substitution allows the modulation of the particle size, drug encapsulation efficiency, drug release rate, and cell uptake efficiency of the nanoparticles. The herein developed hyaluronic acid-deoxycholic acid conjugates are a good candidate for drug delivery and could potentiate therapeutic formulations for doxorubicin–mediated cancer therapy. PMID:25837468

  7. Estimating divergence dates and substitution rates in the Drosophila phylogeny.

    PubMed

    Obbard, Darren J; Maclennan, John; Kim, Kang-Wook; Rambaut, Andrew; O'Grady, Patrick M; Jiggins, Francis M

    2012-11-01

    An absolute timescale for evolution is essential if we are to associate evolutionary phenomena, such as adaptation or speciation, with potential causes, such as geological activity or climatic change. Timescales in most phylogenetic studies use geologically dated fossils or phylogeographic events as calibration points, but more recently, it has also become possible to use experimentally derived estimates of the mutation rate as a proxy for substitution rates. The large radiation of drosophilid taxa endemic to the Hawaiian islands has provided multiple calibration points for the Drosophila phylogeny, thanks to the "conveyor belt" process by which this archipelago forms and is colonized by species. However, published date estimates for key nodes in the Drosophila phylogeny vary widely, and many are based on simplistic models of colonization and coalescence or on estimates of island age that are not current. In this study, we use new sequence data from seven species of Hawaiian Drosophila to examine a range of explicit coalescent models and estimate substitution rates. We use these rates, along with a published experimentally determined mutation rate, to date key events in drosophilid evolution. Surprisingly, our estimate for the date for the most recent common ancestor of the genus Drosophila based on mutation rate (25-40 Ma) is closer to being compatible with independent fossil-derived dates (20-50 Ma) than are most of the Hawaiian-calibration models and also has smaller uncertainty. We find that Hawaiian-calibrated dates are extremely sensitive to model choice and give rise to point estimates that range between 26 and 192 Ma, depending on the details of the model. Potential problems with the Hawaiian calibration may arise from systematic variation in the molecular clock due to the long generation time of Hawaiian Drosophila compared with other Drosophila and/or uncertainty in linking island formation dates with colonization dates. As either source of error will

  8. Estimating Divergence Dates and Substitution Rates in the Drosophila Phylogeny

    PubMed Central

    Obbard, Darren J.; Maclennan, John; Kim, Kang-Wook; Rambaut, Andrew; O’Grady, Patrick M.; Jiggins, Francis M.

    2012-01-01

    An absolute timescale for evolution is essential if we are to associate evolutionary phenomena, such as adaptation or speciation, with potential causes, such as geological activity or climatic change. Timescales in most phylogenetic studies use geologically dated fossils or phylogeographic events as calibration points, but more recently, it has also become possible to use experimentally derived estimates of the mutation rate as a proxy for substitution rates. The large radiation of drosophilid taxa endemic to the Hawaiian islands has provided multiple calibration points for the Drosophila phylogeny, thanks to the "conveyor belt" process by which this archipelago forms and is colonized by species. However, published date estimates for key nodes in the Drosophila phylogeny vary widely, and many are based on simplistic models of colonization and coalescence or on estimates of island age that are not current. In this study, we use new sequence data from seven species of Hawaiian Drosophila to examine a range of explicit coalescent models and estimate substitution rates. We use these rates, along with a published experimentally determined mutation rate, to date key events in drosophilid evolution. Surprisingly, our estimate for the date for the most recent common ancestor of the genus Drosophila based on mutation rate (25–40 Ma) is closer to being compatible with independent fossil-derived dates (20–50 Ma) than are most of the Hawaiian-calibration models and also has smaller uncertainty. We find that Hawaiian-calibrated dates are extremely sensitive to model choice and give rise to point estimates that range between 26 and 192 Ma, depending on the details of the model. Potential problems with the Hawaiian calibration may arise from systematic variation in the molecular clock due to the long generation time of Hawaiian Drosophila compared with other Drosophila and/or uncertainty in linking island formation dates with colonization dates. As either source of error will

  9. Stochastic Demography and the Neutral Substitution Rate in Class-Structured Populations

    PubMed Central

    Lehmann, Laurent

    2014-01-01

    The neutral rate of allelic substitution is analyzed for a class-structured population subject to a stationary stochastic demographic process. The substitution rate is shown to be generally equal to the effective mutation rate, and under overlapping generations it can be expressed as the effective mutation rate in newborns when measured in units of average generation time. With uniform mutation rate across classes the substitution rate reduces to the mutation rate. PMID:24594520

  10. β-Nitro substituted carboxylic acids and their cytotoxicity.

    PubMed

    Csuk, René; Heller, Lucie; Siewert, Bianka; Gutnov, Andrey; Seidelmann, Oliver; Wendisch, Volkmar

    2014-08-15

    β-Nitro-substituted ethyl carboxylates are a new class of cytotoxic agents; they can be easily obtained in fair to good yields in a single-step reaction by a Pd-catalyzed asymmetric conjugate addition of aryl boronic acids to 2-nitro-acrylates. Of all the tested derivatives, 2-(4-chlorophenyl)-3-nitropropionic acid ethyl ester (6) is most cytotoxic especially against the human ovarian cancer cell line A2780 therefore making this compound an interesting candidate for further investigations.

  11. 40 CFR 721.6560 - Acrylic acid, polymer with substituted ethene.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.6560 Acrylic acid, polymer with substituted ethene. (a) Chemical... as acrylic acid, polymer with substituted ethene (PMN P-91-521) is subject to reporting under...

  12. 40 CFR 721.10032 - Acrylic acid, polymer with substituted acrylamides (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.10032 Acrylic acid, polymer with substituted acrylamides (generic). (a... generically as acrylic acid, polymer with substituted acrylamides (PMN P-02-269) is subject to reporting...

  13. 40 CFR 721.6560 - Acrylic acid, polymer with substituted ethene.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.6560 Acrylic acid, polymer with substituted ethene. (a) Chemical... as acrylic acid, polymer with substituted ethene (PMN P-91-521) is subject to reporting under...

  14. 40 CFR 721.10032 - Acrylic acid, polymer with substituted acrylamides (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.10032 Acrylic acid, polymer with substituted acrylamides (generic). (a... generically as acrylic acid, polymer with substituted acrylamides (PMN P-02-269) is subject to reporting...

  15. 40 CFR 721.6560 - Acrylic acid, polymer with substituted ethene.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.6560 Acrylic acid, polymer with substituted ethene. (a) Chemical... as acrylic acid, polymer with substituted ethene (PMN P-91-521) is subject to reporting under...

  16. 40 CFR 721.10032 - Acrylic acid, polymer with substituted acrylamides (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.10032 Acrylic acid, polymer with substituted acrylamides (generic). (a... generically as acrylic acid, polymer with substituted acrylamides (PMN P-02-269) is subject to reporting...

  17. 40 CFR 721.10399 - Benzoic acid azo-substituted pyridine (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Benzoic acid azo-substituted pyridine... Specific Chemical Substances § 721.10399 Benzoic acid azo-substituted pyridine (generic). (a) Chemical... as benzoic acid azo-substituted pyridine (PMN P-10-501) is subject to reporting under this...

  18. 40 CFR 721.10399 - Benzoic acid azo-substituted pyridine (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Benzoic acid azo-substituted pyridine... Specific Chemical Substances § 721.10399 Benzoic acid azo-substituted pyridine (generic). (a) Chemical... as benzoic acid azo-substituted pyridine (PMN P-10-501) is subject to reporting under this...

  19. 40 CFR 721.10399 - Benzoic acid azo-substituted pyridine (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Benzoic acid azo-substituted pyridine... Specific Chemical Substances § 721.10399 Benzoic acid azo-substituted pyridine (generic). (a) Chemical... as benzoic acid azo-substituted pyridine (PMN P-10-501) is subject to reporting under this...

  20. 40 CFR 721.10032 - Acrylic acid, polymer with substituted acrylamides (generic).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.10032 Acrylic acid, polymer with substituted acrylamides (generic). (a... generically as acrylic acid, polymer with substituted acrylamides (PMN P-02-269) is subject to reporting...

  1. 40 CFR 721.6560 - Acrylic acid, polymer with substituted ethene.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.6560 Acrylic acid, polymer with substituted ethene. (a) Chemical... as acrylic acid, polymer with substituted ethene (PMN P-91-521) is subject to reporting under...

  2. 40 CFR 721.10032 - Acrylic acid, polymer with substituted acrylamides (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.10032 Acrylic acid, polymer with substituted acrylamides (generic). (a... generically as acrylic acid, polymer with substituted acrylamides (PMN P-02-269) is subject to reporting...

  3. 40 CFR 721.6560 - Acrylic acid, polymer with substituted ethene.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Acrylic acid, polymer with substituted... Specific Chemical Substances § 721.6560 Acrylic acid, polymer with substituted ethene. (a) Chemical... as acrylic acid, polymer with substituted ethene (PMN P-91-521) is subject to reporting under...

  4. 40 CFR 721.430 - Oxo-substituted amino-al-kanoic acid derivative.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Oxo-substituted amino-al-kanoic acid... Specific Chemical Substances § 721.430 Oxo-substituted amino-al-kanoic acid derivative. (a) Chemical... as oxo-substituted amino al-kan-oic acid derivative (PMN No. P-92-692) is subject to reporting...

  5. 40 CFR 721.10474 - Substituted amino ethane sulfonic acid salt (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Substituted amino ethane sulfonic acid... Specific Chemical Substances § 721.10474 Substituted amino ethane sulfonic acid salt (generic). (a... generically as substituted amino ethane sulfonic acid salt (PMN P-04-107) is subject to reporting under...

  6. 40 CFR 721.10474 - Substituted amino ethane sulfonic acid salt (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Substituted amino ethane sulfonic acid... Specific Chemical Substances § 721.10474 Substituted amino ethane sulfonic acid salt (generic). (a... generically as substituted amino ethane sulfonic acid salt (PMN P-04-107) is subject to reporting under...

  7. 40 CFR 721.430 - Oxo-substituted amino-al-kanoic acid derivative.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Oxo-substituted amino-al-kanoic acid... Specific Chemical Substances § 721.430 Oxo-substituted amino-al-kanoic acid derivative. (a) Chemical... as oxo-substituted amino al-kan-oic acid derivative (PMN No. P-92-692) is subject to reporting...

  8. 40 CFR 721.430 - Oxo-substituted amino-al-kanoic acid derivative.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Oxo-substituted amino-al-kanoic acid... Specific Chemical Substances § 721.430 Oxo-substituted amino-al-kanoic acid derivative. (a) Chemical... as oxo-substituted amino al-kan-oic acid derivative (PMN No. P-92-692) is subject to reporting...

  9. 40 CFR 721.430 - Oxo-substituted amino-al-kanoic acid derivative.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Oxo-substituted amino-al-kanoic acid... Specific Chemical Substances § 721.430 Oxo-substituted amino-al-kanoic acid derivative. (a) Chemical... as oxo-substituted amino al-kan-oic acid derivative (PMN No. P-92-692) is subject to reporting...

  10. 40 CFR 721.430 - Oxo-substituted amino-al-kanoic acid derivative.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Oxo-substituted amino-al-kanoic acid... Specific Chemical Substances § 721.430 Oxo-substituted amino-al-kanoic acid derivative. (a) Chemical... as oxo-substituted amino al-kan-oic acid derivative (PMN No. P-92-692) is subject to reporting...

  11. Monoclonal antibodies recognizing single amino acid substitutions in hemoglobin

    SciTech Connect

    Stanker, L.H.; Branscomb, E.; Vanderlaan, M.; Jensen, R.H.

    1986-06-01

    Four monoclonal antibodies (mAb) to non-human primate hemoglobin referred to as Cap-4, Cap-5, Rh-2, and Rh-4, and two mAb to human hemoglobin, referred to as H-1 and H-3 were isolated and were partially characterized. Binding studies with these mAb on a panel of hemoglobins and isolated ..cap alpha.. and ..beta.. globin chains revealed a unique reactivity pattern for each mAb. Amino acid sequence analysis of the antigens used to generate the binding data suggests that the specific recognition of certain hemoglobin antigens by each mAb is controlled by the presence of a particular amino acid at a specific position within the epitope. The use of synthetic peptides as antigens confirmed this observation for five of the mAb. No synthetic peptides were tested with the sixth mAb, Rh-2. The amino acids required for binding of mAb Cap-4, Cap-5, Rh-4, and Rh-2 to hemoglobin are alanine at ..beta..5, threonine at ..beta..13, glutamine at ..beta..125, and leucine at ..cap alpha..68. The non-human primate hemoglobin antibodies require a specific amino acid that is not present in human hemoglobin. The amino acid required for binding of Cap-4, Cap-5, and Rh-4 could arise by a single base change in the ..beta.. globin gene, whereas the amino acid required for Rh-2 binding could only occur if two base changes occurred. Thus these mAb are candidate probes for a somatic cell mutation assay on the basis of the detection of peripheral blood red cells that possess single amino acid substituted hemoglobin as a result of single base substitutions in the globin genes of precursor cells.

  12. 40 CFR 721.321 - Substituted acrylamides and acrylic acid copolymer (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Substituted acrylamides and acrylic... New Uses for Specific Chemical Substances § 721.321 Substituted acrylamides and acrylic acid copolymer... identified generically as substituted acrylamides and acrylic acid copolymer (PMN P-00-0490) is subject...

  13. 40 CFR 721.321 - Substituted acrylamides and acrylic acid copolymer (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Substituted acrylamides and acrylic... New Uses for Specific Chemical Substances § 721.321 Substituted acrylamides and acrylic acid copolymer... identified generically as substituted acrylamides and acrylic acid copolymer (PMN P-00-0490) is subject...

  14. 40 CFR 721.321 - Substituted acrylamides and acrylic acid copolymer (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Substituted acrylamides and acrylic... New Uses for Specific Chemical Substances § 721.321 Substituted acrylamides and acrylic acid copolymer... identified generically as substituted acrylamides and acrylic acid copolymer (PMN P-00-0490) is subject...

  15. 40 CFR 721.321 - Substituted acrylamides and acrylic acid copolymer (generic).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Substituted acrylamides and acrylic... New Uses for Specific Chemical Substances § 721.321 Substituted acrylamides and acrylic acid copolymer... identified generically as substituted acrylamides and acrylic acid copolymer (PMN P-00-0490) is subject...

  16. 40 CFR 721.321 - Substituted acrylamides and acrylic acid copolymer (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Substituted acrylamides and acrylic... New Uses for Specific Chemical Substances § 721.321 Substituted acrylamides and acrylic acid copolymer... identified generically as substituted acrylamides and acrylic acid copolymer (PMN P-00-0490) is subject...

  17. Electroanalysis of amino acid substitutions in bioengineered acetylcholinesterase.

    PubMed

    Somji, Mehdi; Dounin, Vladimir; Muench, Susanne B; Schulze, Holger; Bachmann, Till T; Kerman, Kagan

    2012-12-01

    This study reports the electrochemical profiling of Nippostrongylus brasiliensis acetylcholinesterase (AChE) wild-type and mutant proteins. An irreversible oxidation signal of electro-active tyrosine (Y), tryptophan (W) and cysteine (C) residues in five mutant proteins along with the wild-type AChE were detected using square-wave voltammetry (SWV) on screen-printed carbon electrodes. Significant differences were observed in the W303L, T65Y and M301W substituted proteins showing a 25-35% higher peak current intensity compared to the Y349Y and F345Y mutants. It was predicted that AChE substituted with electrochemically active residues would produce the greatest signals and this trend was observed in the T65Y, M301W and Y349L mutants. However, conformational changes in the proteins structure as a result of the substitutions appeared to be most influential on peak current intensities. This was demonstrated by the W303L and F345Y mutant enzymes. The current intensity of W303L was greatest despite the removal of its electro-active W residue whereas the F345Y mutant had the lowest peak value despite the addition of an electro-active Y residue. The preliminary results of this study demonstrate that SWV provides a promising tool to probe the presence of electro-active amino acid residues on the surface of a protein produced through bioengineering.

  18. Restoring enzyme activity in nonfunctional low erucic acid Brassica napus fatty acid elongase 1 by a single amino acid substitution.

    PubMed

    Katavic, Vesna; Mietkiewska, Elzbieta; Barton, Dennis L; Giblin, E Michael; Reed, Darwin W; Taylor, David C

    2002-11-01

    Genomic fatty acid elongation 1 (FAE1) clones from high erucic acid (HEA) Brassica napus, Brassica rapa and Brassica oleracea, and low erucic acid (LEA) B. napus cv. Westar, were amplified by PCR and expressed in yeast cells under the control of the strong galactose-inducible promoter. As expected, yeast cells expressing the FAE1 genes from HEA Brassica spp. synthesized very long chain monounsaturated fatty acids that are not normally found in yeast, while fatty acid profiles of yeast cells expressing the FAE1 gene from LEA B. napus were identical to control yeast samples. In agreement with published findings regarding different HEA and LEA B. napus cultivars, comparison of FAE1 protein sequences from HEA and LEA Brassicaceae revealed one crucial amino acid difference: the serine residue at position 282 of the HEA FAE1 sequences is substituted by phenylalanine in LEA B. napus cv. Westar. Using site directed mutagenesis, the phenylalanine 282 residue was substituted with a serine residue in the FAE1 polypeptide from B. napus cv. Westar, the mutated gene was expressed in yeast and GC analysis revealed the presence of very long chain monounsaturated fatty acids (VLCMFAs), indicating that the elongase activity was restored in the LEA FAE1 enzyme by the single amino acid substitution. Thus, for the first time, the low erucic acid trait in canola B. napus can be attributed to a single amino acid substitution which prevents the biosynthesis of the eicosenoic and erucic acids.

  19. Evaluating Rare Amino Acid Substitutions (RGC_CAMs) in a Yeast Model Clade

    PubMed Central

    Polzin, Kenneth; Rokas, Antonis

    2014-01-01

    When inferring phylogenetic relationships, not all sites in a sequence alignment are equally informative. One recently proposed approach that takes advantage of this inequality relies on sites that contain amino acids whose replacement requires multiple substitutions. Identifying these so-called RGC_CAM substitutions (after Rare Genomic Changes as Conserved Amino acids-Multiple substitutions) requires that, first, at any given site in the amino acid sequence alignment, there must be a minimum of two different amino acids; second, each amino acid must be present in at least two taxa; and third, the amino acids must require a minimum of two nucleotide substitutions to replace each other. Although theory suggests that RGC_CAM substitutions are expected to be rare and less likely to be homoplastic, the informativeness of RGC_CAM substitutions has not been extensively evaluated in biological data sets. We investigated the quality of RGC_CAM substitutions by examining their degree of homoplasy and internode certainty in nearly 2.7 million aligned amino acid sites from 5,261 proteins from five species belonging to the yeast Saccharomyces sensu stricto clade whose phylogeny is well-established. We identified 2,647 sites containing RGC_CAM substitutions, a number that contrasts sharply with the 100,887 sites containing RGC_non-CAM substitutions (i.e., changes between amino acids that require only a single nucleotide substitution). We found that RGC_CAM substitutions had significantly lower homoplasy than RGC_non-CAM ones; specifically RGC_CAM substitutions showed a per-site average homoplasy index of 0.100, whereas RGC_non-CAM substitutions had a homoplasy index of 0.215. Internode certainty values were also higher for sites containing RGC_CAM substitutions than for RGC_non-CAM ones. These results suggest that RGC_CAM substitutions possess a strong phylogenetic signal and are useful markers for phylogenetic inference despite their rarity. PMID:24637883

  20. SYNTHESIS AND CHARACTERIZATION OF SUBSTITUTED POLY(STYRENE)-b-POLY(ACRYLIC ACID) BLOCK COPOLYMER MICELLES

    SciTech Connect

    Pickel, Deanna L; Pickel, Joseph M; Devenyi, Jozsef; Britt, Phillip F

    2009-01-01

    Block copolymer micelle synthesis and characterization has been extensively studied. In particular, most studies have focused on the properties of the hydrophilic corona due to the micelle corona structure s impact on the biodistribution and biocompatibility. Unfortunately, less attention has been given to the effect of the core block on the micelle stability, morphology, and the rate of diffusion of small molecules from the core. This investigation is focused on the synthesis of block copolymers composed of meta-substituted styrenes and acrylic acid by Atom Transfer Radical Polymerization. Micelles with cores composed of substituted styrenes having Tgs ranging from -30 to 100 oC have been prepared and the size and shape of these micelles were characterized by Static and Dynamic Light Scattering and TEM. In addition, the critical micelle concentration and rate of diffusion of small molecules from the core were determined by fluorimetry using pyrene as the probe.

  1. Amino acid-bile acid based molecules: extremely narrow surfactant nanotubes formed by a phenylalanine-substituted cholic acid.

    PubMed

    Travaglini, Leana; D'Annibale, Andrea; Schillén, Karin; Olsson, Ulf; Sennato, Simona; Pavel, Nicolae V; Galantini, Luciano

    2012-12-21

    An amino acid-substituted bile acid forms tubular aggregates with inner and outer diameters of about 3 and 6 nm. The diameters are unusually small for surfactant self-assembled tubes. The results enhance the spectrum of applications of supramolecular tubules and open up possibilities for investigating a novel class of biological amphiphiles.

  2. Anaerobic biotransformation of roxarsone and related N-substituted phenylarsonic acids

    USGS Publications Warehouse

    Cortinas, I.; Field, J.A.; Kopplin, M.; Garbarino, J.R.; Gandolfi, A.J.; Sierra-Alvarez, R.

    2006-01-01

    Large quantities of arsenic are introduced into the environment through land application of poultry litter containing the organoarsenical feed additive roxarsone (3-nitro-4-hydroxyphenylarsonic acid). The objective of this study was to evaluate the bioconversion of roxarsone and related N-substituted phenylarsonic acid derivatives under anaerobic conditions. The results demonstrate that roxarsone is rapidly transformed in the absence of oxygen to the corresponding aromatic amine, 4-hydroxy-3-aminophenylarsonic acid (HAPA). The formation of HAPA is attributable to the facile reduction of the nitro group. Electron-donating substrates, such as hydrogen gas, glucose, and lactate, stimulated the rate of nitro group reduction, indicating a microbial role. During long-term incubations, HAPA and the closely related 4-aminophenylarsonic acid (4-APA) were slowly biologically eliminated by up to 99% under methanogenic and sulfate-reducing conditions, whereas little or no removal occurred in heat-killed inoculum controls. Arsenite and, to a lesser extent, arsenate were observed as products of the degradation. Freely soluble forms of the inorganic arsenical species accounted for 19-28% of the amino-substituted phenylarsonic acids removed. This constitutes the first report of a biologically catalyzed rupture of the phenylarsonic group under anaerobic conditions. ?? 2006 American Chemical Society.

  3. Rates of molecular evolution and diversification in plants: chloroplast substitution rates correlate with species-richness in the Proteaceae

    PubMed Central

    2013-01-01

    Background Many factors have been identified as correlates of the rate of molecular evolution, such as body size and generation length. Analysis of many molecular phylogenies has also revealed correlations between substitution rates and clade size, suggesting a link between rates of molecular evolution and the process of diversification. However, it is not known whether this relationship applies to all lineages and all sequences. Here, in order to investigate how widespread this phenomenon is, we investigate patterns of substitution in chloroplast genomes of the diverse angiosperm family Proteaceae. We used DNA sequences from six chloroplast genes (6278bp alignment with 62 taxa) to test for a correlation between diversification and the rate of substitutions. Results Using phylogenetically-independent sister pairs, we show that species-rich lineages of Proteaceae tend to have significantly higher chloroplast substitution rates, for both synonymous and non-synonymous substitutions. Conclusions We show that the rate of molecular evolution in chloroplast genomes is correlated with net diversification rates in this large plant family. We discuss the possible causes of this relationship, including molecular evolution driving diversification, speciation increasing the rate of substitutions, or a third factor causing an indirect link between molecular and diversification rates. The link between the synonymous substitution rate and clade size is consistent with a role for the mutation rate of chloroplasts driving the speed of reproductive isolation. We find no significant differences in the ratio of non-synonymous to synonymous substitutions between lineages differing in net diversification rate, therefore we detect no signal of population size changes or alteration in selection pressures that might be causing this relationship. PMID:23497266

  4. Ideal amino acid exchange forms for approximating substitution matrices.

    PubMed

    Pokarowski, Piotr; Kloczkowski, Andrzej; Nowakowski, Szymon; Pokarowska, Maria; Jernigan, Robert L; Kolinski, Andrzej

    2007-11-01

    We have analyzed 29 published substitution matrices (SMs) and five statistical protein contact potentials (CPs) for comparison. We find that popular, 'classical' SMs obtained mainly from sequence alignments of globular proteins are mostly correlated by at least a value of 0.9. The BLOSUM62 is the central element of this group. A second group includes SMs derived from alignments of remote homologs or transmembrane proteins. These matrices correlate better with classical SMs (0.8) than among themselves (0.7). A third group consists of intermediate links between SMs and CPs - matrices and potentials that exhibit mutual correlations of at least 0.8. Next, we show that SMs can be approximated with a correlation of 0.9 by expressions c(0) + x(i)x(j) + y(i)y(j) + z(i)z(j), 1acids, respectively. The present paper is the continuation of our work (Pokarowski et al., Proteins 2005;59:49-57), where similar approximation were used to derive ideal amino acid interaction forms from CPs. Both approximations allow us to understand general trends in amino acid similarity and can help improve multiple sequence alignments using the fast Fourier transform (MAFFT), fast threading or another methods based on alignments of physicochemical profiles of protein sequences. The use of this approximation in sequence alignments instead of a classical SM yields results that differ by less than 5%. Intermediate links between SMs and CPs, new formulas for approximating these matrices, and the highly significant dependence of classical SMs on coil preferences are new findings.

  5. Work-rate of substitutes in elite soccer: a preliminary study.

    PubMed

    Carling, Christopher; Espié, Vincent; Le Gall, Franck; Bloomfield, Jonathan; Jullien, Hugues

    2010-03-01

    The aim of this study was to investigate the work-rate of substitutes in professional soccer. A computerised player tracking system was used to assess the work-rates of second-half substitutes (11 midfielders and 14 forwards) in a French Ligue 1 club. Total distance, distance covered in five categories of movement intensity and recovery time between high-intensity efforts were evaluated. First- and second-half work-rates of the replaced players were compared. The performance of substitutes was compared to that of the players they replaced, to team-mates in the same position who remained on the pitch after the substitution and in relation to their habitual performances when starting games. No differences in work-rate between first- and second-halves were observed in all players who were substituted. In the second-half, a non-significant trend was observed in midfield substitutes who covered greater distances than the player they replaced whereas no differences were observed in forwards. Midfield substitutes covered a greater overall distance and distance at high-intensities (p<0.01) and had a lower recovery time between high-intensity efforts (p<0.01) compared to other midfield team-mates who remained on the pitch. Forwards covered less distance (p<0.01) in their first 10-min as a substitute compared to their habitual work-rate profile in the opening 10-min when starting matches while this finding was not observed in midfielders. These findings suggest that compared to midfield substitutes, forward substitutes did not utilise their full physical potential. Further investigation is warranted into the reasons behind this finding in order to optimise the work-rate contributions of forward substitutes.

  6. 40 CFR 721.1875 - Boric acid, alkyl and substituted alkyl esters (generic name).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Boric acid, alkyl and substituted alkyl esters (generic name). 721.1875 Section 721.1875 Protection of Environment ENVIRONMENTAL... Significant New Uses for Specific Chemical Substances § 721.1875 Boric acid, alkyl and substituted...

  7. 40 CFR 721.1875 - Boric acid, alkyl and substituted alkyl esters (generic name).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Boric acid, alkyl and substituted alkyl esters (generic name). 721.1875 Section 721.1875 Protection of Environment ENVIRONMENTAL... Significant New Uses for Specific Chemical Substances § 721.1875 Boric acid, alkyl and substituted...

  8. 40 CFR 721.1875 - Boric acid, alkyl and substituted alkyl esters (generic name).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Boric acid, alkyl and substituted alkyl esters (generic name). 721.1875 Section 721.1875 Protection of Environment ENVIRONMENTAL... Significant New Uses for Specific Chemical Substances § 721.1875 Boric acid, alkyl and substituted...

  9. 40 CFR 721.1875 - Boric acid, alkyl and substituted alkyl esters (generic name).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Boric acid, alkyl and substituted alkyl esters (generic name). 721.1875 Section 721.1875 Protection of Environment ENVIRONMENTAL... Significant New Uses for Specific Chemical Substances § 721.1875 Boric acid, alkyl and substituted...

  10. 40 CFR 721.1875 - Boric acid, alkyl and substituted alkyl esters (generic name).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Boric acid, alkyl and substituted alkyl esters (generic name). 721.1875 Section 721.1875 Protection of Environment ENVIRONMENTAL... Significant New Uses for Specific Chemical Substances § 721.1875 Boric acid, alkyl and substituted...

  11. 40 CFR 721.7770 - Alkylphenoxypoly(oxyethylene) sulfuric acid ester, substituted amine salt.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... acid ester, substituted amine salt. 721.7770 Section 721.7770 Protection of Environment ENVIRONMENTAL... ester, substituted amine salt. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as alkyl phenoxypoly(oxyethylene) sulfuric acid ester,...

  12. Antifungal activity of 4-substituted crotonic acid esters.

    PubMed

    Gershon, H; Shanks, L; Gawiak, D E

    1976-08-01

    Twenty-three 4-substituted crotonic acid esters were tested for antifungal activity against Candida albicans, Aspergillus niger, Mucor mucedo, and Trichophyton mentagrophytes. For the analogues of the methyl ester containing substituents in the 4 position, the following order of fungitoxicity was observed: I greater than Br greater than Cl greater than CH3S greater than CH3O greater than F=H. Of the homologues of the esters of the 4-iodo and 4-bromo compounds which included methyl, ethyl, n-propyl, n-butyl, n-pentyl, and n-hexyl, ethyl 4-iodocrotonate was most toxic to the four fungi at pH 7.0 in the presence of 10% beef serum (C. albicans, 18mug/ml, A. niger, 40 mug/ml, M. mucedo, 5 mug/ml, T. mentagrophytes, 4 mug/ml). It is believed that the mechanism of fungitoxicity is due, in part, to a nucleophilic reaction involving SH-containing compounds. This is based on the correlation of fungitoxicity with the order of leaving groups in the nucleophilic reaction and the protection against the toxicity of the test compounds to the fungi by cysteine and glutathione.

  13. An injectable oxidated hyaluronic acid/adipic acid dihydrazide hydrogel as a vitreous substitute.

    PubMed

    Su, Wen-Yu; Chen, Ko-Hua; Chen, Yu-Chun; Lee, Yen-Hsien; Tseng, Ching-Li; Lin, Feng-Huei

    2011-01-01

    Vitrectomy is a common procedure for treating ocular-related diseases. The surgery involves removing the vitreous humor from the center of the eye, and vitreous substitutes are needed to replace the vitreous humor after vitrectomy. In the present study, we developed a colorless, transparent and injectable hydrogel with appropriate refractive index as a vitreous substitute. The hydrogel is formed by oxidated hyaluronic acid (oxi-HA) cross-linked with adipic acid dihydrazide (ADH). Hyaluronic acid (HA) was oxidized by sodium periodate to create aldehyde functional groups, which could be cross-linked by ADH. The refractive index of this hydrogel ranged between 1.3420 and 1.3442, which is quite similar to human vitreous humor (1.3345). The degradation tests demonstrated that the hydrogel could maintain the gel matrix over 35 days, depending on the ADH concentration. In addition, the cytotoxicity was evaluated on retina pigmented epithelium (RPE) cells cultivated following the ISO standard (tests for in vitro cytotoxicity), and the hydrogel was found to be non-toxic. In a preliminary animal study, the oxi-HA/ADH hydrogel was injected into the vitreous cavity of rabbit eyes. The evaluations of slit-lamp observation, intraocular pressure, cornea thickness and histological examination showed no significant abnormal biological reactions for 3 weeks. This study suggests that the injectable oxi-HA/ADH hydrogel should be a potential vitreous substitute. PMID:20843434

  14. Amino acid substitution at the Adh locus of Drosophila is facilitated by small population size.

    PubMed Central

    Ohta, T

    1993-01-01

    The number of amino acid replacement substitutions and that of synonymous substitutions are examined by using DNA sequences of the Adh locus of Drosophila. The ratio of replacement to synonymous substitutions is higher in sequence comparisons between species than in polymorphisms within species. The ratio for the between-species comparisons is highest in the Hawaiian group and lowest in the obscura group. These observations suggest that amino acid substitutions are facilitated by small population size. The result is in accord with the nearly neutral theory of molecular evolution. PMID:8506297

  15. Interactions in the solid state. II: Interaction of sodium bicarbonate with substituted benzoic acids in the presence of moisture.

    PubMed

    Wright, J L; Carstensen, J T

    1986-06-01

    The interaction of an organic acid with sodium bicarbonate in water produces an effervescent reaction. The reaction products are carbon dioxide, water, and the sodium salt of the acid. The kinetic rate-determining step for this reaction is the dehydration of carbonic acid. The solid-solid interaction with known amounts of moisture was followed by quantitatively determining carbon dioxide evolution as a function of time. The aqueous solubilities, diffusion coefficients, dissociation constants, and solid-solid interaction rates of six different substituted benzoic acids were determined. Using a model based on diffusion of the organic acid through the aqueous layer coupled with chemical reaction, predicted rates and levels of carbon dioxide production were compared with experimental results. Included in the model were the effects of the reaction products on the solution properties of the reactants. It was found that high concentrations of substituted sodium benzoate were generated very quickly and affected the solubility of the reactants, diffusion coefficient of the acid, and the carbonic acid dehydration rate constant. Moisture content was found to have a profound influence on the interaction rate. Water provides a medium for diffusion of the reacting species as well as the reaction solvent. PMID:3016234

  16. Interactions in the solid state. II: Interaction of sodium bicarbonate with substituted benzoic acids in the presence of moisture

    SciTech Connect

    Wright, J.L.; Carstensen, J.T.

    1986-06-01

    The interaction of an organic acid with sodium bicarbonate in water produces an effervescent reaction. The reaction products are carbon dioxide, water, and the sodium salt of the acid. The kinetic rate-determining step for this reaction is the dehydration of carbonic acid. The solid-solid interaction with known amounts of moisture was followed by quantitatively determining carbon dioxide evolution as a function of time. The aqueous solubilities, diffusion coefficients, dissociation constants, and solid-solid interaction rates of six different substituted benzoic acids were determined. Using a model based on diffusion of the organic acid through the aqueous layer coupled with chemical reaction, predicted rates and levels of carbon dioxide production were compared with experimental results. Included in the model were the effects of the reaction products on the solution properties of the reactants. It was found that high concentrations of substituted sodium benzoate were generated very quickly and affected the solubility of the reactants, diffusion coefficient of the acid, and the carbonic acid dehydration rate constant. Moisture content was found to have a profound influence on the interaction rate. Water provides a medium for diffusion of the reacting species as well as the reaction solvent.

  17. Effect of D-amino acid substitutions on Ni(II)-assisted peptide bond hydrolysis.

    PubMed

    Ariani, Hanieh H; Polkowska-Nowakowska, Agnieszka; Bal, Wojciech

    2013-03-01

    Previously we demonstrated the sequence-specific hydrolysis of the R1-(Ser/Thr)-peptide bond in Ni(II) complexes of peptides with a general R1-(Ser/Thr)-Xaa-His-Zaa-R2 sequence (R1 and R2 being any sequences) (Kopera, E.; Krezel, A.; Protas, A. M.; Belczyk, A.; Bonna, A.; Wyslouch-Cieszynska, A.; Poznanski, J.; Bal, W. Inorg. Chem. 2010, 49, 6636). In order to refine our understanding of the mechanism of this reaction and to find ways to accelerate it, we undertook a systematic study of effects of d-amino acid substitutions in the template Ac-Gly-Ala-Ser-Arg-His-Trp-Lys-Phe-Leu-NH2 peptide on the hydrolysis rate constants. We found that all stereochemical alterations made around the Ni(II) chelate plane resulted in the decrease of the reaction rate. However, the Ni(II) coordination, a prerequisite to the reaction, was not compromised by these substitutions. We demonstrated that the reaction is only possible when either the side chain of the crucial Ser (or Thr) residue is on the same part of the chelate plane as the next residue in the sequence (Arg), or the side chain of the residue following His (Trp) resides on the opposite side of the plane. The rate of reaction is the fastest when both these conditions are fulfilled. Another novel effect is the strong dependence of the rate of the acyl shift step on the character of the leaving group. PMID:23427909

  18. Theoretical Analysis of Kinetic Isotope Effects on Proton Transfer Reactions between Substituted α-Methoxystyrenes and Substituted Acetic Acids

    PubMed Central

    Wong, Kin Yiu; Richard, John P.; Gao, Jiali

    2009-01-01

    Primary kinetic isotope effects (KIEs) on a series of carboxylic acid-catalyzed protonation reactions of aryl-substituted α-methoxystyrenes (X-1) to form oxocarbenium ions have been computed using the Kleinert variational second-order perturbation theory (KP2) in the framework of Feynman path integrals (PI) along with the potential energy surface obtained at the B3LYP/6-31+G(d,p) level. Good agreement with the experimental data was obtained, demonstrating that this novel computational approach for computing KIEs of organic reactions is a viable alternative to the traditional method employing Bigeleisen equation and harmonic vibrational frequencies. Although tunneling makes relative small contributions to the lowering of the free energy barriers for the carboxylic acid catalyzed protonation reaction, it is necessary to include tunneling contributions to obtain quantitative estimates of the KIEs. Consideration of anharmonicity can further improve the calculated KIEs for the protonation of substituted α-methoxystyrenes by chloroacetic acid, but for the reactions of the parent and 4-NO2 substituted α-methoxystyrene with substituted carboxylic acids, the correction of anharmonicity overestimates the computed KIEs for strong acid catalysts. In agreement with experimental findings, the largest KIEs are found in nearly ergoneutral reactions, ΔGo ≈ 0, where the transition structures are nearly symmetric and the reaction barriers are relatively low. Furthermore, the optimized transition structures are strongly dependent on the free energy for the formation of the carbocation intermediate, i.e., the driving force ΔGo, along with a good correlation of Hammond shift in the transition state structure. PMID:19754046

  19. Theoretical analysis of kinetic isotope effects on proton transfer reactions between substituted alpha-methoxystyrenes and substituted acetic acids.

    PubMed

    Wong, Kin-Yiu; Richard, John P; Gao, Jiali

    2009-10-01

    Primary kinetic isotope effects (KIEs) on a series of carboxylic acid-catalyzed protonation reactions of aryl-substituted alpha-methoxystyrenes (X-1) to form oxocarbenium ions have been computed using the second-order Kleinert variational perturbation theory (KP2) in the framework of Feynman path integrals (PI) along with the potential energy surface obtained at the B3LYP/6-31+G(d,p) level. Good agreement with the experimental data was obtained, demonstrating that this novel computational approach for computing KIEs of organic reactions is a viable alternative to the traditional method employing Bigeleisen equation and harmonic vibrational frequencies. Although tunneling makes relatively small contributions to the lowering of the free energy barriers for the carboxylic acid catalyzed protonation reaction, it is necessary to include tunneling contributions to obtain quantitative estimates of the KIEs. Consideration of anharmonicity can further improve the calculated KIEs for the protonation of substituted alpha-methoxystyrenes by chloroacetic acid, but for the reactions of the parent and 4-NO(2) substituted alpha-methoxystyrene with substituted carboxylic acids, the correction of anharmonicity overestimates the computed KIEs for strong acid catalysts. In agreement with experimental findings, the largest KIEs are found in nearly ergoneutral reactions, DeltaG(o) approximately 0, where the transition structures are nearly symmetric and the reaction barriers are relatively low. Furthermore, the optimized transition structures are strongly dependent on the free energy for the formation of the carbocation intermediate, that is, the driving force DeltaG(o), along with a good correlation of Hammond shift in the transition state structure.

  20. Strong evidence for lineage and sequence specificity of substitution rates and patterns in Drosophila.

    PubMed

    Singh, Nadia D; Arndt, Peter F; Clark, Andrew G; Aquadro, Charles F

    2009-07-01

    Rates of single nucleotide substitution in Drosophila are highly variable within the genome, and several examples illustrate that evolutionary rates differ among Drosophila species as well. Here, we use a maximum likelihood method to quantify lineage-specific substitutional patterns and apply this method to 4-fold degenerate synonymous sites and introns from more than 8,000 genes aligned in the Drosophila melanogaster group. We find that within species, different classes of sequence evolve at different rates, with long introns evolving most slowly and short introns evolving most rapidly. Relative rates of individual single nucleotide substitutions vary approximately 3-fold among lineages, yielding patterns of substitution that are comparatively less GC-biased in the melanogaster species complex relative to Drosophila yakuba and Drosophila erecta. These results are consistent with a model coupling a mutational shift toward reduced GC content, or a shift in mutation-selection balance, in the D. melanogaster species complex, with variation in selective constraint among different classes of DNA sequence. Finally, base composition of coding and intronic sequences is not at equilibrium with respect to substitutional patterns, which primarily reflects the slow rate of the substitutional process. These results thus support the view that mutational and/or selective processes are labile on an evolutionary timescale and that if the process is indeed selection driven, then the distribution of selective constraint is variable across the genome.

  1. 40 CFR 721.7785 - Substituted alkyl aminomethylene polyphosphonic acid, salt (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Substituted alkyl aminomethylene polyphosphonic acid, salt (generic). 721.7785 Section 721.7785 Protection of Environment ENVIRONMENTAL PROTECTION... New Uses for Specific Chemical Substances § 721.7785 Substituted alkyl aminomethylene...

  2. 40 CFR 721.7785 - Substituted alkyl aminomethylene polyphosphonic acid, salt (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Substituted alkyl aminomethylene polyphosphonic acid, salt (generic). 721.7785 Section 721.7785 Protection of Environment ENVIRONMENTAL PROTECTION... New Uses for Specific Chemical Substances § 721.7785 Substituted alkyl aminomethylene...

  3. 40 CFR 721.7785 - Substituted alkyl aminomethylene polyphosphonic acid, salt (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Substituted alkyl aminomethylene polyphosphonic acid, salt (generic). 721.7785 Section 721.7785 Protection of Environment ENVIRONMENTAL PROTECTION... New Uses for Specific Chemical Substances § 721.7785 Substituted alkyl aminomethylene...

  4. 40 CFR 721.7785 - Substituted alkyl aminomethylene polyphosphonic acid, salt (generic).

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Substituted alkyl aminomethylene polyphosphonic acid, salt (generic). 721.7785 Section 721.7785 Protection of Environment ENVIRONMENTAL PROTECTION... New Uses for Specific Chemical Substances § 721.7785 Substituted alkyl aminomethylene...

  5. 40 CFR 721.7785 - Substituted alkyl aminomethylene polyphosphonic acid, salt (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Substituted alkyl aminomethylene polyphosphonic acid, salt (generic). 721.7785 Section 721.7785 Protection of Environment ENVIRONMENTAL PROTECTION... New Uses for Specific Chemical Substances § 721.7785 Substituted alkyl aminomethylene...

  6. 40 CFR 721.7770 - Alkylphenoxypoly(oxyethylene) sulfuric acid ester, substituted amine salt.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... ester, substituted amine salt. (a) Chemical substance and significant new uses subject to reporting. (1... amine salt (PMN P-92-396) is subject to reporting under this section for the significant new uses... acid ester, substituted amine salt. 721.7770 Section 721.7770 Protection of Environment...

  7. 40 CFR 721.7770 - Alkylphenoxypoly(oxyethylene) sulfuric acid ester, substituted amine salt.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... ester, substituted amine salt. (a) Chemical substance and significant new uses subject to reporting. (1... amine salt (PMN P-92-396) is subject to reporting under this section for the significant new uses... acid ester, substituted amine salt. 721.7770 Section 721.7770 Protection of Environment...

  8. 40 CFR 721.7770 - Alkylphenoxypoly(oxyethylene) sulfuric acid ester, substituted amine salt.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... ester, substituted amine salt. (a) Chemical substance and significant new uses subject to reporting. (1... amine salt (PMN P-92-396) is subject to reporting under this section for the significant new uses... acid ester, substituted amine salt. 721.7770 Section 721.7770 Protection of Environment...

  9. 40 CFR 721.7770 - Alkylphenoxypoly(oxyethylene) sulfuric acid ester, substituted amine salt.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... acid ester, substituted amine salt. 721.7770 Section 721.7770 Protection of Environment ENVIRONMENTAL... ester, substituted amine salt. (a) Chemical substance and significant new uses subject to reporting. (1... amine salt (PMN P-92-396) is subject to reporting under this section for the significant new...

  10. Substitution rate comparisons between grasses and palms: synonymous rate differences at the nuclear gene Adh parallel rate differences at the plastid gene rbcL.

    PubMed Central

    Gaut, B S; Morton, B R; McCaig, B C; Clegg, M T

    1996-01-01

    A number of studies have noted that nucleotide substitution rates at the chloroplast-encoded rbcL locus violate the molecular clock principle. Substitution rate variation at this plastid gene is particularly pronounced between palms and grasses; for example, a previous study estimated that substitution rates in rbcL sequences are approximately 5-fold faster in grasses than in palms. To determine whether a proportionate change in substitution rates also occurs in plant nuclear genes, we characterized nucleotide substitution rates in palm and grass sequences for the nuclear gene Adh. In this article, we report that palm sequences evolve at a rate of 2.61 x 10(-9) substitution per synonymous site per year, a rate which is slower than most plant nuclear genes. Grass Adh sequences evolve approximately 2.5-fold faster than palms at synonymous sites. Thus, synonymous rates in nuclear Adh genes show a marked decrease in palms relative to grasses, paralleling the pattern found at the plastid rbcL locus. This shared pattern indicates that synonymous rates are correlated between a nuclear and a plastid gene. Remarkably, nonsynonymous rates do not show this correlation. Nonsynonymous rates vary between two duplicated grass Adh loci, and nonsynonymous rates at the palm Adh locus are not markedly reduced relative to grasses. Images Fig. 3 PMID:8816790

  11. Correlated evolution of nucleotide substitution rates and allelic variation in Mhc-DRB lineages of primates

    PubMed Central

    Garamszegi, László Z; de Groot, Natasja G; Bontrop, Ronald E

    2009-01-01

    Background The major histocompatibility complex (MHC) is a key model of genetic polymorphism. Selection pressure by pathogens or other microevolutionary forces may result in a high rate of non-synonymous substitutions at the codons specifying the contact residues of the antigen binding sites (ABS), and the maintenance of extreme MHC allelic variation at the population/species level. Therefore, selection forces favouring MHC variability for any reason should cause a correlated evolution between substitution rates and allelic polymorphism. To investigate this prediction, we characterised nucleotide substitution rates and allelic polymorphism (i.e. the number of alleles detected in relation to the number of animals screened) of several Mhc class II DRB lineages in 46 primate species, and tested for a correlation between them. Results First, we demonstrate that species-specific and lineage-specific evolutionary constraints favour species- and lineage-dependent substitution rate at the codons specifying the ABS contact residues (i.e. certain species and lineages can be characterised by high substitution rate, while others have low rate). Second, we show that although the degree of the non-synonymous substitution rate at the ABS contact residues was systematically higher than the degree of the synonymous substitution rate, these estimates were strongly correlated when we controlled for species-specific and lineage-specific effects, and also for the fact that different studies relied on different sample size. Such relationships between substitution rates of different types could even be extended to the non-contact residues of the molecule. Third, we provide statistical evidence that increased substitution rate along a MHC gene may lead to allelic variation, as a high substitution rate can be observed in those lineages in which many alleles are maintained. Fourth, we show that the detected patterns were independent of phylogenetic constraints. When we used phylogenetic

  12. Infrared and ultraviolet laser spectroscopy of jet-cooled substituted salicylic acids; substitution effects on the excited state intramolecular proton transfer in salicylic acid

    NASA Astrophysics Data System (ADS)

    Abd El-Hakam Abou El-Nasr, E.; Fujii, A.; Ebata, T.; Mikami, N.

    Substitution effects on the excited state intramolecular proton transfer (ESIPT) in the salicylic acid (SA) frame were studied by electronic and infrared spectroscopy of jet-cooled 5-methoxylsalicylic acid (5-MeOSA), 5-methylsalicylic acid (5-MeSA), 5-fluorosalicylic acid (5-FSA), 6-fluorosalicylic acid (6-FSA), and methyl salicylate (MS). Infrared spectra were measured in the 3 µm region for both the electronic ground (S0) and first excited (S1) states. The electronic excitation/emission spectra of 5-MeSA and 6-FSA showed the typical spectral features of ESIPT, which have been found in the spectra of SA. On the other hand, 5-MeOSA and 5-FSA exhibit a mirror-image relation between their excitation and emission spectra, which has been regarded as a result of the suppression of ESIPT. Despite such a remarkable difference among the electronic spectra, IR spectroscopy shows that a drastic change of the phenolic OH stretching vibration does occur upon electronic excitation of all substituted SAs, that is, the phenolic OH band of all the SAs disappears from the 3 µm region, indicating a large elongation of the phenolic O-H bond (over 0.1 Å) in S1. This result means that the intramolecular hydrogen bond strength is remarkably enhanced by electronic excitation in all the substituted SAs. Substitution effects on ESIPT in dimers are also discussed.

  13. A phylogenetic model for investigating correlated evolution of substitution rates and continuous phenotypic characters.

    PubMed

    Lartillot, Nicolas; Poujol, Raphaël

    2011-01-01

    The comparative approach is routinely used to test for possible correlations between phenotypic or life-history traits. To correct for phylogenetic inertia, the method of independent contrasts assumes that continuous characters evolve along the phylogeny according to a multivariate Brownian process. Brownian diffusion processes have also been used to describe time variations of the parameters of the substitution process, such as the rate of substitution or the ratio of synonymous to nonsynonymous substitutions. Here, we develop a probabilistic framework for testing the coupling between continuous characters and parameters of the molecular substitution process. Rates of substitution and continuous characters are jointly modeled as a multivariate Brownian diffusion process of unknown covariance matrix. The covariance matrix, the divergence times and the phylogenetic variations of substitution rates and continuous characters are all jointly estimated in a Bayesian Monte Carlo framework, imposing on the covariance matrix a prior conjugate to the Brownian process so as to achieve a greater computational efficiency. The coupling between rates and phenotypes is assessed by measuring the posterior probability of positive or negative covariances, whereas divergence dates and phenotypic variations are marginally reconstructed in the context of the joint analysis. As an illustration, we apply the model to a set of 410 mammalian cytochrome b sequences. We observe a negative correlation between the rate of substitution and mass and longevity, which was previously observed. We also find a positive correlation between ω = dN/dS and mass and longevity, which we interpret as an indirect effect of variations of effective population size, thus in partial agreement with the nearly neutral theory. The method can easily be extended to any parameter of the substitution process and to any continuous phenotypic or environmental character. PMID:20926596

  14. 40 CFR 721.10690 - Benzenedicarboxylic acid, polymer with substituted alkanediol, dodecanedioic acid, 1,2-ethanediol...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Benzenedicarboxylic acid, polymer with substituted alkanediol, dodecanedioic acid, 1,2-ethanediol, alkanedioic acid, alkanediol,.alpha.-hydro-.omega.-hydroxypoly , 1,3-isobenzofurandione, methylene diphenyl diisocyanate, 2-oxepanone, 2,2'-oxybis and polymethylene polyphenylene...

  15. Genes Translocated into the Plastid Inverted Repeat Show Decelerated Substitution Rates and Elevated GC Content.

    PubMed

    Li, Fay-Wei; Kuo, Li-Yaung; Pryer, Kathleen M; Rothfels, Carl J

    2016-01-01

    Plant chloroplast genomes (plastomes) are characterized by an inverted repeat (IR) region and two larger single copy (SC) regions. Patterns of molecular evolution in the IR and SC regions differ, most notably by a reduced rate of nucleotide substitution in the IR compared to the SC region. In addition, the organization and structure of plastomes is fluid, and rearrangements through time have repeatedly shuffled genes into and out of the IR, providing recurrent natural experiments on how chloroplast genome structure can impact rates and patterns of molecular evolution. Here we examine four loci (psbA, ycf2, rps7, and rps12 exon 2-3) that were translocated from the SC into the IR during fern evolution. We use a model-based method, within a phylogenetic context, to test for substitution rate shifts. All four loci show a significant, 2- to 3-fold deceleration in their substitution rate following translocation into the IR, a phenomenon not observed in any other, nontranslocated plastid genes. Also, we show that after translocation, the GC content of the third codon position and of the noncoding regions is significantly increased, implying that gene conversion within the IR is GC-biased. Taken together, our results suggest that the IR region not only reduces substitution rates, but also impacts nucleotide composition. This finding highlights a potential vulnerability of correlating substitution rate heterogeneity with organismal life history traits without knowledge of the underlying genome structure. PMID:27401175

  16. Genes Translocated into the Plastid Inverted Repeat Show Decelerated Substitution Rates and Elevated GC Content

    PubMed Central

    Li, Fay-Wei; Kuo, Li-Yaung; Pryer, Kathleen M.; Rothfels, Carl J.

    2016-01-01

    Plant chloroplast genomes (plastomes) are characterized by an inverted repeat (IR) region and two larger single copy (SC) regions. Patterns of molecular evolution in the IR and SC regions differ, most notably by a reduced rate of nucleotide substitution in the IR compared to the SC region. In addition, the organization and structure of plastomes is fluid, and rearrangements through time have repeatedly shuffled genes into and out of the IR, providing recurrent natural experiments on how chloroplast genome structure can impact rates and patterns of molecular evolution. Here we examine four loci (psbA, ycf2, rps7, and rps12 exon 2–3) that were translocated from the SC into the IR during fern evolution. We use a model-based method, within a phylogenetic context, to test for substitution rate shifts. All four loci show a significant, 2- to 3-fold deceleration in their substitution rate following translocation into the IR, a phenomenon not observed in any other, nontranslocated plastid genes. Also, we show that after translocation, the GC content of the third codon position and of the noncoding regions is significantly increased, implying that gene conversion within the IR is GC-biased. Taken together, our results suggest that the IR region not only reduces substitution rates, but also impacts nucleotide composition. This finding highlights a potential vulnerability of correlating substitution rate heterogeneity with organismal life history traits without knowledge of the underlying genome structure. PMID:27401175

  17. Between peptides and bile acids: self-assembly of phenylalanine substituted cholic acids.

    PubMed

    Travaglini, Leana; D'Annibale, Andrea; di Gregorio, Maria Chiara; Schillén, Karin; Olsson, Ulf; Sennato, Simona; Pavel, Nicolae V; Galantini, Luciano

    2013-08-01

    Biocompatible molecules that undergo self-assembly are of high importance in biological and medical applications of nanoscience. Peptides and bile acids are among the most investigated due to their ability to self-organize into many different, often stimuli-sensitive, supramolecular structures. With the aim of preparing molecules mixing the aggregation properties of bile acid and amino acid-based molecules, we report on the synthesis and self-association behavior of two diastereomers obtained by substituting a hydroxyl group of cholic acid with a l-phenylalanine residue. The obtained molecules are amphoteric, and we demonstrate that they show a pH-dependent self-assembly. Both molecules aggregate in globular micelles at high pH, whereas they form tubular superstructures under acid conditions. Unusual narrow nanotubes with outer and inner cross-section diameters of about 6 and 3 nm are formed by the derivatives. The diasteroisomer with α orientation of the substituent forms in addition a wider tubule (17 nm cross-section diameter). The ability to pack in supramolecular tubules is explained in terms of a wedge-shaped bola-form structure of the derivatives. Parallel or antiparallel face-to-face dimers are hypothesized as fundamental building blocks for the formation of the narrow and wide nanotubes, respectively.

  18. Trends in Hanging and Firearm Suicide Rates in Australia: Substitution of Method?

    ERIC Educational Resources Information Center

    De Leo, Diego; Dwyer, Jonathan; Firman, David; Neulinger, Kerryn

    2003-01-01

    This study examined the increase in the rate of suicide by hanging and an apparently simultaneous decrease in the rate of suicide by firearm as hypothetical evidence that Australian males have substituted one method of suicide for another. Individual suicide method choice may be related to independent changes in the social acceptability of each…

  19. Estimating Divergence Times and Substitution Rates in Rhizobia

    PubMed Central

    Chriki-Adeeb, Rim; Chriki, Ali

    2016-01-01

    Accurate estimation of divergence times of soil bacteria that form nitrogen-fixing associations with most leguminous plants is challenging because of a limited fossil record and complexities associated with molecular clocks and phylogenetic diversity of root nodule bacteria, collectively called rhizobia. To overcome the lack of fossil record in bacteria, divergence times of host legumes were used to calibrate molecular clocks and perform phylogenetic analyses in rhizobia. The 16S rRNA gene and intergenic spacer region remain among the favored molecular markers to reconstruct the timescale of rhizobia. We evaluate the performance of the random local clock model and the classical uncorrelated lognormal relaxed clock model, in combination with four tree models (coalescent constant size, birth–death, birth–death incomplete sampling, and Yule processes) on rhizobial divergence time estimates. Bayes factor tests based on the marginal likelihoods estimated from the stepping-stone sampling analyses strongly favored the random local clock model in combination with Yule process. Our results on the divergence time estimation from 16S rRNA gene and intergenic spacer region sequences are compatible with age estimates based on the conserved core genes but significantly older than those obtained from symbiotic genes, such as nodIJ genes. This difference may be due to the accelerated evolutionary rates of symbiotic genes compared to those of other genomic regions not directly implicated in nodulation processes. PMID:27168719

  20. Estimating Divergence Times and Substitution Rates in Rhizobia.

    PubMed

    Chriki-Adeeb, Rim; Chriki, Ali

    2016-01-01

    Accurate estimation of divergence times of soil bacteria that form nitrogen-fixing associations with most leguminous plants is challenging because of a limited fossil record and complexities associated with molecular clocks and phylogenetic diversity of root nodule bacteria, collectively called rhizobia. To overcome the lack of fossil record in bacteria, divergence times of host legumes were used to calibrate molecular clocks and perform phylogenetic analyses in rhizobia. The 16S rRNA gene and intergenic spacer region remain among the favored molecular markers to reconstruct the timescale of rhizobia. We evaluate the performance of the random local clock model and the classical uncorrelated lognormal relaxed clock model, in combination with four tree models (coalescent constant size, birth-death, birth-death incomplete sampling, and Yule processes) on rhizobial divergence time estimates. Bayes factor tests based on the marginal likelihoods estimated from the stepping-stone sampling analyses strongly favored the random local clock model in combination with Yule process. Our results on the divergence time estimation from 16S rRNA gene and intergenic spacer region sequences are compatible with age estimates based on the conserved core genes but significantly older than those obtained from symbiotic genes, such as nodIJ genes. This difference may be due to the accelerated evolutionary rates of symbiotic genes compared to those of other genomic regions not directly implicated in nodulation processes. PMID:27168719

  1. No variation and low synonymous substitution rates in coral mtDNA despite high nuclear variation

    PubMed Central

    Hellberg, Michael E

    2006-01-01

    Background The mitochondrial DNA (mtDNA) of most animals evolves more rapidly than nuclear DNA, and often shows higher levels of intraspecific polymorphism and population subdivision. The mtDNA of anthozoans (corals, sea fans, and their kin), by contrast, appears to evolve slowly. Slow mtDNA evolution has been reported for several anthozoans, however this slow pace has been difficult to put in phylogenetic context without parallel surveys of nuclear variation or calibrated rates of synonymous substitution that could permit quantitative rate comparisons across taxa. Here, I survey variation in the coding region of a mitochondrial gene from a coral species (Balanophyllia elegans) known to possess high levels of nuclear gene variation, and estimate synonymous rates of mtDNA substitution by comparison to another coral (Tubastrea coccinea). Results The mtDNA surveyed (630 bp of cytochrome oxidase subunit I) was invariant among individuals sampled from 18 populations spanning 3000 km of the range of B. elegans, despite high levels of variation and population subdivision for allozymes over these same populations. The synonymous substitution rate between B. elegans and T. coccinea (0.05%/site/106 years) is similar to that in most plants, but 50–100 times lower than rates typical for most animals. In addition, while substitutions to mtDNA in most animals exhibit a strong bias toward transitions, mtDNA from these corals does not. Conclusion Slow rates of mitochondrial nucleotide substitution result in low levels of intraspecific mtDNA variation in corals, even when nuclear loci vary. Slow mtDNA evolution appears to be the basal condition among eukaryotes. mtDNA substitution rates switch from slow to fast abruptly and unidirectionally. This switch may stem from the loss of just one or a few mitochondrion-specific DNA repair or replication genes. PMID:16542456

  2. Caproic acid grafted chitosan cationic nanocomplexes for enhanced gene delivery: effect of degree of substitution.

    PubMed

    Layek, Buddhadev; Singh, Jagdish

    2013-04-15

    This work was designed to investigate the effect of the degree of substitutions of caproic acid on plasmid DNA (pDNA) binding, cellular uptake, biocompatibility, and transfection efficiency of caproic acid grafted chitosan (CGC). The CGC with three substitution degrees (CGC-5, CGC-15, and CGC-25) were synthesized by coupling caproic acid with chitosan. Chemical characterization of graft polymers was performed using FTIR, NMR, and elemental analysis. The CGC polymers showed good pDNA condensing capacity and efficient protection of pDNA from DNase I. The nanosized CGC/pDNA polyplexes exhibited well-defined spherical shapes and stability in serum. Isothermal titration calorimetry demonstrated reduction in CGC-pDNA binding constant with increase in the degree of caproic acid substitution. Caproic acid substitution resulted in 2-7-fold higher cellular uptake in HEK 293 cells mainly via the clathrin-mediated pathway without affecting biocompatibility. In vitro transfection study suggested a dependence of transfection efficiency on the variability of caproic acid substitution. The CGC-15 polymer exhibited 31-fold and 1.33-fold higher gene expression compared to chitosan and the marketed non-viral vector FuGENEHD, respectively. These finding suggests that the CGC-15 graft polymer is a promising non-viral gene delivery vector due to its superior transfection efficiency and biocompatibility.

  3. 40 CFR 747.195 - Triethanolamine salt of a substituted organic acid.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... Agency, at 40 CFR 747.195, as published in the Federal Register of June 14, 1984. A copy of the... organic acid. 747.195 Section 747.195 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Substances § 747.195 Triethanolamine salt of a substituted organic acid. This section identifies...

  4. 40 CFR 747.195 - Triethanolamine salt of a substituted organic acid.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... Agency, at 40 CFR 747.195, as published in the Federal Register of June 14, 1984. A copy of the... organic acid. 747.195 Section 747.195 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Substances § 747.195 Triethanolamine salt of a substituted organic acid. This section identifies...

  5. 40 CFR 747.195 - Triethanolamine salt of a substituted organic acid.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... Agency, at 40 CFR 747.195, as published in the Federal Register of June 14, 1984. A copy of the... organic acid. 747.195 Section 747.195 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Substances § 747.195 Triethanolamine salt of a substituted organic acid. This section identifies...

  6. 40 CFR 747.195 - Triethanolamine salt of a substituted organic acid.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... Agency, at 40 CFR 747.195, as published in the Federal Register of June 14, 1984. A copy of the... organic acid. 747.195 Section 747.195 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Substances § 747.195 Triethanolamine salt of a substituted organic acid. This section identifies...

  7. 40 CFR 747.195 - Triethanolamine salt of a substituted organic acid.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... Agency, at 40 CFR 747.195, as published in the Federal Register of June 14, 1984. A copy of the... organic acid. 747.195 Section 747.195 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED... Substances § 747.195 Triethanolamine salt of a substituted organic acid. This section identifies...

  8. The INFVo perceptual rating scale for substitution voicing: development and reliability.

    PubMed

    Moerman, Mieke; Martens, Jean-Pierre; Crevier-Buchman, Lise; de Haan, Else; Grand, Stephanie; Tessier, Christophe; Woisard, Virginie; Dejonckere, Philippe

    2006-05-01

    In this paper, an experimental study of inter-judge consistency for the different dimensions of a recently proposed new scale for the rating of substitution voices is presented. The IINFVo rating scale tries to score five parameters, namely impression, intelligibility, noise, fluency and voicing. Each parameter is scored between 0 (very good substitution voicing) and 10 (very deviant substitution voicing) on a visual analogue scale. Inter-judge consistencies were measured among semi-professional as well as among professional jury members. The consistencies among semi-professionals, expressed as Pearson correlation coefficients, ranged from moderate to good (0.57-0.68), whereas those among professionals were good to excellent (0.82-0.87) and compared favourably to consistency figures published for traditional perceptual evaluation scales such as the GRBAS scale for laryngeal dysphonia. Since there is a strong correlation between the scores of impression and intelligibility, and since intelligibility is hard to score by non-native listeners, we suggest taking the mean of the two scores as the "impression" of a modified dimensional INFVo rating scale. Our experiments demonstrate that the INFVo rating scale has good potential to become a routine perceptual evaluation method in a multidimensional assessment protocol for substitution voicing.

  9. 40 CFR 721.6181 - Fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl ether...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Fatty acid, reaction product with... Specific Chemical Substances § 721.6181 Fatty acid, reaction product with substituted oxirane, formaldehyde... as fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl...

  10. 40 CFR 721.6181 - Fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl ether...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Fatty acid, reaction product with... Specific Chemical Substances § 721.6181 Fatty acid, reaction product with substituted oxirane, formaldehyde... as fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl...

  11. 40 CFR 721.6181 - Fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl ether...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Fatty acid, reaction product with... Specific Chemical Substances § 721.6181 Fatty acid, reaction product with substituted oxirane, formaldehyde... as fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl...

  12. 40 CFR 721.6181 - Fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl ether...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Fatty acid, reaction product with... Specific Chemical Substances § 721.6181 Fatty acid, reaction product with substituted oxirane, formaldehyde... as fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl...

  13. 40 CFR 721.6181 - Fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl ether...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Fatty acid, reaction product with... Specific Chemical Substances § 721.6181 Fatty acid, reaction product with substituted oxirane, formaldehyde... as fatty acid, reaction product with substituted oxirane, formaldehyde-phenol polymer glycidyl...

  14. Structure and NLO properties of halogen (F, Cl) substituted formic acid dimers.

    PubMed

    Umadevi, P; Senthilkumar, L; Gayathri, M; Kolandaivel, P

    2014-11-11

    In this work, using ab initio and density functional theory (DFT) methods halogen substituted formic acid (FA) dimer is studied. The dimer stability is due to the hydrogen bonds, either conventional (OH⋯O, OH⋯F, OH⋯Cl) or non-conventional (CH⋯O, CH⋯F, CH⋯Cl). Among all the dimers, trans-trans form is more stable than the trans-cis, and cis-cis form. Basis set extrapolated counterpoise corrected interaction energy results for the FA dimer are in excellent agreement with BSSE corrected MP2 interaction energy. Symmetry Adopted Perturbation Theory (SAPT) analysis reveals that the electrostatic effect plays a dominant role in stabilization among the dimers with maximum interaction energy. Chlorine substituted FA dimer has high hyperpolarizability, which makes them excellent candidate for nonlinear optical materials (NLO). The halogen substituted formic acid dimers have higher stability and polarizability value than the unsubstituted formic acid dimer. The hyperpolarizability values depend on the geometrical structures of halogenated formic acid dimers than the type of hydrogen bonds. The small excitation energy and HOMO-LUMO gap in the halogenated formic acid dimer has led to the strong nonlinear optical response. The depolarization ratio and Rayleigh scattering increases in formic acid dimer after the halogen atom substitution.

  15. Substitution Model Adequacy and Assessing the Reliability of Estimates of Virus Evolutionary Rates and Time Scales.

    PubMed

    Duchêne, Sebastián; Di Giallonardo, Francesca; Holmes, Edward C

    2016-01-01

    Determining the time scale of virus evolution is central to understanding their origins and emergence. The phylogenetic methods commonly used for this purpose can be misleading if the substitution model makes incorrect assumptions about the data. Empirical studies consider a pool of models and select that with the highest statistical fit. However, this does not allow the rejection of all models, even if they poorly describe the data. An alternative is to use model adequacy methods that evaluate the ability of a model to predict hypothetical future observations. This can be done by comparing the empirical data with data generated under the model in question. We conducted simulations to evaluate the sensitivity of such methods with nucleotide, amino acid, and codon data. These effectively detected underparameterized models, but failed to detect mutational saturation and some instances of nonstationary base composition, which can lead to biases in estimates of tree topology and length. To test the applicability of these methods with real data, we analyzed nucleotide and amino acid data sets from the genus Flavivirus of RNA viruses. In most cases these models were inadequate, with the exception of a data set of relatively closely related sequences of Dengue virus, for which the GTR+Γ nucleotide and LG+Γ amino acid substitution models were adequate. Our results partly explain the lack of consensus over estimates of the long-term evolutionary time scale of these viruses, and indicate that assessing the adequacy of substitution models should be routinely used to determine whether estimates are reliable.

  16. Determination of substitution positions in hyaluronic acid hydrogels using NMR and MS based methods.

    PubMed

    Wende, Frida J; Gohil, Suresh; Mojarradi, Hotan; Gerfaud, Thibaud; Nord, Lars I; Karlsson, Anders; Boiteau, Jean-Guy; Kenne, Anne Helander; Sandström, Corine

    2016-01-20

    In hydrogels of cross-linked polysaccharides, the total amount of cross-linker and the degree of cross-linking influence the properties of the hydrogel. The substitution position of the cross-linker on the polysaccharide is another parameter that can influence hydrogel properties; hence methods for detailed structural analysis of the substitution pattern are required. NMR and LC-MS methods were developed to determine the positions and amounts of substitution of 1,4-butanediol diglycidyl ether (BDDE) on hyaluronic acid (HA), and for the first time it is shown that BDDE can react with any of the four available hydroxyl groups of the HA disaccharide repeating unit. This was achieved by studying di-, tetra-, and hexasaccharides obtained from degradation of BDDE cross-linked HA hydrogel by chondroitinase. Furthermore, amount of linker substitution at each position was shown to be dependent on the size of the oligosaccharides. For the disaccharide, substitutions were predominantly at ΔGlcA-OH2 and GlcNAc-OH6 while in the tetra- and hexasaccharides, it was mainly at the reducing end GlcNAc-OH4. In the disaccharide there was no substitution at this position. Since chondroitinase is able to completely hydrolyse non-substituted HA into unsaturated disaccharides, these results indicate that the enzyme is prevented to cleave on the non-reducing side of an oligosaccharide substituted at the reducing end GlcNAc-OH4. The procedure can be adopted for the determination of substitution positions in other types of polymers. PMID:26572480

  17. Surface enhanced Raman spectra of carbonate, hydrocarbonate, and substituted acetic acids on silver hydrosols

    NASA Astrophysics Data System (ADS)

    Kai, Sun; Chaozhi, Wan; Guangzhi, Xu

    1989-01-01

    The SERS spectra of carbonate, hydrocarbonate and several substituted acetic acids absorbed on silver hydrosols are recorded. The greatest enhancement of E' modes is shown in the spectrum of carbonate, from which the carbonate is deduced to be absorbed in an "end on" configuration, rather than flat on the surface. The spectrum of the hydrocarbonate solution shows the most enhanced bands at about 925 and 620 cm -1, which cannot be explained clearly. All the substituted acids have a most enhanced bands at about 1630 cm -1, revealing that the acids are initially adsorbed in a single bonding state through the carboxyl group. The change in the SERS spectra of the acids with time indicates that a bidentate bridging adsorbed state may be formed after some time.

  18. Hydrolysis of α-Chloro-Substituted 2- and 4-Pyridones: Rate Enhancement by Zwitterionic Structure

    PubMed Central

    Tan, Ronald C.; Vien, Janie Q. T.; Wu, Weiming

    2011-01-01

    The hydrolysis of α-chloro-N-methyl-4-pyridone was found to be more than five times faster than that of α-chloro-N-methyl-2-pyridone. Structural studies of 2- and 4-pyridones have revealed the higher polarity and greater extent of zwitterionic content in 4-pyridone. The results are thus consistent with the hypothesis that polarization and higher zwitterionic content in the heterocyclic structures enhances the rate of hydrolysis in α-substituted pyridone and uracil derivatives. PMID:22178555

  19. Hydrolysis of α-chloro-substituted 2- and 4-pyridones: rate enhancement by zwitterionic structure.

    PubMed

    Tan, Ronald C; Vien, Janie Q T; Wu, Weiming

    2012-01-15

    The hydrolysis of α-chloro-N-methyl-4-pyridone was found to be more than five times faster than that of α-chloro-N-methyl-2-pyridone. Structural studies of 2- and 4-pyridones have revealed the higher polarity and greater extent of zwitterionic content in 4-pyridone. The results are thus consistent with the hypothesis that polarization and higher zwitterionic content in the heterocyclic structures enhances the rate of hydrolysis in α-substituted pyridone and uracil derivatives. PMID:22178555

  20. Polyvinyl alcohol and amino acids as substitutes for bovine serum albumin in culture media for mouse preimplantation embryos.

    PubMed

    Biggers, J D; Summers, M C; McGinnis, L K

    1997-01-01

    The effect of replacing bovine serum albumin (BSA) in a simple defined medium (KSOM) with polyvinyl alcohol (PVA) and/or amino acids on the percentages of mouse zygotes that develop to at least the blastocyst stage and that hatch at least partially or completely is reported. Blastocysts could form when BSA was replaced with only PVA, but at a moderately reduced rate; however, partial hatching, and hence complete hatching, were severely impaired when BSA was replaced with only PVA. The substitution of BSA with amino acids alone resulted in a high rate of blastocyst formation and moderate impairment of hatching. The addition of PVA to BSA-free KSOM supplemented with amino acids had no extra effect. BSA had significant effects when added to BSA-free KSOM supplemented with amino acids. The BSA caused a significant increase in the rate of partial hatching, and may even have had a small effect on the rate of blastocyst formation. The results also showed that glucose, at a high concentration of 5.56 mM, does not inhibit the development of mouse zygotes to hatched blastocysts when cultured in KSOM supplemented with amino acids. PMID:9286737

  1. Reactivity of substituted charged phenyl radicals toward components of nucleic acids.

    PubMed

    Ramírez-Arizmendi, Luis E; Heidbrink, Jenny L; Guler, Leonard P; Kenttämaa, Hilkka I

    2003-02-26

    Reactions of differently substituted phenyl radicals with components of nucleic acids have been investigated in the gas phase. A positively charged group located meta with respect to the radical site was employed to allow manipulation of the radicals in a Fourier-transform ion cyclotron resonance mass spectrometer. All of these electrophilic radicals react with sugars via exclusive hydrogen atom abstraction, with adenine and uracil almost exclusively via addition (likely at the C8 and C5 carbons, respectively), and with the nucleoside thymidine by hydrogen atom abstraction and addition at C5 in the base moiety (followed by elimination of (*)CH(3)). These findings parallel the reactivity of the phenyl radical with components of nucleic acids in solution, except that the selectivity for addition is different. Like HO(*), the electrophilic charged phenyl radicals appear to favor addition to the C5-end of the C5-C6 double bond of thymine and thymidine, whereas the phenyl radical preferentially adds to C6. The charged phenyl radicals do not predominantly add to thymine, as the neutral phenyl radical and HO(*), but mainly react by hydrogen atom abstraction from the methyl group (some addition to C5 in the base followed by loss of (*)CH(3) also occurs). Adenine appears to be the preferred target among the nucleobases, while uracil is the least favored. A systematic increase in the electrophilicity of the radicals by modification of the radicals' structures was found to facilitate all reactions, but the addition even more than hydrogen atom abstraction. Therefore, the least reactive radicals are most selective toward hydrogen atom abstraction, while the most reactive radicals also efficiently add to the base. Traditional enthalpy arguments do not rationalize the rate variations. Instead, the rates reflect the radicals' electron affinities used as a measure for their ability to polarize the transition state of each reaction.

  2. 10-Boronic acid substituted camptothecin as prodrug of SN-38.

    PubMed

    Wang, Lei; Xie, Shao; Ma, Longjun; Chen, Yi; Lu, Wei

    2016-06-30

    Malignant tumor cells have been found to have high levels of reactive oxygen species such as hydrogen peroxide (H2O2), supporting the hypothesis that a prodrug could be activated by intracellular H2O2 and lead to a potential antitumor therapy. In this study, the 7-ethyl-10-boronic acid camptothecin (B1) was synthesized for the first time as prodrug of SN-38, by linking a cleavable aryl carbon-boron bond to the SN-38. Prodrug B1 selectively activated by H2O2, converted rapidly to the active form SN-38 under favorable oxidative conditions in cancer cells with elevated levels of H2O2. The cell survival assay showed that prodrug B1 was equally or more effective in inhibiting the growth of six different cancer cells, as compared to SN-38. Unexpectedly, prodrug B1 displayed even more potent Topo I inhibitory activity than SN-38, suggesting that it was not only a prodrug of SN-38 but also a typical Topo I inhibitor. Prodrug B1 also demonstrated a significant antitumor activity at 2.0 mg/kg in a xenograft model using human brain star glioblastoma cell lines U87MG. PMID:27060760

  3. Too packed to change: side-chain packing and site-specific substitution rates in protein evolution.

    PubMed

    Marcos, María Laura; Echave, Julian

    2015-01-01

    In protein evolution, due to functional and biophysical constraints, the rates of amino acid substitution differ from site to site. Among the best predictors of site-specific rates are solvent accessibility and packing density. The packing density measure that best correlates with rates is the weighted contact number (WCN), the sum of inverse square distances between a site's C α and the C α of the other sites. According to a mechanistic stress model proposed recently, rates are determined by packing because mutating packed sites stresses and destabilizes the protein's active conformation. While WCN is a measure of C α packing, mutations replace side chains. Here, we consider whether a site's evolutionary divergence is constrained by main-chain packing or side-chain packing. To address this issue, we extended the stress theory to model side chains explicitly. The theory predicts that rates should depend solely on side-chain contact density. We tested this prediction on a data set of structurally and functionally diverse monomeric enzymes. We compared side-chain contact density with main-chain contact density measures and with relative solvent accessibility (RSA). We found that side-chain contact density is the best predictor of rate variation among sites (it explains 39.2% of the variation). Moreover, the independent contribution of main-chain contact density measures and RSA are negligible. Thus, as predicted by the stress theory, site-specific evolutionary rates are determined by side-chain packing.

  4. Natural derivatives of diphenolic acid as substitutes for bisphenol-A

    NASA Astrophysics Data System (ADS)

    Ertl, Johanna; Cerri, Elisa; Rizzuto, Matteo; Caretti, Daniele

    2014-05-01

    Diphenolic acid had been originally used in the first epoxy resins and was later on forgotten as it was substituted by the cheaper bisphenol A. But in the recent years major health concerns have been raised as bisphenol A has a pseudo-hormonal effect on the body, playing the role of estrogen it can cause a severe impact on the organism, especially in males. Moreover it is produced from acetone and phenol, both from fossil, and thus limited resources. On the contrary, diphenolic acid is synthesized from levulinic acid and phenol. Levulinic acid being directly produced by hydrolysis of biomass. By substituting the fossil phenol with natural phenols from lignin or plant extraction we are able to synthesize a fully renewable substitute for bisphenol A. The reactions to yield an epoxy resin have been examined and the reactivity with epichlorohydrin is satisfying. Moreover, some of the derivatives of diphenolic acid have interesting curing properties and preliminary results show excellent properties of the cured resin, including thermal stability and pencil hardness.

  5. Amino acid substitutions in inherited albumin variants from Amerindian and Japanese populations

    SciTech Connect

    Takahashi, N.; Takahashi, Y.; Isobe, T.; Putnam, F.W.; Fujita, M.; Satoh, C.; Neel, J.V.

    1987-11-01

    The authors report an effort to determine the basis for the altered migration of seven inherited albumin variants detected by one-dimensional electrophoresis in population surveys involving tribal Amerindians and Japanese children. An amino acid substitution has thus far been determined for four of the variants. The randomness in the albumin polypeptide of these and the other sixteen independently ascertained amino acid substitutions of albumin and proalbumin thus far established was analyzed; the clustering of eight of these at two positions in the six-amino acid propeptide sequence seems noteworthy. By comparison with other proteins studied by electrophoresis, albumin exhibits average variability. It is a paradox that individuals who, for genetic reasons, lack albumin exhibit no obvious ill effects; yet, electrophoretic variants of albumin are no more numerous than are variants of proteins, the absence of which results in severe disease.

  6. The probability of speciation on an interaction network with unequal substitution rates.

    PubMed

    Olofsson, Peter; Livingstone, Kevin; Humphreys, Joshua; Steinman, Douglas

    2016-08-01

    Speciation is characterized by the development of reproductive isolating barriers between diverging groups. A seminal paper of a mathematical model of speciation was published by Orr (1995), extended by Livingstone et al. (2012) to incorporate interaction networks. Here, we further develop the model to take into account the possibility of different substitution rates for network nodes of different connectivity. Mathematically, this amounts to sampling nodes from an undirected graph where the inclusion probability for a given node depends on its degree (number of connecting edges). We establish formulas for the rate of speciation and identify a crucial parameter that is a measure of the deviation from simple random sampling. PMID:27177943

  7. Modeling protein evolution with several amino acid replacement matrices depending on site rates.

    PubMed

    Le, Si Quang; Dang, Cuong Cao; Gascuel, Olivier

    2012-10-01

    Most protein substitution models use a single amino acid replacement matrix summarizing the biochemical properties of amino acids. However, site evolution is highly heterogeneous and depends on many factors that influence the substitution patterns. In this paper, we investigate the use of different substitution matrices for different site evolutionary rates. Indeed, the variability of evolutionary rates corresponds to one of the most apparent heterogeneity factors among sites, and there is no reason to assume that the substitution patterns remain identical regardless of the evolutionary rate. We first introduce LG4M, which is composed of four matrices, each corresponding to one discrete gamma rate category (of four). These matrices differ in their amino acid equilibrium distributions and in their exchangeabilities, contrary to the standard gamma model where only the global rate differs from one category to another. Next, we present LG4X, which also uses four different matrices, but leaves aside the gamma distribution and follows a distribution-free scheme for the site rates. All these matrices are estimated from a very large alignment database, and our two models are tested using a large sample of independent alignments. Detailed analysis of resulting matrices and models shows the complexity of amino acid substitutions and the advantage of flexible models such as LG4M and LG4X. Both significantly outperform single-matrix models, providing gains of dozens to hundreds of log-likelihood units for most data sets. LG4X obtains substantial gains compared with LG4M, thanks to its distribution-free scheme for site rates. Since LG4M and LG4X display such advantages but require the same memory space and have comparable running times to standard models, we believe that LG4M and LG4X are relevant alternatives to single replacement matrices. Our models, data, and software are available from http://www.atgc-montpellier.fr/models/lg4x.

  8. Computational study of the rate constants and free energies of intramolecular radical addition to substituted anilines.

    PubMed

    Gansäuer, Andreas; Seddiqzai, Meriam; Dahmen, Tobias; Sure, Rebecca; Grimme, Stefan

    2013-01-01

    The intramolecular radical addition to aniline derivatives was investigated by DFT calculations. The computational methods were benchmarked by comparing the calculated values of the rate constant for the 5-exo cyclization of the hexenyl radical with the experimental values. The dispersion-corrected PW6B95-D3 functional provided very good results with deviations for the free activation barrier compared to the experimental values of only about 0.5 kcal mol(-1) and was therefore employed in further calculations. Corrections for intramolecular London dispersion and solvation effects in the quantum chemical treatment are essential to obtain consistent and accurate theoretical data. For the investigated radical addition reaction it turned out that the polarity of the molecules is important and that a combination of electrophilic radicals with preferably nucleophilic arenes results in the highest rate constants. This is opposite to the Minisci reaction where the radical acts as nucleophile and the arene as electrophile. The substitution at the N-atom of the aniline is crucial. Methyl substitution leads to slower addition than phenyl substitution. Carbamates as substituents are suitable only when the radical center is not too electrophilic. No correlations between free reaction barriers and energies (ΔG (‡) and ΔG R) are found. Addition reactions leading to indanes or dihydrobenzofurans are too slow to be useful synthetically.

  9. Computational study of the rate constants and free energies of intramolecular radical addition to substituted anilines.

    PubMed

    Gansäuer, Andreas; Seddiqzai, Meriam; Dahmen, Tobias; Sure, Rebecca; Grimme, Stefan

    2013-01-01

    The intramolecular radical addition to aniline derivatives was investigated by DFT calculations. The computational methods were benchmarked by comparing the calculated values of the rate constant for the 5-exo cyclization of the hexenyl radical with the experimental values. The dispersion-corrected PW6B95-D3 functional provided very good results with deviations for the free activation barrier compared to the experimental values of only about 0.5 kcal mol(-1) and was therefore employed in further calculations. Corrections for intramolecular London dispersion and solvation effects in the quantum chemical treatment are essential to obtain consistent and accurate theoretical data. For the investigated radical addition reaction it turned out that the polarity of the molecules is important and that a combination of electrophilic radicals with preferably nucleophilic arenes results in the highest rate constants. This is opposite to the Minisci reaction where the radical acts as nucleophile and the arene as electrophile. The substitution at the N-atom of the aniline is crucial. Methyl substitution leads to slower addition than phenyl substitution. Carbamates as substituents are suitable only when the radical center is not too electrophilic. No correlations between free reaction barriers and energies (ΔG (‡) and ΔG R) are found. Addition reactions leading to indanes or dihydrobenzofurans are too slow to be useful synthetically. PMID:24062821

  10. Computational study of the rate constants and free energies of intramolecular radical addition to substituted anilines

    PubMed Central

    Seddiqzai, Meriam; Dahmen, Tobias; Sure, Rebecca

    2013-01-01

    Summary The intramolecular radical addition to aniline derivatives was investigated by DFT calculations. The computational methods were benchmarked by comparing the calculated values of the rate constant for the 5-exo cyclization of the hexenyl radical with the experimental values. The dispersion-corrected PW6B95-D3 functional provided very good results with deviations for the free activation barrier compared to the experimental values of only about 0.5 kcal mol−1 and was therefore employed in further calculations. Corrections for intramolecular London dispersion and solvation effects in the quantum chemical treatment are essential to obtain consistent and accurate theoretical data. For the investigated radical addition reaction it turned out that the polarity of the molecules is important and that a combination of electrophilic radicals with preferably nucleophilic arenes results in the highest rate constants. This is opposite to the Minisci reaction where the radical acts as nucleophile and the arene as electrophile. The substitution at the N-atom of the aniline is crucial. Methyl substitution leads to slower addition than phenyl substitution. Carbamates as substituents are suitable only when the radical center is not too electrophilic. No correlations between free reaction barriers and energies (ΔG ‡ and ΔG R) are found. Addition reactions leading to indanes or dihydrobenzofurans are too slow to be useful synthetically. PMID:24062821

  11. Microheterogeneity of antithrombin III: effect of single amino acid substitutions and relationship with functional abnormalities.

    PubMed

    De Stefano, V; Leone, G; Mastrangelo, S; Lane, D A; Girolami, A; de Moerloose, P; Sas, G; Abildgaard, U; Blajchman, M; Rodeghiero, F

    1994-02-01

    Microheterogeneity of antithrombin III (AT-III) was investigated by crossed immunoelectrofocusing (CIEF) on eleven molecular variants. A normal pattern was found in five variants while two different abnormal CIEF patterns were found in the other four and two variants, respectively. Point mutations causing a major pI change (exceeding 4.0) of the amino acid substituted lead to alterations in the overall microheterogeneity. The variants thus substituted share a first type of abnormal CIEF pattern with alterations throughout the pH range, regardless of the location of the mutation (reactive site and adjacent regions or heparin binding region). Minor amino acid pI changes in these regions do not alter the AT-III overall microheterogeneity, whatever the resulting functional defect. However, if the mutation is placed in the region around positions 404 or 429, then even minor changes of the amino acid pI seem able to alter the overall charge, leading to a second type of abnormal CIEF pattern with the main alteration at pH 4.8-4.6. Neuraminidase treatment leads to disappearance of microheterogeneity except for the variants with the Arg393 to Cys substitution. Addition of thrombin induces CIEF modifications specifically related to the functional defect. A normal formation of thrombin-antithrombin complexes induces a shift towards the more acid pH range, whereas in the variants substituted at the reactive site the CIEF pattern is substantially unaffected by thrombin; variants substituted at positions 382-384 show a maximal thrombin-induced increase of the isoforms at pI 4.8-4.6. Therefore mutant antithrombins with different functional abnormalities but sharing a common CIEF pattern were well distinguished.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8180341

  12. Radiation chemistry of salicylic and methyl substituted salicylic acids: Models for the radiation chemistry of pharmaceutical compounds

    NASA Astrophysics Data System (ADS)

    Ayatollahi, Shakiba; Kalnina, Daina; Song, Weihua; Turks, Maris; Cooper, William J.

    2013-11-01

    Salicylic acid and its derivatives are components of many medications and moieties found in numerous pharmaceutical compounds. They have been used as models for various pharmaceutical compounds in pharmacological studies, for the treatment of pharmaceuticals and personal care products (PPCPs), and, reactions with natural organic matter (NOM). In this study, the radiation chemistry of benzoic acid, salicylic acid and four methyl substituted salicylic acids (MSA) is reported. The absolute bimolecular reaction rate constants for hydroxyl radical reaction with benzoic and salicylic acids as well as 3-methyl-, 4-methyl-, 5-methyl-, and 6-methyl-salicylic acid were determined (5.86±0.54)×109, (1.07±0.07)×1010, (7.48±0.17)×109, (7.31±0.29)×109, (5.47±0.25)×109, (6.94±0.10)×109 (M-1 s-1), respectively. The hydrated electron reaction rate constants were measured (3.02±0.10)×109, (8.98±0.27)×109, (5.39±0.21)×109, (4.33±0.17)×109, (4.72±0.15)×109, (1.42±0.02)×109 (M-1 s-1), respectively. The transient absorption spectra for the six model compounds were examined and their role as model compounds for the radiation chemistry of pharmaceuticals investigated.

  13. Adipose reduction by beta,beta'-tetramethyl-substituted hexadecanedioic acid (MEDICA 16).

    PubMed

    Tzur, R; Smith, E; Bar-Tana, J

    1989-01-01

    Treatment of normal rats kept on a balanced laboratory chow diet with beta,beta'-tetramethyl-substituted hexadecanedioic acid (MEDICA 16) (Bar-Tana et al., 1985, J. Biol. Chem, 260, 8404-8410) resulted in an acute reduction in adiposity, which was already established during the first week of treatment and was sustained as long as the drug was administered. Adipose reduction consisted of 30-80 percent decrease in the perirenal, omental, epididymal, parametrial and subcutaneous fat with a concomitant 50 percent decrease in total body neutral lipid mass. The reduction in adiposity was accounted for by a respective decrease in the lipid content of individual adipocytes together with a transient or sustained decrease in the number of adipocytes of selected adipose tissues. The decrease in the lipid content resulted from (a) an extensive hypotriglyceridemia in MEDICA 16-treated rats; (b) inhibition of adipose lipogenesis by MEDICA 16; (c) increased sensitivity to catecholamines-. ACTH- and forskolin-induced lipolysis in MEDICA 16 adipocytes. Adipose reduction by MEDICA 16 was not compromised by a decrease in overall net caloric intake but was accompanied by a 40 percent increase in resting metabolic rate. PMID:2670791

  14. Omega-3 Fatty Acid Formulations in Cardiovascular Disease: Dietary Supplements are Not Substitutes for Prescription Products.

    PubMed

    Fialkow, Jonathan

    2016-08-01

    Omega-3 fatty acid products are available as prescription formulations (icosapent ethyl, omega-3-acid ethyl esters, omega-3-acid ethyl esters A, omega-3-carboxylic acids) and dietary supplements (predominantly fish oils). Most dietary supplements and all but one prescription formulation contain mixtures of the omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Products containing both EPA and DHA may raise low-density lipoprotein cholesterol (LDL-C). In clinical trials, the EPA-only prescription product, icosapent ethyl, did not raise LDL-C compared with placebo. To correct a common misconception, it is important to note that omega-3 fatty acid dietary supplements are not US FDA-approved over-the-counter drugs and are not required to demonstrate safety and efficacy prior to marketing. Conversely, prescription products are supported by extensive clinical safety and efficacy investigations required for FDA approval and have active and ongoing safety monitoring programs. While omega-3 fatty acid dietary supplements may have a place in the supplementation of diet, they generally contain lower levels of EPA and DHA than prescription products and are not approved or intended to treat disease. Perhaps due to the lack of regulation of dietary supplements, EPA and DHA levels may vary widely within and between brands, and products may also contain unwanted cholesterol or fats or potentially harmful components, including toxins and oxidized fatty acids. Accordingly, omega-3 fatty acid dietary supplements should not be substituted for prescription products. Similarly, prescription products containing DHA and EPA should not be substituted for the EPA-only prescription product, as DHA may raise LDL-C and thereby complicate the management of patients with dyslipidemia.

  15. Omega-3 Fatty Acid Formulations in Cardiovascular Disease: Dietary Supplements are Not Substitutes for Prescription Products.

    PubMed

    Fialkow, Jonathan

    2016-08-01

    Omega-3 fatty acid products are available as prescription formulations (icosapent ethyl, omega-3-acid ethyl esters, omega-3-acid ethyl esters A, omega-3-carboxylic acids) and dietary supplements (predominantly fish oils). Most dietary supplements and all but one prescription formulation contain mixtures of the omega-3 fatty acids eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Products containing both EPA and DHA may raise low-density lipoprotein cholesterol (LDL-C). In clinical trials, the EPA-only prescription product, icosapent ethyl, did not raise LDL-C compared with placebo. To correct a common misconception, it is important to note that omega-3 fatty acid dietary supplements are not US FDA-approved over-the-counter drugs and are not required to demonstrate safety and efficacy prior to marketing. Conversely, prescription products are supported by extensive clinical safety and efficacy investigations required for FDA approval and have active and ongoing safety monitoring programs. While omega-3 fatty acid dietary supplements may have a place in the supplementation of diet, they generally contain lower levels of EPA and DHA than prescription products and are not approved or intended to treat disease. Perhaps due to the lack of regulation of dietary supplements, EPA and DHA levels may vary widely within and between brands, and products may also contain unwanted cholesterol or fats or potentially harmful components, including toxins and oxidized fatty acids. Accordingly, omega-3 fatty acid dietary supplements should not be substituted for prescription products. Similarly, prescription products containing DHA and EPA should not be substituted for the EPA-only prescription product, as DHA may raise LDL-C and thereby complicate the management of patients with dyslipidemia. PMID:27138439

  16. Comment on "Transitions to asexuality result in excess amino acid substitutions".

    PubMed

    Butlin, Roger

    2006-09-01

    Paland and Lynch (Reports, 17 February 2006, p. 990) showed that in Daphnia pulex, the ratio of amino acid replacement to silent substitution in mitochondrial genes is higher in asexual lineages than in sexual lineages. If this base-composition bias is maintained by selection, it too should alter following transitions in reproductive mode. Analysis reveals no such change in the genomes of D. pulex.

  17. Massive difference in synonymous substitution rates among mitochondrial, plastid, and nuclear genes of Phaeocystis algae.

    PubMed

    Smith, David Roy; Arrigo, Kevin R; Alderkamp, Anne-Carlijn; Allen, Andrew E

    2014-02-01

    We are just beginning to understand how mutation rates differ among mitochondrial, plastid, and nuclear genomes. In most seed plants the mitochondrial mutation rate is estimated to be lower than those of the plastid and nucleus, whereas in the red alga Porphyra the opposite is true, and in certain green algae all three genomes appear to have similar rates of mutation. Relative rate statistics of organelle vs nuclear genes, however, are lacking for lineages that acquired their plastids through secondary endosymbiosis, but recent organelle DNA analyses suggest that they may differ drastically from what is observed in lineages with primary plastids, such as green plants and red algae. Here, by measuring synonymous nucleotide substitutions, we approximate the relative mutation rates within the haptophyte genus Phaeocystis, which has a red-algal-derived, secondary plastid. Synonymous-site divergence data indicate that for Phaeocystis antarctica and P. globosa the mitochondrial mutation rate is 10 and 3 times that of the plastid and nucleus, respectively. This differs drastically from relative rate estimates for primary-plastid-bearing lineages and presents a much more dynamic view of organelle vs nuclear mutation rates across the eukaryotic domain. PMID:24216019

  18. The causes of synonymous rate variation in the rodent genome. Can substitution rates be used to estimate the sex bias in mutation rate?

    PubMed Central

    Smith, N G; Hurst, L D

    1999-01-01

    Miyata et al. have suggested that the male-to-female mutation rate ratio (alpha) can be estimated by comparing the neutral substitution rates of X-linked (X), Y-linked (Y), and autosomal (A) genes. Rodent silent site X/A comparisons provide very different estimates from X/Y comparisons. We examine three explanations for this discrepancy: (1) statistical biases and artifacts, (2) nonneutral evolution, and (3) differences in mutation rate per germline replication. By estimating errors and using a variety of methodologies, we tentatively reject explanation 1. Our analyses of patterns of codon usage, synonymous rates, and nonsynonymous rates suggest that silent sites in rodents are evolving neutrally, and we can therefore reject explanation 2. We find both base composition and methylation differences between the different sets of chromosomes, a result consistent with explanation 3, but these differences do not appear to explain the observed discrepancies in estimates of alpha. Our finding of significantly low synonymous substitution rates in genomically imprinted genes suggests a link between hemizygous expression and an adaptive reduction in the mutation rate, which is consistent with explanation 3. Therefore our results provide circumstantial evidence in favor of the hypothesis that the discrepancies in estimates of alpha are due to differences in the mutation rate per germline replication between different parts of the genome. This explanation violates a critical assumption of the method of Miyata et al., and hence we suggest that estimates of alpha, obtained using this method, need to be treated with caution. PMID:10353908

  19. An amino acid substitution in the pyruvate dehydrogenase E1{alpha} gene, affecting mitochondrial import of the precursor protein

    SciTech Connect

    Takakubo, F.; Thorburn, D.R.; Dahl, H.H.M.

    1995-10-01

    A mutation in the mitochondrial targeting sequence was characterized in a male patient with X chromosome-linked pyruvate dehydrogenase E1{alpha} deficiency. The mutation was a base substitution of G by C at nucleotide 134 in the mitochondrial targeting sequence of the PDHA1 gene, resulting in an arginine-to-proline substitution at codon 10 (R10P). Pyruvate dehydrogenase activity in cultured skin fibroblasts was 28% of the control value, and immunoblot analysis revealed a decreased level of pyruvate dehydrogenase E1{alpha}immunoreactivity. Chimeric constructs in which the normal and mutant pyruvate dehydrogenase E1{alpha} targeting sequences were attached to the mitochondrial matrix protein ornithine transcarbamylase were synthesized in a cell free translation system, and mitochondrial import of normal and mutant proteins was compared in vitro. The results show that ornithine transcarbamylase targeted by the mutant pyruvate dehydrogenase E1{alpha} sequence was translocated into the mitochondrial matrix at a reduced rate, suggesting that defective import is responsible for the reduced pyruvate dehydrogenase level in mitochondria. The mutation was also present in an affected brother and the mildly affected mother. The clinical presentations of this X chromosome-linked disorder in affected family members are discussed. To our knowledge, this is the first report of an amino acid substitution in a mitochondrial targeting sequence resulting in a human genetic disease. 58 refs., 5 figs., 1 tab.

  20. Evaluating EDTA as a substitute for phosphoric acid-etching of enamel and dentin.

    PubMed

    Imbery, Terence A; Kennedy, Matthew; Janus, Charles; Moon, Peter C

    2012-01-01

    Matrix metalloproteinases (MMPs) are proteolytic enzymes released when dentin is acid-etched. The enzymes are capable of destroying unprotected collagen fibrils that are not encapsulated by the dentin adhesive. Chlorhexidine applied after etching inhibits the activation of released MMPs, whereas neutral ethylenediamine tetra-acetic acid (EDTA) prevents the release of MMPs. The purpose of this study was to determine if conditioning enamel and dentin with EDTA can be a substitute for treating acid-etching enamel and dentin with chlorhexidine. A column of composite resin was bonded to enamel and dentin after conditioning. Shear bond strengths were evaluated after 48 hours and after accelerated aging for three hours in 12% sodium hypochlorite. Shear bond strengths ranged from 15.6 MP a for accelerated aged EDTA enamel specimens to 26.8 MPa for dentin conditioned with EDTA and tested after 48 hours. A three-way ANOVA and a Tukey HSD test found statistically significant differences among the eight groups and the three independent variables (P < 0.05). EDTA was successfully substituted for phosphoric acid-etched enamel and dentin treated with chlorhexidine. Interactions of conditioning agent and aging were significant for dentin but not for enamel. In an effort to reduce the detrimental effects of MMPs, conditioning enamel and dentin with EDTA is an alternative to treating acid-etched dentin and enamel with chlorhexidine.

  1. Substitution rate calibration of small subunit ribosomal RNA identifies chlorarachniophyte endosymbionts as remnants of green algae.

    PubMed Central

    Van de Peer, Y; Rensing, S A; Maier, U G; De Wachter, R

    1996-01-01

    Chlorarachniophytes are amoeboid algae with chlorophyll a and b containing plastids that are surrounded by four membranes instead of two as in plants and green algae. These extra membranes form important support for the hypothesis that chlorarachniophytes have acquired their plastids by the ingestion of another eukaryotic plastid-containing alga. Chlorarachniophytes also contain a small nucleus-like structure called the nucleomorph situated between the two inner and the two outer membranes surrounding the plastid. This nucleomorph is a remnant of the endosymbiont's nucleus and encodes, among other molecules, small subunit ribosomal RNA. Previous phylogenetic analyses on the basis of this molecule provided unexpected and contradictory evidence for the origin of the chlorarachniophyte endosymbiont. We developed a new method for measuring the substitution rates of the individual nucleotides of small subunit ribosomal RNA. From the resulting substitution rate distribution, we derived an equation that gives a more realistic relationship between sequence dissimilarity and evolutionary distance than equations previously available. Phylogenetic trees constructed on the basis of evolutionary distances computed by this new method clearly situate the chlorarachniophyte nucleomorphs among the green algae. Moreover, this relationship is confirmed by transversion analysis of the Chlorarachnion plastid small subunit ribosomal RNA. PMID:8755544

  2. Vapor Phase Dehydration of Glycerol to Acrolein Over SBA-15 Supported Vanadium Substituted Phosphomolybdic Acid Catalyst.

    PubMed

    Viswanadham, Balaga; Srikanth, Amirineni; Kumar, Vanama Pavan; Chary, Komandur V R

    2015-07-01

    Vapor phase dehydration of glycerol to acrolein was investigated over heteropolyacid (HPA) catalysts containing vanadium substituted phosphomolybdic acid (H4PMo11VO40) supported on mesoporous SBA-15. A series of HPA catalysts with HPA loadings varying from 10-50 wt% were prepared by impregnation method on SBA-15 support. The catalysts were characterized by X-ray diffraction, Raman spectroscopy, Fourier Transform infrared spectroscopy, temperature-programmed desorption of NH3, pyridine adsorbed FT-IR spectroscopy, scanning electron microscopy, pore size distribution and specific surface area measurements. The nature of acidic sites was examined by pyridine adsorbed FT-IR spectroscopy. XRD results suggest that the active phase containing HPA was highly dispersed at lower loadings on the support. FT-IR and Raman spectra results confirm that the presence of primary Keggin ion structure of HPA on the support and it was not affected during the preparation of catalysts. Pore size distribution results reveal that all the samples show unimodel pore size distribution with well depicted mesoporous structure. NH3-TPD results suggest that the acidity of catalysts increased with increase of HPA loading. The findings of acidity measurements by FT-IR spectra of pyridine adsorption reveals that the catalysts consist both the Brønsted and Lewis acidic sites and the amount of Brønsted acidic sites are increasing with HPA loading. SBA-15 supported vanadium substituted phosphomolybdic acid catalysts are found to be highly active during the dehydration reaction and exhibited 100% conversion of glycerol (10 wt% of glycerol) and the acrolein selectivity was appreciably changed with HPA active phase loading. The catalytic functionalities during glycerol dehydration are well correlated with surface acidity of the catalysts.

  3. Simulation of the in vivo resorption rate of β-tricalcium phosphate bone graft substitutes implanted in a sheep model.

    PubMed

    Bashoor-Zadeh, Mahdieh; Baroud, Gamal; Bohner, Marc

    2011-09-01

    A few years ago, a model was proposed to predict the effect of the pore architecture of a bone graft substitute on its cell-mediated resorption rate. The aim of the present study was to compare the predictions of the model with the in vivo resorption rate of four β-tricalcium phosphate bone graft substitutes implanted in a sheep model. The simulation algorithm contained two main steps: (1) detection of the pores that could be accessed by blood vessels of 50 μm in diameter, and (2) removal of one solid layer at the surface of these pores. This process was repeated until full resorption occurred. Since the pore architecture was complex, μCT data and fuzzy imaging techniques were combined to reconstruct the precise bone graft substitute geometry and then image processing algorithms were developed to perform the resorption simulation steps. The proposed algorithm was verified by comparing its results with the analytical results of a simple geometry and experimental in-vivo data of β-TCP bone substitutes with more complex geometry. An excellent correlation (r(2)>0.9 for all 4 bone graft substitutes) was found between simulation results and in-vivo data, suggesting that this resorption model could be used to (i) better understand the in vivo behavior of bone graft substitutes resorbed by cell-mediation, and (ii) optimize the pore architecture of a bone graft substitute, for example to maximize its resorption rate.

  4. 40 CFR 721.10381 - Cyclic carboxylic acid, polymer with dihydroxy dialkyl ether, hydroxy substituted alkane and...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Cyclic carboxylic acid, polymer with dihydroxy dialkyl ether, hydroxy substituted alkane and carboxylic acid anhydride, methacrylate terminated... Specific Chemical Substances § 721.10381 Cyclic carboxylic acid, polymer with dihydroxy dialkyl...

  5. 40 CFR 721.10381 - Cyclic carboxylic acid, polymer with dihydroxy dialkyl ether, hydroxy substituted alkane and...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Cyclic carboxylic acid, polymer with dihydroxy dialkyl ether, hydroxy substituted alkane and carboxylic acid anhydride, methacrylate terminated... Specific Chemical Substances § 721.10381 Cyclic carboxylic acid, polymer with dihydroxy dialkyl...

  6. 40 CFR 721.10381 - Cyclic carboxylic acid, polymer with dihydroxy dialkyl ether, hydroxy substituted alkane and...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Cyclic carboxylic acid, polymer with dihydroxy dialkyl ether, hydroxy substituted alkane and carboxylic acid anhydride, methacrylate terminated... Specific Chemical Substances § 721.10381 Cyclic carboxylic acid, polymer with dihydroxy dialkyl...

  7. Antibody-specific model of amino acid substitution for immunological inferences from alignments of antibody sequences.

    PubMed

    Mirsky, Alexander; Kazandjian, Linda; Anisimova, Maria

    2015-03-01

    Antibodies are glycoproteins produced by the immune system as a dynamically adaptive line of defense against invading pathogens. Very elegant and specific mutational mechanisms allow B lymphocytes to produce a large and diversified repertoire of antibodies, which is modified and enhanced throughout all adulthood. One of these mechanisms is somatic hypermutation, which stochastically mutates nucleotides in the antibody genes, forming new sequences with different properties and, eventually, higher affinity and selectivity to the pathogenic target. As somatic hypermutation involves fast mutation of antibody sequences, this process can be described using a Markov substitution model of molecular evolution. Here, using large sets of antibody sequences from mice and humans, we infer an empirical amino acid substitution model AB, which is specific to antibody sequences. Compared with existing general amino acid models, we show that the AB model provides significantly better description for the somatic evolution of mice and human antibody sequences, as demonstrated on large next generation sequencing (NGS) antibody data. General amino acid models are reflective of conservation at the protein level due to functional constraints, with most frequent amino acids exchanges taking place between residues with the same or similar physicochemical properties. In contrast, within the variable part of antibody sequences we observed an elevated frequency of exchanges between amino acids with distinct physicochemical properties. This is indicative of a sui generis mutational mechanism, specific to antibody somatic hypermutation. We illustrate this property of antibody sequences by a comparative analysis of the network modularity implied by the AB model and general amino acid substitution models. We recommend using the new model for computational studies of antibody sequence maturation, including inference of alignments and phylogenetic trees describing antibody somatic hypermutation in

  8. Structural consequences of amino acid substitutions causing Tay-Sachs disease.

    PubMed

    Ohno, Kazuki; Saito, Seiji; Sugawara, Kanako; Sakuraba, Hitoshi

    2008-08-01

    To determine the structural changes in the alpha-subunit of beta-hexosaminidase due to amino acid substitutions causing Tay-Sachs disease, we built structural models of mutant alpha-subunits resulting from 33 missense mutations (24 infantile and 9 late-onset), and analyzed the influence of each amino acid replacement on the structure by calculating the number of atoms affected and determining the solvent-accessible surface area of the corresponding amino acid residue in the wild-type alpha-subunit. In the infantile Tay-Sachs group, the number of atoms influenced by a mutation was generally larger than that in the late-onset Tay-Sachs group in both the main chain and the side chain, and residues associated with the mutations found in the infantile Tay-Sachs group tended to be less solvent-accessible than those in the late-onset Tay-Sachs group. Furthermore, color imaging determined the distribution and degree of the structural changes caused by representative amino acid substitutions, and that there were also differences between the infantile and late-onset Tay-Sachs disease groups. Structural study is useful for elucidating the basis of Tay-Sachs disease.

  9. Structural consequences of amino acid substitutions causing Tay-Sachs disease.

    PubMed

    Ohno, Kazuki; Saito, Seiji; Sugawara, Kanako; Sakuraba, Hitoshi

    2008-08-01

    To determine the structural changes in the alpha-subunit of beta-hexosaminidase due to amino acid substitutions causing Tay-Sachs disease, we built structural models of mutant alpha-subunits resulting from 33 missense mutations (24 infantile and 9 late-onset), and analyzed the influence of each amino acid replacement on the structure by calculating the number of atoms affected and determining the solvent-accessible surface area of the corresponding amino acid residue in the wild-type alpha-subunit. In the infantile Tay-Sachs group, the number of atoms influenced by a mutation was generally larger than that in the late-onset Tay-Sachs group in both the main chain and the side chain, and residues associated with the mutations found in the infantile Tay-Sachs group tended to be less solvent-accessible than those in the late-onset Tay-Sachs group. Furthermore, color imaging determined the distribution and degree of the structural changes caused by representative amino acid substitutions, and that there were also differences between the infantile and late-onset Tay-Sachs disease groups. Structural study is useful for elucidating the basis of Tay-Sachs disease. PMID:18490185

  10. Spectroscopic determination of acid dissociation constants of N-substituted-6-acylbenzothiazolone derivatives.

    PubMed

    Sıdır, Yadigar Gülseven; Sıdır, Isa; Berber, Halil

    2011-05-26

    The acid dissociation constants of twelve novel drug precursor N-substituted-6-acylbenzothiazolone derivatives were determined by using the UV-vis spectroscopic technique. The protonation and deprotonation behaviors of the investigated molecules were researched from the super basic to super acid regions (i.e., 8 mol·L(-1) KOH to 98% H(2)SO(4)) including the pH region. It is observed that all of the molecules are protonated in the super acidic region. The calculated relative stability values of possible tautomer structures indicate that the keto form of investigated molecules is favored over the enol form. It was predicted that protonation occurs at the amide (oxo) group found in the keto form.

  11. The Synthesis and Characterization of Substituted Polyaniline Hollow Spheres doped with a Polymeric Acid

    NASA Astrophysics Data System (ADS)

    Sui, Jing; Zhang, Lijuan; Travas-Sejdic, Jadranka; Kilmartin, Paul A.

    2009-07-01

    Self-assembled poly(o-toluidine) (POT) and poly(o-anisidine) (POA) hollow spheres were prepared by oxidative polymerization using ammonium persulfate as the oxidant in the presence of 5% by weight of a polymeric acid, poly(methyl vinyl ether-alt-maleic acid) (PMVEA). The substituent at the ortho position had a significant effect on the size of the hollow nanospheres as determined by SEM and TEM. The nanospheres were of a very uniform size in the presence of the polymeric acid, with average diameters of 338±25 nm for POT and 210±20 nm for POA. The POT and POA hollow spheres were also characterized by FTIR and UV-Vis spectroscopy, which confirmed the chemical identity of the substituted polyanilines.

  12. Aggregation of dialkyl-substituted diphosphonic acids and its effect on metal ion extraction.

    SciTech Connect

    Chiarizia, R.; Barrans, R. E., Jr.; Ferraro, J. R. Herlinger, A. W.; McAlister, D. R.

    1999-10-22

    Solvent extraction reagents containing the diphosphonic acid group exhibit an extraordinary affinity for tri-, tetra- and hexavalent actinides. Their use has been considered for actinide separation and pre-concentration procedures. Solvent extraction data obtained with P,P{prime}-di(2-ethylhexyl) methane-, ethane- and butanediphosphonic acids exhibit features that are difficult to explain without Knowledge of the aggregation state of the extractants. Information about the aggregation of the dialkyl-substituted diphosphonic acids in aromatic diluents has been obtained using the complementary techniques of vapor pressure osmometry (VPO), small angle neutron scattering (SANS), infrared spectroscopy and molecular mechanics. The results from these techniques provide an understanding of the aggregation behavior of these extractants that is fully compatible with the solvent extraction data. The most important results and their relevance to solvent extraction are reviewed in this paper.

  13. Disentangling the perturbational effects of amino acid substitutions in the DNA-binding domain of p53.

    PubMed

    Wacey, A I; Cooper, D N; Liney, D; Hovig, E; Krawczak, M

    1999-01-01

    The spectrum of somatic cancer-associated missense mutations in the human TP53 gene was studied in order to assess the potential structural and functional importance of various intra-molecular properties associated with these substitutions. Relating the observed frequency of particular amino acid substitutions in the p53 DNA-binding domain to their expected frequency, as calculated from DNA sequence-dependent mutation rates, yielded estimates of their relative clinical observation likelihood (RCOL). Several biophysical properties were found to display significant covariation with RCOL values. Thus RCOL values were observed to decrease with increasing solvent accessibility of the substituted residue and with increasing distance from the p53 DNA-binding and Zn2+ -binding sites. The number of adverse steric interactions introduced by an amino acid replacement was found to be positively correlated with its RCOL value, irrespective of the magnitude of the interactions. A gain in hydrogen bond number was found to be only half as likely to come to clinical attention as mutations involving either a reduction or no change in hydrogen bond number. When the difference in potential energy between the wild-type and mutant DNA-binding domains was considered, RCOL values exhibited a minimum around changes of zero. Finally, classification of mutated residues in terms of their protein/solvent environment yielded, for somatic p53 mutations, RCOL values that resembled those previously determined for inherited mutations of human factor IX causing haemophilia B, suggesting that similar mechanisms may be responsible for the mutation-related perturbation of biological function in different protein folds.

  14. Accounting for variation of substitution rates through time in Bayesian phylogeny reconstruction of Sapotoideae (Sapotaceae).

    PubMed

    Smedmark, Jenny E E; Swenson, Ulf; Anderberg, Arne A

    2006-06-01

    We used Bayesian phylogenetic analysis of 5 kb of chloroplast DNA data from 68 Sapotaceae species to clarify phylogenetic relationships within Sapotoideae, one of the two major clades within Sapotaceae. Variation in substitution rates through time was shown to be a very important aspect of molecular evolution for this data set. Relative rates tests indicated that changes in overall rate have taken place in several lineages during the history of the group and Bayes factors strongly supported a covarion model, which allows the rate of a site to vary over time, over commonly used models that only allow rates to vary across sites. Rate variation over time was actually found to be a more important model component than rate variation across sites. The covarion model was originally developed for coding gene sequences and has so far only been tested for this type of data. The fact that it performed so well with the present data set, consisting mainly of data from noncoding spacer regions, suggests that it deserves a wider consideration in model based phylogenetic inference. Repeatability of phylogenetic results was very difficult to obtain with the more parameter rich models, and analyses with identical settings often supported different topologies. Overparameterization may be the reason why the MCMC did not sample from the posterior distribution in these cases. The problem could, however, be overcome by using less parameter rich evolutionary models, and adjusting the MCMC settings. The phylogenetic results showed that two taxa, previously thought to belong in Sapotoideae, are not part of this group. Eberhardtia aurata is the sister of the two major Sapotaceae clades, Chrysophylloideae and Sapotoideae, and Neohemsleya usambarensis belongs in Chrysophylloideae. Within Sapotoideae two clades, Sideroxyleae and Sapoteae, were strongly supported. Bayesian analysis of the character history of some floral morphological traits showed that the ancestral type of flower in

  15. Decarboxylation of substituted cinnamic acids by lactic acid bacteria isolated during malt whisky fermentation.

    PubMed

    van Beek, S; Priest, F G

    2000-12-01

    Seven strains of Lactobacillus isolated from malt whisky fermentations and representing Lactobacillus brevis, L. crispatus, L. fermentum, L. hilgardii, L. paracasei, L. pentosus, and L. plantarum contained genes for hydroxycinnamic acid (p-coumaric acid) decarboxylase. With the exception of L. hilgardii, these bacteria decarboxylated p-coumaric acid and/or ferulic acid, with the production of 4-vinylphenol and/or 4-vinylguaiacol, respectively, although the relative activities on the two substrates varied between strains. The addition of p-coumaric acid or ferulic acid to cultures of L. pentosus in MRS broth induced hydroxycinnamic acid decarboxylase mRNA within 5 min, and the gene was also induced by the indigenous components of malt wort. In a simulated distillery fermentation, a mixed culture of L. crispatus and L. pentosus in the presence of Saccharomyces cerevisiae decarboxylated added p-coumaric acid more rapidly than the yeast alone but had little activity on added ferulic acid. Moreover, we were able to demonstrate the induction of hydroxycinnamic acid decarboxylase mRNA under these conditions. However, in fermentations with no additional hydroxycinnamic acid, the bacteria lowered the final concentration of 4-vinylphenol in the fermented wort compared to the level seen in a pure-yeast fermentation. It seems likely that the combined activities of bacteria and yeast decarboxylate p-coumaric acid and then reduce 4-vinylphenol to 4-ethylphenol more effectively than either microorganism alone in pure cultures. Although we have shown that lactobacilli participate in the metabolism of phenolic compounds during malt whisky fermentations, the net result is a reduction in the concentrations of 4-vinylphenol and 4-vinylguaiacol prior to distillation.

  16. Structure-Activity Relationship Studies of Amino Acid Substitutions in Radiolabeled Neurotensin Conjugates.

    PubMed

    Mascarin, Alba; Valverde, Ibai E; Mindt, Thomas L

    2016-01-01

    Radiolabeled derivatives of the peptide neurotensin (NT) and its binding sequence NT(8-13) have been studied as potential imaging probes and therapeutics for NT-1-receptor-positive cancer. However, a direct comparison of reported NT analogues, even if radiolabeled with the same radionuclide, is difficult because different techniques and models have been used for preclinical evaluations. In an effort to identify a suitable derivative of NT(8-13) for radiotracer development, we herein report a side-by-side in vitro comparison of radiometallated NT derivatives bearing some of the most commonly reported amino acid substitutions in their sequence. Performed investigations include cell internalization experiments, determinations of receptor affinity, measurements of the distribution coefficient, and blood serum stability studies. Of the [(177)Lu]-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-labeled examples studied, analogues of NT(8-13) containing a short hydrophilic tetraethylene glycol (PEG4 ) spacer between the peptide and the radiometal complex, and a minimum number of substitutions of amino acid residues, exhibited the most promising properties in vitro. PMID:26593062

  17. Structure-Activity Relationship Studies of Amino Acid Substitutions in Radiolabeled Neurotensin Conjugates.

    PubMed

    Mascarin, Alba; Valverde, Ibai E; Mindt, Thomas L

    2016-01-01

    Radiolabeled derivatives of the peptide neurotensin (NT) and its binding sequence NT(8-13) have been studied as potential imaging probes and therapeutics for NT-1-receptor-positive cancer. However, a direct comparison of reported NT analogues, even if radiolabeled with the same radionuclide, is difficult because different techniques and models have been used for preclinical evaluations. In an effort to identify a suitable derivative of NT(8-13) for radiotracer development, we herein report a side-by-side in vitro comparison of radiometallated NT derivatives bearing some of the most commonly reported amino acid substitutions in their sequence. Performed investigations include cell internalization experiments, determinations of receptor affinity, measurements of the distribution coefficient, and blood serum stability studies. Of the [(177)Lu]-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA)-labeled examples studied, analogues of NT(8-13) containing a short hydrophilic tetraethylene glycol (PEG4 ) spacer between the peptide and the radiometal complex, and a minimum number of substitutions of amino acid residues, exhibited the most promising properties in vitro.

  18. Amino Acid Substitutions in Cold-Adapted Proteins from Halorubrum lacusprofundi, an Extremely Halophilic Microbe from Antarctica

    PubMed Central

    DasSarma, Shiladitya; Capes, Melinda D.; Karan, Ram; DasSarma, Priya

    2013-01-01

    The halophilic Archaeon Halorubrum lacusprofundi, isolated from the perennially cold and hypersaline Deep Lake in Antarctica, was recently sequenced and compared to 12 Haloarchaea from temperate climates by comparative genomics. Amino acid substitutions for 604 H. lacusprofundi proteins belonging to conserved haloarchaeal orthologous groups (cHOGs) were determined and found to occur at 7.85% of positions invariant in proteins from mesophilic Haloarchaea. The following substitutions were observed most frequently: (a) glutamic acid with aspartic acid or alanine; (b) small polar residues with other small polar or non-polar amino acids; (c) small non-polar residues with other small non-polar residues; (d) aromatic residues, especially tryptophan, with other aromatic residues; and (e) some larger polar residues with other similar residues. Amino acid substitutions for a cold-active H. lacusprofundi β-galactosidase were then examined in the context of a homology modeled structure at residues invariant in homologous enzymes from mesophilic Haloarchaea. Similar substitutions were observed as in the genome-wide approach, with the surface accessible regions of β-galactosidase displaying reduced acidity and increased hydrophobicity, and internal regions displaying mainly subtle changes among smaller non-polar and polar residues. These findings are consistent with H. lacusprofundi proteins displaying amino acid substitutions that increase structural flexibility and protein function at low temperature. We discuss the likely mechanisms of protein adaptation to a cold, hypersaline environment on Earth, with possible relevance to life elsewhere. PMID:23536799

  19. Reversal of the surface charge asymmetry in purple membrane due to single amino acid substitutions.

    PubMed Central

    Hsu, K C; Rayfield, G W; Needleman, R

    1996-01-01

    Twenty-seven mutant bacteriorhodopsin's were screened to determine the PKa for reversal of the permanent electric dipole moment. The photoelectric response of an aqueous purple-membrane suspension was used to determine the direction of the purple-membrane dipole moment as a function of pH. The pK(a) for the dipole reversal of wild-type bacteriorhodopsin is 4.5. Six of the 27 mutant bacteriorhodopsin's were found to have a pK(a) for dipole reversal larger than that of wild-type bacteriorhodopsin. Two of these mutants, L93T and L93W, involve a neutral amino acid substitution in the interior of the protein. The direction of the purple-membrane permanent electric dipole moment is determined by the purple-membrane surface charge asymmetry. We conclude that these two substitutions, which do not involve charge replacement, alter the pK(a) for the reversal of the purple-membrane surface charge asymmetry. We suggest that these changes to the pK(a) are due to altered protein folding at the surface of the purple-membrane induced by single-site substitutions in the protein interior. PMID:9172760

  20. Fourier transform infrared spectroscopic imaging parameters describing acid phosphate substitution in biologic hydroxyapatite.

    PubMed

    Spevak, Lyudmila; Flach, Carol R; Hunter, Tracey; Mendelsohn, Richard; Boskey, Adele

    2013-05-01

    Acid phosphate substitution into mineralized tissues is an important determinant of their mechanical properties and their response to treatment. This study identifies and validates Fourier transform infrared spectroscopic imaging (FTIRI) spectral parameters that provide information on the acid phosphate (HPO4) substitution into hydroxyapatite in developing mineralized tissues. Curve fitting and Fourier self-deconvolution were used to identify subband positions in model compounds (with and without HPO4). The intensity of subbands at 1127 and 1110 cm(-1) correlated with the acid phosphate content in these models. Peak height ratios of these subbands to the ν3 vibration at 1096 cm(-1) found in stoichiometric apatite were evaluated in the model compounds and mixtures thereof. FTIRI spectra of bones and teeth at different developmental ages were analyzed using these spectral parameters. Factor analysis (a chemometric technique) was also conducted on the tissue samples and resulted in factor loadings with spectral features corresponding to the HPO4 vibrations described above. Images of both factor correlation coefficients and the peak height ratios 1127/1096 and 1112/1096 cm(-1) demonstrated higher acid phosphate content in younger vs. more mature regions in the same specimen. Maps of the distribution of acid phosphate content will be useful for characterizing the extent of new bone formation, the areas of potential decreased strength, and the effects of therapies such as those used in metabolic bone diseases (osteoporosis, chronic kidney disease) on mineral composition. Because of the wider range of values obtained with the 1127/1096 cm(-1) parameter compared to the 1110/1096 cm(-1) parameter and the smaller scatter in the slope, it is suggested that this ratio should be the parameter of choice.

  1. Evolutionary dynamics of the plastid inverted repeat: the effects of expansion, contraction, and loss on substitution rates.

    PubMed

    Zhu, Andan; Guo, Wenhu; Gupta, Sakshi; Fan, Weishu; Mower, Jeffrey P

    2016-03-01

    Rates of nucleotide substitution were previously shown to be several times slower in the plastid inverted repeat (IR) compared with single-copy (SC) regions, suggesting that the IR provides enhanced copy-correction activity. To examine the generality of this synonymous rate dependence on the IR, we compared plastomes from 69 pairs of closely related species representing 52 families of angiosperms, gymnosperms, and ferns. We explored the breadth of IR boundary shifts in land plants and demonstrate that synonymous substitution rates are, on average, 3.7 times slower in IR genes than in SC genes. In addition, genes moved from the SC into the IR exhibit lower synonymous rates consistent with other IR genes, while genes moved from the IR into the SC exhibit higher rates consistent with other SC genes. Surprisingly, however, several plastid genes from Pelargonium, Plantago, and Silene have highly accelerated synonymous rates despite their IR localization. Together, these results provide strong evidence that the duplicative nature of the IR reduces the substitution rate within this region. The anomalously fast-evolving genes in Pelargonium, Plantago, and Silene indicate localized hypermutation, potentially induced by a higher level of error-prone double-strand break repair in these regions, which generates substitutional rate variation. PMID:26574731

  2. Identification and biophysical characterization of a very-long-chain-fatty-acid-substituted phosphatidylinositol in yeast subcellular membranes

    PubMed Central

    2004-01-01

    Morphological analysis of a conditional yeast mutant in acetyl-CoA carboxylase acc1ts/mtr7, the rate-limiting enzyme of fatty acid synthesis, suggested that the synthesis of C26 VLCFAs (very-long-chain fatty acids) is important for maintaining the structure and function of the nuclear membrane. To characterize this C26-dependent pathway in more detail, we have now examined cells that are blocked in pathways that require C26. In yeast, ceramide synthesis and remodelling of GPI (glycosylphosphatidylinositol)-anchors are two pathways that incorporate C26 into lipids. Conditional mutants blocked in either ceramide synthesis or the synthesis of GPI anchors do not display the characteristic alterations of the nuclear envelope observed in acc1ts, indicating that the synthesis of another C26-containing lipid may be affected in acc1ts mutant cells. Lipid analysis of isolated nuclear membranes revealed the presence of a novel C26-substituted PI (phosphatidylinositol). This C26-PI accounts for approx. 1% of all the PI species, and is present in both the nuclear and the plasma membrane. Remarkably, this C26-PI is the only C26-containing glycerophospholipid that is detectable in wild-type yeast, and the C26-substitution is highly specific for the sn-1 position of the glycerol backbone. To characterize the biophysical properties of this lipid, it was chemically synthesized. In contrast to PIs with normal long-chain fatty acids (C16 or C18), the C26-PI greatly reduced the bilayer to hexagonal phase transition of liposomes composed of 1,2-dielaidoyl-sn-glycero-3-phosphoethanolamine (DEPE). The biophysical properties of this lipid are thus consistent with a possible role in stabilizing highly curved membrane domains. PMID:15270698

  3. 40 CFR 721.9220 - Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Reaction products of secondary alkyl... Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric acid... substances identified generically as reaction products of secondary alkyl amines with a...

  4. 40 CFR 721.9220 - Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Reaction products of secondary alkyl... Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric acid... substances identified generically as reaction products of secondary alkyl amines with a...

  5. 40 CFR 721.9220 - Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Reaction products of secondary alkyl... Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric acid... substances identified generically as reaction products of secondary alkyl amines with a...

  6. 40 CFR 721.9220 - Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Reaction products of secondary alkyl... Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric acid... substances identified generically as reaction products of secondary alkyl amines with a...

  7. 40 CFR 721.9220 - Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Reaction products of secondary alkyl... Reaction products of secondary alkyl amines with a substituted benzenesulfonic acid and sulfuric acid... substances identified generically as reaction products of secondary alkyl amines with a...

  8. Effect of ring substitution on the metabolic fate and anti-inflammatory activity of some prodolic acid analogs.

    PubMed

    Robinson, W T; Martel, R R; Ferdinandi, E; Kraml, M

    1977-12-01

    Prodolic acid, 1-n-propyl-1,3,4,9-tetrahydropyrano[3,4-b]indole-1-acetic acid, exhibits potent anti-inflammatory activity in adjuvant arthritic rats. The potency of prodolic acid is enhanced by indole ring substitution. This increase correlated well with higher and sustained drug concentrations in the serum of normal animals. Pharmacokinetic studies demonstrated that ring substitution prolonged the serum half-life without affecting the absorption or volume of distribution. Because, in the rat, indole ring hydroxylation is a major pathway for the disposition of prodolic acid, we ascribe the increased pharmacological activity of ring substituted derivatives to the interference of substituents with the hydroxylation reaction. PMID:925966

  9. Comparative mitochondrial genomics of snakes: extraordinary substitution rate dynamics and functionality of the duplicate control region

    PubMed Central

    Jiang, Zhi J; Castoe, Todd A; Austin, Christopher C; Burbrink, Frank T; Herron, Matthew D; McGuire, Jimmy A; Parkinson, Christopher L; Pollock, David D

    2007-01-01

    Background The mitochondrial genomes of snakes are characterized by an overall evolutionary rate that appears to be one of the most accelerated among vertebrates. They also possess other unusual features, including short tRNAs and other genes, and a duplicated control region that has been stably maintained since it originated more than 70 million years ago. Here, we provide a detailed analysis of evolutionary dynamics in snake mitochondrial genomes to better understand the basis of these extreme characteristics, and to explore the relationship between mitochondrial genome molecular evolution, genome architecture, and molecular function. We sequenced complete mitochondrial genomes from Slowinski's corn snake (Pantherophis slowinskii) and two cottonmouths (Agkistrodon piscivorus) to complement previously existing mitochondrial genomes, and to provide an improved comparative view of how genome architecture affects molecular evolution at contrasting levels of divergence. Results We present a Bayesian genetic approach that suggests that the duplicated control region can function as an additional origin of heavy strand replication. The two control regions also appear to have different intra-specific versus inter-specific evolutionary dynamics that may be associated with complex modes of concerted evolution. We find that different genomic regions have experienced substantial accelerated evolution along early branches in snakes, with different genes having experienced dramatic accelerations along specific branches. Some of these accelerations appear to coincide with, or subsequent to, the shortening of various mitochondrial genes and the duplication of the control region and flanking tRNAs. Conclusion Fluctuations in the strength and pattern of selection during snake evolution have had widely varying gene-specific effects on substitution rates, and these rate accelerations may have been functionally related to unusual changes in genomic architecture. The among-lineage and

  10. Physicochemical and acid gelation properties of commercial UHT-treated plant-based milk substitutes and lactose free bovine milk.

    PubMed

    Mäkinen, Outi E; Uniacke-Lowe, Thérèse; O'Mahony, James A; Arendt, Elke K

    2015-02-01

    Physicochemical and acid gelation properties of UHT-treated commercial soy, oat, quinoa, rice and lactose-free bovine milks were studied. The separation profiles were determined using a LUMiSizer dispersion analyser. Soy, rice and quinoa milks formed both cream and sediment layers, while oat milk sedimented but did not cream. Bovine milk was very stable to separation while all plant milks separated at varying rates; rice and oat milks being the most unstable products. Particle sizes in plant-based milk substitutes, expressed as volume mean diameters (d4.3), ranged from 0.55μm (soy) to 2.08μm (quinoa) while the average size in bovine milk was 0.52μm. Particles of plant-based milk substitutes were significantly more polydisperse compared to those of bovine milk. Upon acidification with glucono-δ-lactone (GDL), bovine, soy and quinoa milks formed structured gels with maximum storage moduli of 262, 187 and 105Pa, respectively while oat and rice milks did not gel. In addition to soy products currently on the market, quinoa may have potential in dairy-type food applications. PMID:25172757

  11. Calcite crystal growth rate inhibition by polycarboxylic acids

    USGS Publications Warehouse

    Reddy, M.M.; Hoch, A.R.

    2001-01-01

    Calcite crystal growth rates measured in the presence of several polycarboxyclic acids show that tetrahydrofurantetracarboxylic acid (THFTCA) and cyclopentanetetracarboxylic acid (CPTCA) are effective growth rate inhibitors at low solution concentrations (0.01 to 1 mg/L). In contrast, linear polycarbocylic acids (citric acid and tricarballylic acid) had no inhibiting effect on calcite growth rates at concentrations up to 10 mg/L. Calcite crystal growth rate inhibition by cyclic polycarboxyclic acids appears to involve blockage of crystal growth sites on the mineral surface by several carboxylate groups. Growth morphology varied for growth in the absence and in the presence of both THFTCA and CPTCA. More effective growth rate reduction by CPTCA relative to THFTCA suggests that inhibitor carboxylate stereochemical orientation controls calcite surface interaction with carboxylate inhibitors. ?? 20O1 Academic Press.

  12. Efficient continuous-flow synthesis of novel 1,2,3-triazole-substituted β-aminocyclohexanecarboxylic acid derivatives with gram-scale production

    PubMed Central

    Ötvös, Sándor B; Georgiádes, Ádám; Mándity, István M; Kiss, Lóránd

    2013-01-01

    Summary The preparation of novel multi-substituted 1,2,3-triazole-modified β-aminocyclohexanecarboxylic acid derivatives in a simple and efficient continuous-flow procedure is reported. The 1,3-dipolar cycloaddition reactions were performed with copper powder as a readily accessible Cu(I) source. Initially, high reaction rates were achieved under high-pressure/high-temperature conditions. Subsequently, the reaction temperature was lowered to room temperature by the joint use of both basic and acidic additives to improve the safety of the synthesis, as azides were to be handled as unstable reactants. Scale-up experiments were also performed, which led to the achievement of gram-scale production in a safe and straightforward way. The obtained 1,2,3-triazole-substituted β-aminocyclohexanecarboxylates can be regarded as interesting precursors for drugs with possible biological effects. PMID:23946850

  13. Evolution of moth sex pheromone composition by a single amino acid substitution in a fatty acid desaturase.

    PubMed

    Buček, Aleš; Matoušková, Petra; Vogel, Heiko; Šebesta, Petr; Jahn, Ullrich; Weißflog, Jerrit; Svatoš, Aleš; Pichová, Iva

    2015-10-13

    For sexual communication, moths primarily use blends of fatty acid derivatives containing one or more double bonds in various positions and configurations, called sex pheromones (SPs). To study the molecular basis of novel SP component (SPC) acquisition, we used the tobacco hornworm (Manduca sexta), which uses a blend of mono-, di-, and uncommon triunsaturated fatty acid (3UFA) derivatives as SP. We identified pheromone-biosynthetic fatty acid desaturases (FADs) MsexD3, MsexD5, and MsexD6 abundantly expressed in the M. sexta female pheromone gland. Their functional characterization and in vivo application of FAD substrates indicated that MsexD3 and MsexD5 biosynthesize 3UFAs via E/Z14 desaturation from diunsaturated fatty acids produced by previously characterized Z11-desaturase/conjugase MsexD2. Site-directed mutagenesis of sequentially highly similar MsexD3 and MsexD2 demonstrated that swapping of a single amino acid in the fatty acyl substrate binding tunnel introduces E/Z14-desaturase specificity to mutated MsexD2. Reconstruction of FAD gene phylogeny indicates that MsexD3 was recruited for biosynthesis of 3UFA SPCs in M. sexta lineage via gene duplication and neofunctionalization, whereas MsexD5 representing an alternative 3UFA-producing FAD has been acquired via activation of a presumably inactive ancestral MsexD5. Our results demonstrate that a change as small as a single amino acid substitution in a FAD enzyme might result in the acquisition of new SP compounds. PMID:26417103

  14. Evolution of moth sex pheromone composition by a single amino acid substitution in a fatty acid desaturase

    PubMed Central

    Buček, Aleš; Matoušková, Petra; Vogel, Heiko; Šebesta, Petr; Jahn, Ullrich; Weißflog, Jerrit; Svatoš, Aleš; Pichová, Iva

    2015-01-01

    For sexual communication, moths primarily use blends of fatty acid derivatives containing one or more double bonds in various positions and configurations, called sex pheromones (SPs). To study the molecular basis of novel SP component (SPC) acquisition, we used the tobacco hornworm (Manduca sexta), which uses a blend of mono-, di-, and uncommon triunsaturated fatty acid (3UFA) derivatives as SP. We identified pheromone-biosynthetic fatty acid desaturases (FADs) MsexD3, MsexD5, and MsexD6 abundantly expressed in the M. sexta female pheromone gland. Their functional characterization and in vivo application of FAD substrates indicated that MsexD3 and MsexD5 biosynthesize 3UFAs via E/Z14 desaturation from diunsaturated fatty acids produced by previously characterized Z11-desaturase/conjugase MsexD2. Site-directed mutagenesis of sequentially highly similar MsexD3 and MsexD2 demonstrated that swapping of a single amino acid in the fatty acyl substrate binding tunnel introduces E/Z14-desaturase specificity to mutated MsexD2. Reconstruction of FAD gene phylogeny indicates that MsexD3 was recruited for biosynthesis of 3UFA SPCs in M. sexta lineage via gene duplication and neofunctionalization, whereas MsexD5 representing an alternative 3UFA-producing FAD has been acquired via activation of a presumably inactive ancestral MsexD5. Our results demonstrate that a change as small as a single amino acid substitution in a FAD enzyme might result in the acquisition of new SP compounds. PMID:26417103

  15. Effects of ion substitution on bile acid-dependent and -independent bile formation by rat liver.

    PubMed Central

    Van Dyke, R W; Stephens, J E; Scharschmidt, B F

    1982-01-01

    To characterize the transport mechanisms responsible for formation of canalicular bile, we have examined the effects of ion substitution on bile acid-dependent and bile acid-independent bile formation by the isolated perfused rat liver. Complete replacement of perfusate sodium with choline and lithium abolished taurocholate-induced choleresis and reduced biliary taurocholate output by greater than 70%. Partial replacement of perfusate sodium (25 of 128 mM) by choline reduced bile acid-independent bile formation by 30% and replacement of the remaining sodium (103 mM) by choline reduced bile acid-independent bile formation by an additional 64%. In contrast, replacement of the remaining sodium (103 mM) by lithium reduced bile acid-independent bile formation by only an additional 20%, while complete replacement of sodium (128 mM) by lithium reduced bile formation by only 17%, and lithium replaced sodium as the predominant biliary cation. Replacement of perfusate bicarbonate by Tricine, a zwitterionic amino acid buffer, decreased bile acid-independent bile formation by greater than or equal to 50% and decreased biliary bicarbonate output by approximately 60%, regardless of the accompanying cation. In separate experiments, replacement of sodium by lithium essentially abolished Na,K-ATPase activity measured either as ouabain-suppressible ATP hydrolysis in rat liver or kidney homogenates, or as ouabain-suppressible 86Rb uptake by cultured rat hepatocytes. These studies indicate that bile acid(taurocholate)-dependent bile formation by rat liver exhibits a specific requirement for sodium, a finding probably attributable to the role(s) of sodium in hepatic sodium-coupled taurocholate uptake and/or in maintenance of Na,K-ATPase activity. The surprising finding that bile acid-independent bile formation was substantially unaltered by complete replacement of sodium with the permeant cation lithium does not appear to be explained by Na,K-ATPase-mediated lithium transport. Although

  16. Anisotropic scattering rate in Fe-substituted Bi2Sr2Ca(Cu1-xFex)2O8+δ

    DOE PAGES

    Naamneh, M.; Lubashevsky, Y.; Lahoud, E.; Gu, G.; Kanigel, A.

    2015-05-27

    We measured the electronic structure of Fe substituted Bi2212 using Angle Resolved Photoemission Spectroscopy (ARPES). We find that the substitution does not change the momentum dependence of the superconducting gap but induces a very anisotropic enhancement of the scattering rate. A comparison of the effect of Fe substitution to that of Zn substitution suggests that the Fe reduces Tc so effectively because it supresses very strongly the coherence weight around the anti-nodes.

  17. An amino acid substitution (L925V) associated with resistance to pyrethroids in Varroa destructor.

    PubMed

    González-Cabrera, Joel; Davies, T G Emyr; Field, Linda M; Kennedy, Peter J; Williamson, Martin S

    2013-01-01

    The Varroa mite, Varroa destructor, is an important pest of honeybees and has played a prominent role in the decline in bee colony numbers over recent years. Although pyrethroids such as tau-fluvalinate and flumethrin can be highly effective in removing the mites from hives, their intensive use has led to many reports of resistance. To investigate the mechanism of resistance in UK Varroa samples, the transmembrane domain regions of the V. destructor voltage-gated sodium channel (the main target site for pyrethroids) were PCR amplified and sequenced from pyrethroid treated/untreated mites collected at several locations in Central/Southern England. A novel amino acid substitution, L925V, was identified that maps to a known hot spot for resistance within the domain IIS5 helix of the channel protein; a region that has also been proposed to form part of the pyrethroid binding site. Using a high throughput diagnostic assay capable of detecting the mutation in individual mites, the L925V substitution was found to correlate well with resistance, being present in all mites that had survived tau-fluvalinate treatment but in only 8 % of control, untreated samples. The potential for using this assay to detect and manage resistance in Varroa-infected hives is discussed.

  18. An amino acid substitution (Gly853-->Glu) in the collagen alpha 1(II) chain produces hypochondrogenesis.

    PubMed

    Bogaert, R; Tiller, G E; Weis, M A; Gruber, H E; Rimoin, D L; Cohn, D H; Eyre, D R

    1992-11-01

    The spondyloepiphyseal dysplasia subclassification of bone dysplasias includes achondrogenesis, hypochondrogenesis, and spondyloepiphyseal dysplasia congenita. The phenotypic expression of these disorders ranges from mild to perinatal lethal forms. We report the detection and partial characterization of a defect in type II collagen in a perinatal lethal form of hypochondrogenesis. Electrophoresis in sodium dodecyl sulfate-polyacrylamide of CB peptides (where CB represents cyanogen bromide) from type II collagen of the diseased cartilage showed a doublet band for peptide alpha 1(II)CB10 and evidence for post-translational overmodification of the major peptides (CB8, CB10, and CB11) seen as a retarded electrophoretic mobility. Peptide CB10 was digested by endoproteinase Asp-N; and on reverse-phase high pressure liquid chromatography, fragments of abnormal mobility were noted. Sequence analysis of a unique peptide D12 revealed a single amino acid substitution (Gly-->Glu) at position 853 of the triple helical domain. This was confirmed by sequence analysis of amplified COL2A1 cDNA, which revealed a single nucleotide substitution (GGA-->GAA) in 5 of 10 clones. Electron micrographs of the diseased cartilage showed a sparse extracellular matrix and chondrocytes containing dilated rough endoplasmic reticulum, which suggested impaired assembly and secretion of the mutant protein. This case further documents the molecular basis of the spondyloepiphyseal dysplasia spectrum of chondrodysplasias as mutations in COL2A1. PMID:1429602

  19. Slow and Fast Evolving Endosymbiont Lineages: Positive Correlation between the Rates of Synonymous and Non-Synonymous Substitution

    PubMed Central

    Silva, Francisco J.; Santos-Garcia, Diego

    2015-01-01

    The availability of complete genome sequences of bacterial endosymbionts with strict vertical transmission to the host progeny opens the possibility to estimate molecular evolutionary rates in different lineages and understand the main biological mechanisms influencing these rates. We have compared the rates of evolution for non-synonymous and synonymous substitutions in nine bacterial endosymbiont lineages, belonging to four clades (Baumannia, Blochmannia, Portiera, and Sulcia). The main results are the observation of a positive correlation between both rates with differences among lineages of up to three orders of magnitude and that the substitution rates decrease over long endosymbioses. To explain these results we propose three mechanisms. The first, variations in the efficiencies of DNA replication and DNA repair systems, is unable to explain most of the observed differences. The second, variations in the generation time among bacterial lineages, would be based on the accumulation of fewer DNA replication errors per unit time in organisms with longer generation times. The third, a potential control of the endosymbiont DNA replication and repair systems through the transfer of nuclear-encoded proteins, could explain the lower rates in long-term obligate endosymbionts. Because the preservation of the genomic integrity of the harbored obligate endosymbiont would be advantageous for the insect host, biological mechanisms producing a general reduction in the rates of nucleotide substitution per unit of time would be a target for natural selection. PMID:26617602

  20. Cu(II) /TEMPO-promoted one-pot synthesis of highly substituted pyrimidines from amino acid esters.

    PubMed

    Zhou, Nini; Xie, Tao; Li, Zhongle; Xie, Zhixiang

    2014-12-22

    A novel, Cu(OAc)2/TEMPO promoted one-step approach for the preparation of fully substituted pyrimidines from readily available amino acid esters has been described. In this reaction, the amino acid esters act as the only N-C sources for the construction of corresponding pyrimidines. The mechanism of this process includes oxidative dehydrogenation, the generation of an imine radical, and a formal [3+3] cycloaddition. This methodology proves to be a high atom-economic and straightforward strategy for the synthesis of pyrimidines and diverse substrates which are substituted by various functional groups have been afforded in moderate to good yield. PMID:25377658

  1. Structural and catalytic effects of an invariant purine substitution in the hammerhead ribozyme: implications for the mechanism of acid-base catalysis.

    PubMed

    Schultz, Eric P; Vasquez, Ernesto E; Scott, William G

    2014-09-01

    The hammerhead ribozyme catalyzes RNA cleavage via acid-base catalysis. Whether it does so by general acid-base catalysis, in which the RNA itself donates and abstracts protons in the transition state, as is typically assumed, or by specific acid-base catalysis, in which the RNA plays a structural role and proton transfer is mediated by active-site water molecules, is unknown. Previous biochemical and crystallographic experiments implicate an invariant purine in the active site, G12, as the general base. However, G12 may play a structural role consistent with specific base catalysis. To better understand the role of G12 in the mechanism of hammerhead catalysis, a 2.2 Å resolution crystal structure of a hammerhead ribozyme from Schistosoma mansoni with a purine substituted for G12 in the active site of the ribozyme was obtained. Comparison of this structure (PDB entry 3zd4), in which A12 is substituted for G, with three previously determined structures that now serve as important experimental controls, allows the identification of structural perturbations that are owing to the purine substitution itself. Kinetic measurements for G12 purine-substituted schistosomal hammerheads confirm a previously observed dependence of rate on the pK(a) of the substituted purine; in both cases inosine, which is similar to G in pK(a) and hydrogen-bonding properties, is unexpectedly inactive. Structural comparisons indicate that this may primarily be owing to the lack of the exocyclic 2-amino group in the G12A and G12I substitutions and its structural effect upon both the nucleotide base and phosphate of A9. The latter involves the perturbation of a previously identified and well characterized metal ion-binding site known to be catalytically important in both minimal and full-length hammerhead ribozyme sequences. The results permit it to be suggested that G12 plays an important role in stabilizing the active-site structure. This result, although not inconsistent with the potential

  2. Disproportional plastome-wide increase of substitution rates and relaxed purifying selection in genes of carnivorous Lentibulariaceae.

    PubMed

    Wicke, Susann; Schäferhoff, Bastian; dePamphilis, Claude W; Müller, Kai F

    2014-03-01

    Carnivorous Lentibulariaceae exhibit the most sophisticated implementation of the carnivorous syndrome in plants. Their unusual lifestyle coincides with distinct genomic peculiarities such as the smallest angiosperm nuclear genomes and extremely high nucleotide substitution rates across all genomic compartments. Here, we report the complete plastid genomes from each of the three genera Pinguicula, Utricularia, and Genlisea, and investigate plastome-wide changes in their molecular evolution as the carnivorous syndrome unfolds. We observe a size reduction by up to 9% mostly due to the independent loss of genes for the plastid NAD(P)H dehydrogenase and altered proportions of plastid repeat DNA, as well as a significant plastome-wide increase of substitution rates and microstructural changes. Protein-coding genes across all gene classes show a disproportional elevation of nonsynonymous substitutions, particularly in Utricularia and Genlisea. Significant relaxation of purifying selection relative to noncarnivores occurs in the plastid-encoded fraction of the photosynthesis ATP synthase complex, the photosystem I, and in several other photosynthesis and metabolic genes. Shifts in selective regimes also affect housekeeping genes including the plastid-encoded polymerase, for which evidence for relaxed purifying selection was found once during the transition to carnivory, and a second time during the diversification of the family. Lentibulariaceae significantly exhibit enhanced rates of nucleotide substitution in most of the 130 noncoding regions. Various factors may underlie the observed patterns of relaxation of purifying selection and substitution rate increases, such as reduced net photosynthesis rates, alternative paths of nutrient uptake (including organic carbon), and impaired DNA repair mechanisms.

  3. Disproportional plastome-wide increase of substitution rates and relaxed purifying selection in genes of carnivorous Lentibulariaceae.

    PubMed

    Wicke, Susann; Schäferhoff, Bastian; dePamphilis, Claude W; Müller, Kai F

    2014-03-01

    Carnivorous Lentibulariaceae exhibit the most sophisticated implementation of the carnivorous syndrome in plants. Their unusual lifestyle coincides with distinct genomic peculiarities such as the smallest angiosperm nuclear genomes and extremely high nucleotide substitution rates across all genomic compartments. Here, we report the complete plastid genomes from each of the three genera Pinguicula, Utricularia, and Genlisea, and investigate plastome-wide changes in their molecular evolution as the carnivorous syndrome unfolds. We observe a size reduction by up to 9% mostly due to the independent loss of genes for the plastid NAD(P)H dehydrogenase and altered proportions of plastid repeat DNA, as well as a significant plastome-wide increase of substitution rates and microstructural changes. Protein-coding genes across all gene classes show a disproportional elevation of nonsynonymous substitutions, particularly in Utricularia and Genlisea. Significant relaxation of purifying selection relative to noncarnivores occurs in the plastid-encoded fraction of the photosynthesis ATP synthase complex, the photosystem I, and in several other photosynthesis and metabolic genes. Shifts in selective regimes also affect housekeeping genes including the plastid-encoded polymerase, for which evidence for relaxed purifying selection was found once during the transition to carnivory, and a second time during the diversification of the family. Lentibulariaceae significantly exhibit enhanced rates of nucleotide substitution in most of the 130 noncoding regions. Various factors may underlie the observed patterns of relaxation of purifying selection and substitution rate increases, such as reduced net photosynthesis rates, alternative paths of nutrient uptake (including organic carbon), and impaired DNA repair mechanisms. PMID:24344209

  4. Acid mine drainage simulated leaching behavior of goethite and cobalt substituted goethite

    NASA Astrophysics Data System (ADS)

    Penprase, S. B.; Kimball, B. E.

    2015-12-01

    Though most modern day mining aims to eliminate the seepage of acid mine drainage (AMD) to the local watershed, historical mines regularly receive little to no remediation, and often release acidic, metal-rich drainage and particles to the environment. Treatment of AMD often includes neutralizing pH to facilitate the precipitation of Fe-oxides and dissolved trace metals, thereby forming Trace Metal Substituted (TMS) forms of known minerals, such as goethite (α-FeOOH). The stability of TMS precipitates is not fully understood. As a result, we conducted a 20 day leach experiment using laboratory synthesized pure (Gt) and cobalt-substituted (CoGt) goethites with a dilute ultrapure HCl solution (pH = 3.61) at T = 23.3±2.5ºC. Leached solids were characterized using X-ray diffraction (XRD) and scanning electron microscopy paired with energy dispersive spectroscopy (SEM-EDS). Leach solutions were sampled for pH and conductivity, and dissolved chemistry was determined with Inductively Coupled Plasma Mass Spectroscopy (ICP-MS). Preliminary results indicate Gt and CoGt filtered leach solutions experienced constant pH (Gt = 3.9 ± 0.1, CoGt = 6.8 ± 0.2) and conductivity (Gt = 69 ± 6.6 μS/cm, CoGt = 81 ± 16 μS/cm) for t = 0-20 days. Micro-focused XRD results indicate that leached solids did not change in mineralogy throughout the experiment, and SEM images show minor disintegration along mineral grain edges, but little overall change in shape. Preliminary ICP-MS results show lower dissolved Fe concentrations for CoGt (1.1 ± 1.1 ppb) compared to Gt (17 ± 8.9 ppb) over time. Dissolved Co concentrations ranged from 560 - 830 ppb and increased over time. Compared to leaching of pure Gt, leaching of CoGt generated significantly higher pH, slightly higher conductivity, and significantly less dissolved Fe. During the CoGt leach, Co was preferentially leached over Fe. The differences in leaching behavior between pure and TMS goethite in the laboratory have implications for

  5. Mutational Biases Drive Elevated Rates of Substitution at Regulatory Sites across Cancer Types

    PubMed Central

    Semple, Colin A.

    2016-01-01

    Disruption of gene regulation is known to play major roles in carcinogenesis and tumour progression. Here, we comprehensively characterize the mutational profiles of diverse transcription factor binding sites (TFBSs) across 1,574 completely sequenced cancer genomes encompassing 11 tumour types. We assess the relative rates and impact of the mutational burden at the binding sites of 81 transcription factors (TFs), by comparing the abundance and patterns of single base substitutions within putatively functional binding sites to control sites with matched sequence composition. There is a strong (1.43-fold) and significant excess of mutations at functional binding sites across TFs, and the mutations that accumulate in cancers are typically more disruptive than variants tolerated in extant human populations at the same sites. CTCF binding sites suffer an exceptionally high mutational load in cancer (3.31-fold excess) relative to control sites, and we demonstrate for the first time that this effect is seen in essentially all cancer types with sufficient data. The sub-set of CTCF sites involved in higher order chromatin structures has the highest mutational burden, suggesting a widespread breakdown of chromatin organization. However, we find no evidence for selection driving these distinctive patterns of mutation. The mutational load at CTCF-binding sites is substantially determined by replication timing and the mutational signature of the tumor in question, suggesting that selectively neutral processes underlie the unusual mutation patterns. Pervasive hyper-mutation within transcription factor binding sites rewires the regulatory landscape of the cancer genome, but it is dominated by mutational processes rather than selection. PMID:27490693

  6. Mutational Biases Drive Elevated Rates of Substitution at Regulatory Sites across Cancer Types.

    PubMed

    Kaiser, Vera B; Taylor, Martin S; Semple, Colin A

    2016-08-01

    Disruption of gene regulation is known to play major roles in carcinogenesis and tumour progression. Here, we comprehensively characterize the mutational profiles of diverse transcription factor binding sites (TFBSs) across 1,574 completely sequenced cancer genomes encompassing 11 tumour types. We assess the relative rates and impact of the mutational burden at the binding sites of 81 transcription factors (TFs), by comparing the abundance and patterns of single base substitutions within putatively functional binding sites to control sites with matched sequence composition. There is a strong (1.43-fold) and significant excess of mutations at functional binding sites across TFs, and the mutations that accumulate in cancers are typically more disruptive than variants tolerated in extant human populations at the same sites. CTCF binding sites suffer an exceptionally high mutational load in cancer (3.31-fold excess) relative to control sites, and we demonstrate for the first time that this effect is seen in essentially all cancer types with sufficient data. The sub-set of CTCF sites involved in higher order chromatin structures has the highest mutational burden, suggesting a widespread breakdown of chromatin organization. However, we find no evidence for selection driving these distinctive patterns of mutation. The mutational load at CTCF-binding sites is substantially determined by replication timing and the mutational signature of the tumor in question, suggesting that selectively neutral processes underlie the unusual mutation patterns. Pervasive hyper-mutation within transcription factor binding sites rewires the regulatory landscape of the cancer genome, but it is dominated by mutational processes rather than selection. PMID:27490693

  7. Identification of Amino Acid Substitutions Supporting Antigenic Change of Influenza A(H1N1)pdm09 Viruses

    PubMed Central

    Koel, Björn F.; Mögling, Ramona; Chutinimitkul, Salin; Fraaij, Pieter L.; Burke, David F.; van der Vliet, Stefan; de Wit, Emmie; Bestebroer, Theo M.; Rimmelzwaan, Guus F.; Osterhaus, Albert D. M. E.; Smith, Derek J.; Fouchier, Ron A. M.

    2015-01-01

    ABSTRACT The majority of currently circulating influenza A(H1N1) viruses are antigenically similar to the virus that caused the 2009 influenza pandemic. However, antigenic variants are expected to emerge as population immunity increases. Amino acid substitutions in the hemagglutinin protein can result in escape from neutralizing antibodies, affect viral fitness, and change receptor preference. In this study, we constructed mutants with substitutions in the hemagglutinin of A/Netherlands/602/09 in an attenuated backbone to explore amino acid changes that may contribute to emergence of antigenic variants in the human population. Our analysis revealed that single substitutions affecting the loop that consists of amino acid positions 151 to 159 located adjacent to the receptor binding site caused escape from ferret and human antibodies elicited after primary A(H1N1)pdm09 virus infection. The majority of these substitutions resulted in similar or increased replication efficiency in vitro compared to that of the virus carrying the wild-type hemagglutinin and did not result in a change of receptor preference. However, none of the substitutions was sufficient for escape from the antibodies in sera from individuals that experienced both seasonal and pandemic A(H1N1) virus infections. These results suggest that antibodies directed against epitopes on seasonal A(H1N1) viruses contribute to neutralization of A(H1N1)pdm09 antigenic variants, thereby limiting the number of possible substitutions that could lead to escape from population immunity. IMPORTANCE Influenza A viruses can cause significant morbidity and mortality in humans. Amino acid substitutions in the hemagglutinin protein can result in escape from antibody-mediated neutralization. This allows the virus to reinfect individuals that have acquired immunity to previously circulating strains through infection or vaccination. To date, the vast majority of A(H1N1)pdm09 strains remain antigenically similar to the virus

  8. Amino-acid substitution in alpha-spectrin commonly coinherited with nondominant hereditary spherocytosis.

    PubMed

    Tse, W T; Gallagher, P G; Jenkins, P B; Wang, Y; Benoit, L; Speicher, D; Winkelmann, J C; Agre, P; Forget, B G; Marchesi, S L

    1997-03-01

    Nondominant hereditary spherocytosis (ndHS) is a disorder characterized in some patients by severe hemolytic anemia and marked deficiency of erythrocyte spectrin. This report describes the identification of a variant spectrin chain, alpha-spectrin Bughill or alpha(BH), that is associated with this disorder in a number of patients. Tryptic maps of spectrin from affected individuals revealed an acidic shift in isoelectric point of the alphaII domain peptides at 46 kD and 35 kD. A point mutation at codon 970 of the alpha-spectrin gene (GCT-->GAT), that changes the encoded amino acid from an alanine to an aspartic acid, was identified in genomic DNA of affected patients. The alpha(BH) variant was present in 8 patients with ndHS from five different kindreds but was absent in 4 patients from two other kindreds. The 8 ndHS patients with the alpha(BH) variant appeared to be homozygous for the alpha(BH) variant by analysis of peptide maps of limited tryptic digests of erythrocyte spectrin. However, following genomic DNA analysis, only 2 of these patients were true homozygotes, whereas 6 were found to be doubly heterozygous for the alpha(BH) allele and a second, presumably abnormal, alpha-spectrin gene. These results suggest that, in these 6 patients, the second alpha-spectrin allele is in fact associated with one or more genetic defect(s), causing decreased accumulation of alpha-spectrin. The pattern of transmission of the alpha(BH) allele in certain families suggests that the alpha(BH) amino-acid substitution is not itself responsible for ndHS but is more likely a polymorphic variant that, in some but not all cases, is in linkage disequilibrium with another uncharacterized alpha-spectrin gene defect that itself is a cause of ndHS. PMID:9067503

  9. Intelligibility of laryngectomees' substitute speech: automatic speech recognition and subjective rating.

    PubMed

    Schuster, Maria; Haderlein, Tino; Nöth, Elmar; Lohscheller, Jörg; Eysholdt, Ulrich; Rosanowski, Frank

    2006-02-01

    Substitute speech after laryngectomy is characterized by restricted aero-acoustic properties in comparison with laryngeal speech and has therefore lower intelligibility. Until now, an objective means to determine and quantify the intelligibility has not existed, although the intelligibility can serve as a global outcome parameter of voice restoration after laryngectomy. An automatic speech recognition system was applied on recordings of a standard text read by 18 German male laryngectomees with tracheoesophageal substitute speech. The system was trained with normal laryngeal speakers and not adapted to severely disturbed voices. Substitute speech was compared to laryngeal speech of a control group. Subjective evaluation of intelligibility was performed by a panel of five experts and compared to automatic speech evaluation. Substitute speech showed lower syllables/s and lower word accuracy than laryngeal speech. Automatic speech recognition for substitute speech yielded word accuracy between 10.0 and 50% (28.7+/-12.1%) with sufficient discrimination. It complied with experts' subjective evaluations of intelligibility. The multi-rater kappa of the experts alone did not differ from the multi-rater kappa of experts and the recognizer. Automatic speech recognition serves as a good means to objectify and quantify global speech outcome of laryngectomees. For clinical use, the speech recognition system will be adapted to disturbed voices and can also be applied in other languages. PMID:16001246

  10. Acidic Dicationic Iron(II) Dihydrogen Complexes and Compounds Related by H(2) Substitution.

    PubMed

    Landau, Shaun E.; Morris, Robert H.; Lough, Alan J.

    1999-12-27

    [trans-Fe(H(2))(CO)(dppe)(2)](2+) (3) (dppe = 1,2-bis(diphenylphosphino)ethane) was generated by protonation of [trans-FeH(CO)(dppe)(2)](+) in CD(2)Cl(2). [trans-Fe(H(2))(CO)(depe)(2)](2+) (6) (depe = 1,2-bis(diethylphosphino)ethane) was generated by the treatment of [trans-FeCl(CO)(depe)(2)](+) in CD(2)Cl(2) with AgSbF(6) under 1 atm of H(2). Complex 3 is more acidic than trifluoromethanesulfonic acid (HOTf) in CD(2)Cl(2), while 6 is suspected to be less acidic than [Et(2)OH](+). 3[OTf](2) is stable to H(2) loss under reduced pressure for several hours, an indication of strong three-center (Fe-H(2)), two-electron sigma-bonding. Both complexes 3 and 6 undergo H(2) substitution reactions. There is evidence of the formation of [trans-Fe(H(2)O)(CO)(dppe)(2)](2+) and [trans-Fe(OTf)(CO)(dppe)(2)](+), although these complexes could not be isolated. [trans-FeY(CO)(depe)(2)]Y complexes (Y(-) = [BF(4)](-), 7[BF(4)]; Y(-) = [OTf](-), 8[OTf]) were isolated from the corresponding reactions of [trans-FeH(CO)(depe)(2)]Y with [Et(2)OH][BF(4)] or HOTf. 7[BF(4)] was structurally characterized by single-crystal X-ray diffraction. Attempts to grow crystals of 8[OTf] yielded salts containing the complex [trans-Fe(H(2)O)(CO)(depe)(2)](2+) (9), which were structurally characterized by single-crystal X-ray diffraction. Coordination of [BF(4)](-) in 7[BF(4)] was demonstrated, by variable-temperature (31)P{(1)H} NMR spectroscopy, to be dynamic. Dissolving 7[BF(4)] in methanol results in nucleophilic substitution at B to yield the new complex [trans-FeF(CO)(depe)(2)](+) (10). An attempt to grow crystals of 10[BF(4)] from the reaction mixture resulted in crystals of [H(2)(depe)][BF(4)], which were structurally characterized by single-crystal X-ray diffraction.

  11. A unique tumor antigen produced by a single amino acid substitution.

    PubMed

    Monach, P A; Meredith, S C; Siegel, C T; Schreiber, H

    1995-01-01

    Mice immunized against a cancer recognize antigens unique to that cancer, but the molecular structures of such antigens are unknown. We isolated CD4+ T cell clones recognizing an antigen uniquely expressed on the UV-induced tumor 6132A; some clones inhibited the growth of tumors bearing the specific antigen. A T cell hybridoma was used to purify this antigen from nuclear extracts by RP-HPLC and SDS-PAGE using T cell immunoblot assays. A partial amino acid sequence was nearly identical to a sequence in ribosomal protein L9. The cDNA sequence of L9 from 6132A PRO cells differed from the normal sequence at one nucleotide; this mutation encoded histidine instead of leucine at position 47. A synthetic peptide containing this mutation was over 1000-fold more stimulatory of T cells than was the wild-type peptide. These results indicate that this unique tumor antigen is derived from a single amino acid substitution in a cellular protein.

  12. Role of a single amino acid substitution of VP3 H142D for increased acid resistance of foot-and-mouth disease virus serotype A.

    PubMed

    Biswal, Jitendra K; Das, Biswajit; Sharma, Gaurav K; Khulape, Sagar A; Pattnaik, Bramhadev

    2016-04-01

    Foot-and-mouth disease virus (FMDV) particles lose infectivity due to their dissociation into pentamers at pH value below 6.5. After the uptake of FMDV by receptor-mediated endocytosis, the acid-dependent dissociation process is required for the release of FMDV genome inside endosomes. Nevertheless, dissociation of FMDV particles in mildly acidic conditions renders the inactivated FMD vaccine less effective. To improve the acid stability of inactivated FMD vaccine during the manufacturing process, a serotype A IND 40/2000 (in-use vaccine strain) mutant with increased resistance to acid inactivation was generated through reverse genetics approach. Based upon the earlier reports, the crucial amino acid residue, H142 of VP3 capsid protein was substituted separately to various amino acid residues Arg (R), Phe (F), Ala (A), and Asp (D) on the full-genome length cDNA clone. While the H142 → R or H142 → F or H142 → A substitutions resulted in non-infectious FMDV, H142 → D mutation on VP3 protein (H3142D) resulted in the generation of mutant virus with enhanced resistance to acid-induced inactivation. In addition, H3142D substitution did not alter the replication ability and antigenicity of mutant as compared to the parental virus. However, the virus competition experiments revealed that the H3142D substitution conferred a loss of fitness for the mutant virus. Results from this study demonstrate that the H3142D substitution is the molecular determinant of acid-resistant phenotype in FMDV serotype A. PMID:26873406

  13. Orthogonal selectivity with cinnamic acids in 3-substituted benzofuran synthesis through C-H olefination of phenols.

    PubMed

    Agasti, Soumitra; Sharma, Upendra; Naveen, Togati; Maiti, Debabrata

    2015-03-28

    A palladium catalyzed intermolecular annulation of cinnamic acids and phenols has been achieved for the selective synthesis of 3-substituted benzofurans. Isotope labeling, competition experiments, kinetic studies, and intermediate trapping have supported a sequence of C-C bond formation and decarboxylation followed by the C-O cyclization pathway.

  14. Many amino acid substitution variants identified in DNA repair genes during human population screenings are predicted to impact protein function

    SciTech Connect

    Xi, T; Jones, I M; Mohrenweiser, H W

    2003-11-03

    Over 520 different amino acid substitution variants have been previously identified in the systematic screening of 91 human DNA repair genes for sequence variation. Two algorithms were employed to predict the impact of these amino acid substitutions on protein activity. Sorting Intolerant From Tolerant (SIFT) classified 226 of 508 variants (44%) as ''Intolerant''. Polymorphism Phenotyping (PolyPhen) classed 165 of 489 amino acid substitutions (34%) as ''Probably or Possibly Damaging''. Another 9-15% of the variants were classed as ''Potentially Intolerant or Damaging''. The results from the two algorithms are highly associated, with concordance in predicted impact observed for {approx}62% of the variants. Twenty one to thirty one percent of the variant proteins are predicted to exhibit reduced activity by both algorithms. These variants occur at slightly lower individual allele frequency than do the variants classified as ''Tolerant'' or ''Benign''. Both algorithms correctly predicted the impact of 26 functionally characterized amino acid substitutions in the APE1 protein on biochemical activity, with one exception. It is concluded that a substantial fraction of the missense variants observed in the general human population are functionally relevant. These variants are expected to be the molecular genetic and biochemical basis for the associations of reduced DNA repair capacity phenotypes with elevated cancer risk.

  15. Synthesis of sterically encumbered C10-arylated benzo[h]quinolines using ortho-substituted aryl boronic acids.

    PubMed

    Weimar, Marko; Fuchter, Matthew J

    2013-01-01

    The challenging coupling of 10-halobenzo[h]quinolines with ortho-substituted aryl boronic acids has been achieved using Pd(OAc)(2)/P(O)Ph(3) as the catalytic system. High yields were obtained for diversely functionalised substrates under mild reaction conditions. PMID:23069777

  16. [Cu(OH)(TMEDA)]₂Cl₂-catalyzed regioselective 2-arylation of 5-substituted tetrazoles with boronic acids under mild conditions.

    PubMed

    Onaka, Takuya; Umemoto, Hideaki; Miki, Yasuyoshi; Nakamura, Akira; Maegawa, Tomohiro

    2014-07-18

    A mild and regioselective 2-arylation of 5-substituted tetrazoles is described. The reaction proceeds regioselectively with a variety of arylboronic acids in the presence of [Cu(OH)(TMEDA)]2Cl2 to afford 2,5-disubstituted tetrazoles. This is the first report of highly regioselective arylation of 5-alkyltetrazoles. PMID:24962401

  17. Highly regio- and diastereoselective synthesis of CF3-substituted lactones via photoredox-catalyzed carbolactonization of alkenoic acids.

    PubMed

    Yasu, Yusuke; Arai, Yusuke; Tomita, Ren; Koike, Takashi; Akita, Munetaka

    2014-02-01

    Trifluoromethylative lactonization of both terminal and internal alkenoic acids by photoredox catalysis has been developed. The use of a Ru photocatalyst and Umemoto's reagent as a CF3 source is key in the present carbolactonization. This is the first example of a highly endo- and diastereoselective synthesis of CF3-substituted five-, six-, and seven-membered ring lactones from internal alkenoic acids. PMID:24422891

  18. Allele-Selective Inhibition of Huntingtin and Ataxin-3 Expression by RNA Duplexes Containing Unlocked Nucleic Acid (UNA) Substitutions

    PubMed Central

    Aiba, Yuichiro; Hu, Jiaxin; Liu, Jing; Xiang, Qin; Martinez, Carlos; Corey, David R.

    2014-01-01

    Unlocked nucleic acid (UNA) is an acyclic analog of RNA that can be introduced into RNA or DNA oligonucleotides. The increased flexibility conferred by the acyclic structure fundamentally affects the strength of base-pairing, createing opportunities for improved applications and new insights into molecular recognition. Here we test how UNA substitutions affect allele-selective inhibition of trinucleotide-repeat genes Huntingtin (HTT) and Ataxin-3 (ATX-3) expression. We find that the either the combination of mismatched bases and UNA substitutions or UNA substitutions alone can improve potency and selectivity. Inhibition is potent and selectivities of > 40-fold for inhibiting mutant versus wild-type expression can be achieved. Surprisingly, even though UNA preserves the potential for complete base-pairing, the introduction of UNA substitutions at central positions within fully complementary duplexes leads to >19-fold selectivity. Like mismatched bases, the introduction of central UNA bases disrupts the potential for cleavage of substrate by Argonaute 2 (AGO2) during gene silencing. UNA-substituted duplexes are as effective as other strategies for allele-selective silencing of trinucleotide repeat disease genes. Modulation of AGO2 activity by the introduction of UNA substitutions demonstrates that backbone flexibility is as important as base-pairing for catalysis of fully complementary duplex substrates. UNA can be used to tailor RNA silencing for optimal properties and allele-selective action. PMID:24266403

  19. The fundamental theorem of neutral evolution: rates of substitution and mutation should factor in premeiotic clusters.

    PubMed

    Woodruff, R C; Thomson, J N

    2005-11-01

    Mutations do not always arise as single events. Many new mutations actually occur in the cell lineage before germ cell formation or meiosis and are therefore replicated pre-meiotically. The increased likelihood of substitutions caused by these clusters of new mutant alleles can change the fundamental theorem of neutral evolution.

  20. Improved Synthesis of 5-Substituted 1H-Tetrazoles via the [3+2] Cycloaddition of Nitriles and Sodium Azide Catalyzed by Silica Sulfuric Acid

    PubMed Central

    Du, Zhenting; Si, Changmei; Li, Youqiang; Wang, Yin; Lu, Jing

    2012-01-01

    A silica supported sulfuric acid catalyzed [3+2] cycloaddition of nitriles and sodium azide to form 5-substituted 1H-tetrazoles is described. The protocol can provide a series of 5-substituted 1H-tetrazoles using silica sulfuric acid from nitriles and sodium azide in DMF in 72%–95% yield. PMID:22606004

  1. Vapor pressures of substituted polycarboxylic acids are much lower than previously reported

    NASA Astrophysics Data System (ADS)

    Huisman, A. J.; Krieger, U. K.; Zuend, A.; Marcolli, C.; Peter, T.

    2013-07-01

    The partitioning of compounds between the aerosol and gas phase is a primary focus in the study of the formation and fate of secondary organic aerosol. We present measurements of the vapor pressure of 2-methylmalonic (isosuccinic) acid, 2-hydroxymalonic (tartronic) acid, 2-methylglutaric acid, 3-hydroxy-3-carboxy-glutaric (citric) acid and DL-2,3-dihydroxysuccinic (DL-tartaric) acid, which were obtained from the evaporation rate of supersaturated liquid particles levitated in an electrodynamic balance. Our measurements indicate that the pure component liquid vapor pressures at 298.15 K for tartronic, citric and tartaric acids are much lower than the same quantity that was derived from solid state measurements in the only other room temperature measurement of these materials (made by Booth et al., 2010). This strongly suggests that empirical correction terms in a recent vapor pressure estimation model to account for the inexplicably high vapor pressures of these and similar compounds should be revisited, and that due caution should be used when the estimated vapor pressures of these and similar compounds are used as inputs for other studies.

  2. Relative Reaction Rates of Sulfamic Acid and Hydroxylamine with Nitric Acid

    SciTech Connect

    Karraker, D.G.

    2001-03-28

    This report describes a study of comparative reaction rates where the reductant is in excess, as in the 1B bank in the Purex process. The results of this work apply to planned plant tests to partially substitute HAN for the ferrous sulfamate reductant in the Purex 1B bank.

  3. Whole-cell biocatalytic production of variously substituted β-aryl- and β-heteroaryl-β-amino acids.

    PubMed

    Ratnayake, Nishanka Dilini; Theisen, Chelsea; Walter, Tyler; Walker, Kevin D

    2016-01-10

    Biologically-active β-peptides and pharmaceuticals that contain key β-amino acids are emerging as target therapeutics; thus, synthetic strategies to make substituted, enantiopure β-amino acids are increasing. Here, we use whole-cell Escherichia coli (OD600 ∼ 35) engineered to express a Pantoea agglomerans phenylalanine aminomutase (PaPAM) biocatalyst. In either 5 mL, 100mL, or 1L of M9 minimal medium containing α-phenylalanine (20mM), the cells produced ∼ 1.4 mg mL(-1) of β-phenylalanine in each volume. Representative pilot-scale 5-mL cultures, fermentation reactions converted 18 variously substituted α-arylalanines to their (S)-β-aryl-β-amino acids in vivo and were not toxic to cells at mid- to late-stage growth. The β-aryl-β-amino acids made ranged from 0.043 mg (p-nitro-β-phenylalanine, 4% converted yield) to 1.2mg (m-bromo-β-phenylalanine, 96% converted yield) over 6h in 5 mL. The substituted β-amino acids made herein can be used in redox and Stille-coupling reactions to make synthetic building blocks, or as bioisosteres in drug design.

  4. ESTIMATION OF CARBOXYLIC ACID ESTER HYDROLYSIS RATE CONSTANTS

    EPA Science Inventory

    SPARC chemical reactivity models were extended to calculate hydrolysis rate constants for carboxylic acid esters from molecular structure. The energy differences between the initial state and the transition state for a molecule of interest are factored into internal and external...

  5. Heart Rate Response and Lactic Acid Concentration in Squash Players.

    ERIC Educational Resources Information Center

    Beaudin, Paula; And Others

    1978-01-01

    It was concluded that playing squash is an activity that results in heart rate responses of sufficient intensity to elicit aerobic training effects without producing high lactic acid concentration in the blood. (MM)

  6. Amino acid substitutions in genetic variants of human serum albumin and in sequences inferred from molecular cloning

    SciTech Connect

    Takahashi, N.; Takahashi, Y.; Blumberg, B.S.; Putnam, F.W.

    1987-07-01

    The structural changes in four genetic variants of human serum albumin were analyzed by tandem high-pressure liquid chromatography (HPLC) of the tryptic peptides, HPLC mapping and isoelectric focusing of the CNBr fragments, and amino acid sequence analysis of the purified peptides. Lysine-372 of normal (common) albumin A was changed to glutamic acid both in albumin Naskapi, a widespread polymorphic variant of North American Indians, and in albumin Mersin found in Eti Turks. The two variants also exhibited anomalous migration in NaDodSO/sub 4//PAGE, which is attributed to a conformational change. The identity of albumins Naskapi and Mersin may have originated through descent from a common mid-Asiatic founder of the two migrating ethnic groups, or it may represent identical but independent mutations of the albumin gene. In albumin Adana, from Eti Turks, the substitution site was not identified but was localized to the region from positions 447 through 548. The substitution of aspartic acid-550 by glycine was found in albumin Mexico-2 from four individuals of the Pima tribe. Although only single-point substitutions have been found in these and in certain other genetic variants of human albumin, five differences exist in the amino acid sequences inferred from cDNA sequences by workers in three other laboratories. However, our results on albumin A and on 14 different genetic variants accord with the amino acid sequence of albumin deduced from the genomic sequence. The apparent amino acid substitutions inferred from comparison of individual cDNA sequences probably reflect artifacts in cloning or in cDNA sequence analysis rather than polymorphism of the coding sections of the albumin gene.

  7. Amino acid substitutions in genetic variants of human serum albumin and in sequences inferred from molecular cloning.

    PubMed

    Takahashi, N; Takahashi, Y; Blumberg, B S; Putnam, F W

    1987-07-01

    The structural changes in four genetic variants of human serum albumin were analyzed by tandem high-pressure liquid chromatography (HPLC) of the tryptic peptides, HPLC mapping and isoelectric focusing of the CNBr fragments, and amino acid sequence analysis of the purified peptides. Lysine-372 of normal (common) albumin A was changed to glutamic acid both in albumin Naskapi, a widespread polymorphic variant of North American Indians, and in albumin Mersin found in Eti Turks. The two variants also exhibited anomalous migration in NaDodSO4/PAGE, which is attributed to a conformational change. The identity of albumins Naskapi and Mersin may have originated through descent from a common mid-Asiatic founder of the two migrating ethnic groups, or it may represent identical but independent mutations of the albumin gene. In albumin Adana, from Eti Turks, the substitution site was not identified but was localized to the region from positions 447 through 548. The substitution of aspartic acid-550 by glycine was found in albumin Mexico-2 from four individuals of the Pima tribe. Although only single-point substitutions have been found in these and in certain other genetic variants of human albumin, five differences exist in the amino acid sequences inferred from cDNA sequences by workers in three other laboratories. However, our results on albumin A and on 14 different genetic variants accord with the amino acid sequence of albumin deduced from the genomic sequence. The apparent amino acid substitutions inferred from comparison of individual cDNA sequences probably reflect artifacts in cloning or in cDNA sequence analysis rather than polymorphism of the coding sections of the albumin gene.

  8. Predicting the functional consequences of cancer-associated amino acid substitutions

    PubMed Central

    Shihab, Hashem A.; Gough, Julian; Cooper, David N.; Day, Ian N. M.; Gaunt, Tom R.

    2013-01-01

    Motivation: The number of missense mutations being identified in cancer genomes has greatly increased as a consequence of technological advances and the reduced cost of whole-genome/whole-exome sequencing methods. However, a high proportion of the amino acid substitutions detected in cancer genomes have little or no effect on tumour progression (passenger mutations). Therefore, accurate automated methods capable of discriminating between driver (cancer-promoting) and passenger mutations are becoming increasingly important. In our previous work, we developed the Functional Analysis through Hidden Markov Models (FATHMM) software and, using a model weighted for inherited disease mutations, observed improved performances over alternative computational prediction algorithms. Here, we describe an adaptation of our original algorithm that incorporates a cancer-specific model to potentiate the functional analysis of driver mutations. Results: The performance of our algorithm was evaluated using two separate benchmarks. In our analysis, we observed improved performances when distinguishing between driver mutations and other germ line variants (both disease-causing and putatively neutral mutations). In addition, when discriminating between somatic driver and passenger mutations, we observed performances comparable with the leading computational prediction algorithms: SPF-Cancer and TransFIC. Availability and implementation: A web-based implementation of our cancer-specific model, including a downloadable stand-alone package, is available at http://fathmm.biocompute.org.uk. Contact: fathmm@biocompute.org.uk Supplementary information: Supplementary data are available at Bioinformatics online. PMID:23620363

  9. High-Temperature Decomposition of Brønsted Acid Sites in Gallium-Substituted Zeolites

    SciTech Connect

    K Al-majnouni; N Hould; W Lonergan; D Vlachos; R Lobo

    2011-12-31

    The dehydroxylation of Broensted acid sites (BAS) in Ga-substituted zeolites was investigated at temperatures up to 850 C using X-ray absorption spectroscopy (XAS), Fourier transform infrared spectroscopy (FTIR), and mass spectrometry-temperature programmed desorption (MS-TPD). X-ray absorption near-edge spectroscopy (XANES) revealed that the majority of gallium has tetrahedral coordination even after complete dehydroxylation. The interatomic gallium-oxygen distance and gallium coordination number determined by extended X-ray absorption fine structure (EXAFS) are consistent with gallium in tetrahedral coordination at low T (< 550 C). Upon heating Ga-Beta and Ga-ZSM5 to 850 C, analysis of the EXAFS showed that 70 and 80% of the gallium was still in tetrahedral coordination. The remainder of the gallium was found to be in octahedral coordination. No trigonal Ga atoms were observed. FTIR measurements carried out at similar temperatures show that the intensity of the OH vibration due to BAS has been eliminated. MS-TPD revealed that hydrogen in addition to water evolved from the samples during dehydroxylation. This shows that dehydrogenation in addition to dehydration is a mechanism that contributes to BAS decomposition. Dehydrogenation was further confirmed by exposing the sample to hydrogen to regenerate some of the BAS as monitored by FTIR and MS-TPD.

  10. Change in oligomerization specificity of the p53 tetramerization domain by hydrophobic amino acid substitutions.

    PubMed Central

    Stavridi, E. S.; Chehab, N. H.; Caruso, L. C.; Halazonetis, T. D.

    1999-01-01

    The tumor suppressor function of the wild-type p53 protein is transdominantly inhibited by tumor-derived mutant p53 proteins. Such transdominant inhibition limits the prospects for gene therapy approaches that aim to introduce wild-type p53 into cancer cells. The molecular mechanism for transdominant inhibition involves sequestration of wild-type p53 subunits into inactive wild-type/mutant hetero-tetramers. Thus, p53 proteins, whose oligomerization specificity is altered so they cannot interact with tumor-derived mutant p53, would escape transdominant inhibition. Aided by the known three-dimensional structure of the p53 tetramerization domain and by trial and error we designed a novel domain with seven amino acid substitutions in the hydrophobic core. A full-length p53 protein bearing this novel domain formed homo-tetramers and had tumor suppressor function, but did not hetero-oligomerize with tumor-derived mutant p53 and resisted transdominant inhibition. Thus, hydrophobic core residues influence the oligomerization specificity of the p53 tetramerization domain. PMID:10493578

  11. Calf rearing in the tropics: growth rates and utilisation of milk substitute diets.

    PubMed

    Black, D H

    1982-08-01

    Twelve Brahman (Bos indicus) and 36 Holstein (Bos taurus) calves in Trinidad were fed on an amount of milk substitute equivalent to either 10 or 20% of their body weight each day. The diets contained either 10, 20 or 30% added fat. Nutrient balances were studied for 21 days. The liveweight gains of the calves depended on the quantity of milk fed and the breed of calf (Holstein greater than Brahman) but were independent of fat level. The Brahman calves were reluctant to accept milk substitute from a bucket. The digestibility of dry matter, nitrogen and fat was slightly higher for the Holstein than for the Brahman calves but there was little difference between the ME requirements for the 2 breeds. PMID:7123668

  12. Degradation of chloro- and methyl-substituted benzoic acids by a genetically modified microorganism

    SciTech Connect

    Mueller, R.; Deckwer, W.D.; Hecht, V.

    1996-09-05

    Degradation of 3-chlorobenzoic acid (3CB), 4-chlorobenzoic acid (4CB), and 4-methylbenzoic acid (4MB) as single substrates (carbon sources) and as a substrate mixture were studied in batch and continuous culture using the genetically modified microorganism Pseudomonas sp. B13 FR1 SN45P. The strain was able to mineralize the single compounds as well as the substrate mixture completely. Conversion of the three compounds in the substrate mixture proceeded simultaneously. Maximum specific substrate conversion rates were calculated to be 0.9 g g{sup {minus}1} h{sup {minus}1} for 3 CB and 4CB and 1.1 g g{sup {minus}1} h{sup {minus}1} for 4MB. Mass balances indicated the transient accumulation of pathway intermediates during batch cultivations. Hence, the rate limiting step in the degradative pathway is not the initial microbial attack of the original substrate or its transport through the cell membrane. Degradation rates on 3CB were comparable to those of the parent strain Pseudomonas sp. B13. The stability of the degradation pathways of strain Pseudomonas sp. B13 FR1 SN45P could be demonstrated in a continuous cultivation over 3.5 months (734 generation times) on 3CB, 4MB, and 4CB, which were used a single carbon sources one after the other.

  13. Photophysical properties and photochemistry of substituted cinnamates and cinnamic acids for UVB blocking: effect of hydroxy, nitro, and fluoro substitutions at ortho, meta, and para positions.

    PubMed

    Promkatkaew, Malinee; Suramitr, Songwut; Karpkird, Thitinun; Wanichwecharungruang, Supason; Ehara, Masahiro; Hannongbua, Supa

    2014-03-01

    Photophysical properties and photochemistry of various substituted cinnamates and cinnamic acids for ultraviolet B blocking were investigated experimentally and theoretically. This series includes monohydroxy, -nitro, and -fluoro derivatives. The absorption spectra were satisfactorily reproduced by the direct SAC-CI method with respect to the peak position and intensity. The transition character of the low-lying two ππ* and σπ* states for these 18 derivatives was analyzed. The para derivatives have a different transition character of the ππ* transitions compared with those of the ortho and meta derivatives. To elucidate the relaxation mechanism, the emission spectra were observed with oxygen quenching and the photostability was examined experimentally. The calculated radiative lifetimes indicate that the ortho- and meta-substituted derivatives have longer lifetimes for emission than the para derivatives. The potential energy curves of the first and second singlet excited states of the hydroxy derivatives as well as the vertical singlet and triplet transitions were examined to investigate the relaxation qualitatively. The ortho and meta derivatives have an energy barrier or flat surface in S1 resulting in fluorescence, whereas the para derivatives show nonradiative decay without an energy barrier. The para-hydroxy derivative was found to be an excellent UV absorber based on its broad absorption in the UVB/UVA regions, less emission, and higher photostability.

  14. Electronic Effects of 11β Substituted 17β-Estradiol Derivatives and Instrumental Effects on the Relative Gas Phase Acidity

    NASA Astrophysics Data System (ADS)

    Bourgoin-Voillard, Sandrine; Fournier, Françoise; Afonso, Carlos; Zins, Emilie-Laure; Jacquot, Yves; Pèpe, Claude; Leclercq, Guy; Tabet, Jean-Claude

    2012-12-01

    Numerous studies have highlighted the role of the proton donor characteristics of the phenol group of 17β-estradiol (E2) in its association with the estrogen receptor alpha (ERα). Since the substitutions at position C(11) have been reported to modulate this association, we hypothesized that such substitutions may modify the phenol acidity. Hence, phenol gas-phase acidity of nine C(11)-substituted E2-derivatives were evaluated using the extended Cooks' kinetic method, which is a method widely used to determine thermochemical properties by mass spectrometry. To enhance accuracy in data collection we recorded data from several instruments, including quadrupole ion trap, triple quadrupole, and hybrid QqTOF. Indeed, we report for the first time the use of the QqTOF instrument to provide a novel means to improve data accuracy by giving access to an intermediate effective temperature range. All experimental gas-phase acidity values were supported by theoretical calculations. Our results confirmed the ability of distant substituents at C(11) to modulate the phenol acidity through electrostatic interactions, electron withdrawing inductive effects, and mesomeric effects. However, no relationship was found between the phenol gas-phase acidity of investigated steroids and their binding affinity for ERα assessed in solution. Thus, our results highlight that the intrinsic properties of the hormone do not influence sufficiently the stabilization of the hormone/ERα complex. It is more likely that such stabilization would be more related to factors depending on the environment within the binding pocket such as hydrophobic, steric as well as direct intermolecular electrostatic effects between ERα residues and the substituted steroidal estrogens.

  15. Imidase catalyzing desymmetric imide hydrolysis forming optically active 3-substituted glutaric acid monoamides for the synthesis of gamma-aminobutyric acid (GABA) analogs.

    PubMed

    Nojiri, Masutoshi; Hibi, Makoto; Shizawa, Hiroaki; Horinouchi, Nobuyuki; Yasohara, Yoshihiko; Takahashi, Satomi; Ogawa, Jun

    2015-12-01

    The recent use of optically active 3-substituted gamma-aminobutyric acid (GABA) analogs in human therapeutics has identified a need for an efficient, stereoselective method of their synthesis. Here, bacterial strains were screened for enzymes capable of stereospecific hydrolysis of 3-substituted glutarimides to generate (R)-3-substituted glutaric acid monoamides. The bacteria Alcaligenes faecalis NBRC13111 and Burkholderia phytofirmans DSM17436 were discovered to hydrolyze 3-(4-chlorophenyl) glutarimide (CGI) to (R)-3-(4-chlorophenyl) glutaric acid monoamide (CGM) with 98.1% enantiomeric excess (e.e.) and 97.5% e.e., respectively. B. phytofirmans DSM17436 could also hydrolyze 3-isobutyl glutarimide (IBI) to produce (R)-3-isobutyl glutaric acid monoamide (IBM) with 94.9% e.e. BpIH, an imidase, was purified from B. phytofirmans DSM17436 and found to generate (R)-CGM from CGI with specific activity of 0.95 U/mg. The amino acid sequence of BpIH had a 75% sequence identity to that of allantoinase from A. faecalis NBRC13111 (AfIH). The purified recombinant BpIH and AfIH catalyzed (R)-selective hydrolysis of CGI and IBI. In addition, a preliminary investigation of the enzymatic properties of BpIH and AfIH revealed that both enzymes were stable in the range of pH 6-10, with an optimal pH of 9.0, stable at temperatures below 40 °C, and were not metalloproteins. These results indicate that the use of this class of hydrolase to generate optically active 3-substituted glutaric acid monoamide could simplify the production of specific chiral GABA analogs for drug therapeutics.

  16. A New Hierarchy of Phylogenetic Models Consistent with Heterogeneous Substitution Rates

    PubMed Central

    Woodhams, Michael D.; Fernández-Sánchez, Jesús; Sumner, Jeremy G.

    2015-01-01

    When the process underlying DNA substitutions varies across evolutionary history, some standard Markov models underlying phylogenetic methods are mathematically inconsistent. The most prominent example is the general time-reversible model (GTR) together with some, but not all, of its submodels. To rectify this deficiency, nonhomogeneous Lie Markov models have been identified as the class of models that are consistent in the face of a changing process of DNA substitutions regardless of taxon sampling. Some well-known models in popular use are within this class, but are either overly simplistic (e.g., the Kimura two-parameter model) or overly complex (the general Markov model). On a diverse set of biological data sets, we test a hierarchy of Lie Markov models spanning the full range of parameter richness. Compared against the benchmark of the ever-popular GTR model, we find that as a whole the Lie Markov models perform well, with the best performing models having 8–10 parameters and the ability to recognize the distinction between purines and pyrimidines. PMID:25858352

  17. Long-term care and hospital utilisation by older people: an analysis of substitution rates.

    PubMed

    Forder, Julien

    2009-11-01

    Older people are intensive users of hospital and long-term care services. This paper explores the extent to which these services are substitutes. A small area analysis was used with both care home and (tariff cost-weighted) hospital utilisation for older people aggregated to electoral wards in England.Health and social-care structural equations were specified using a theoretical model. The estimation accounted for the skewed and censored nature of the data. For health utilisation, both a fixed effects instrumental variables GMM model and a generalised estimating equations (GEE) model were fitted, the later on a log dependent variable with predicted values of social care utilisation used to account for endogeneity (bootstrapping was used to derive standard errors). In addition to a GMM model, the social-care estimation used both two-part and tobit models (also with predicted health utilisation and bootstrapping).The results indicate that for each additional pound1 spent on care homes, hospital expenditure falls by pound0.35. Also, pound1 additional hospital spend corresponds to just over pound0.35 reduction on care home spend. With these cost substitution effects offsetting, a transfer of resources to care homes is efficient if the resultant outcome gain is greater than the outcome loss from reduced hospital use.

  18. Reactivity of Cations and Zwitterions Formed in Photochemical and Acid-Catalyzed Reactions from m-Hydroxycycloalkyl-Substituted Phenol Derivatives.

    PubMed

    Cindro, Nikola; Antol, Ivana; Mlinarić-Majerski, Kata; Halasz, Ivan; Wan, Peter; Basarić, Nikola

    2015-12-18

    Three m-substituted phenol derivatives, each with a labile benzylic alcohol group and bearing either protoadamantyl 4, homoadamantyl 5, or a cyclohexyl group 6, were synthesized and their thermal acid-catalyzed and photochemical solvolytic reactivity studied, using preparative irradiations, fluorescence measurements, nanosecond laser flash photolysis, and quantum chemical calculations. The choice of m-hydroxy-substitution was driven by the potential for these phenolic systems to generate m-quinone methides on photolysis, which could ultimately drive the excited-state pathway, as opposed to forming simple benzylic carbocations in the corresponding thermal route. Indeed, thermal acid-catalyzed reactions gave the corresponding cations, which undergo rearrangement and elimination from 4, only elimination from 5, and substitution and elimination from 6. On the other hand, upon photoexcitation of 4-6 to S1 in a polar protic solvent, proton dissociation from the phenol, coupled with elimination of the benzylic OH (as hydroxide ion) gave zwitterions (formal m-quinone methides). The zwitterions exhibit reactivity different from the corresponding cations due to a difference in charge distribution, as shown by DFT calculations. Thus, protoadamantyl zwitterion has a less nonclassical character than the corresponding cation, so it does not undergo 1,2-shift of the carbon atom, as observed in the acid-catalyzed reaction. PMID:26595342

  19. Enantioconvergent Nucleophilic Substitution Reaction of Racemic Alkyne-Dicobalt Complex (Nicholas Reaction) Catalyzed by Chiral Brønsted Acid.

    PubMed

    Terada, Masahiro; Ota, Yusuke; Li, Feng; Toda, Yasunori; Kondoh, Azusa

    2016-08-31

    Catalytic enantioselective syntheses enable a practical approach to enantioenriched molecules. While most of these syntheses have been accomplished by reaction at the prochiral sp(2)-hybridized carbon atom, little attention has been paid to enantioselective nucleophilic substitution at the sp(3)-hybridized carbon atom. In particular, substitution at the chiral sp(3)-hybridized carbon atom of racemic electrophiles has been rarely exploited. To establish an unprecedented enantioselective substitution reaction of racemic electrophiles, enantioconvergent Nicholas reaction of an alkyne-dicobalt complex derived from racemic propargylic alcohol was developed using a chiral phosphoric acid catalyst. In the present enantioconvergent process, both enantiomers of the racemic alcohol were transformed efficiently to a variety of thioethers with high enantioselectivity. The key to achieving success is dynamic kinetic asymmetric transformation (DYKAT) of enantiomeric cationic intermediates generated via dehydroxylation of the starting racemic alcohol under the influence of the chiral phosphoric acid catalyst. The present fascinating DYKAT involves the efficient racemization of these enantiomeric intermediates and effective resolution of these enantiomers through utilization of the chiral conjugate base of the phosphoric acid. PMID:27490239

  20. Enantioconvergent Nucleophilic Substitution Reaction of Racemic Alkyne-Dicobalt Complex (Nicholas Reaction) Catalyzed by Chiral Brønsted Acid.

    PubMed

    Terada, Masahiro; Ota, Yusuke; Li, Feng; Toda, Yasunori; Kondoh, Azusa

    2016-08-31

    Catalytic enantioselective syntheses enable a practical approach to enantioenriched molecules. While most of these syntheses have been accomplished by reaction at the prochiral sp(2)-hybridized carbon atom, little attention has been paid to enantioselective nucleophilic substitution at the sp(3)-hybridized carbon atom. In particular, substitution at the chiral sp(3)-hybridized carbon atom of racemic electrophiles has been rarely exploited. To establish an unprecedented enantioselective substitution reaction of racemic electrophiles, enantioconvergent Nicholas reaction of an alkyne-dicobalt complex derived from racemic propargylic alcohol was developed using a chiral phosphoric acid catalyst. In the present enantioconvergent process, both enantiomers of the racemic alcohol were transformed efficiently to a variety of thioethers with high enantioselectivity. The key to achieving success is dynamic kinetic asymmetric transformation (DYKAT) of enantiomeric cationic intermediates generated via dehydroxylation of the starting racemic alcohol under the influence of the chiral phosphoric acid catalyst. The present fascinating DYKAT involves the efficient racemization of these enantiomeric intermediates and effective resolution of these enantiomers through utilization of the chiral conjugate base of the phosphoric acid.

  1. Decreased Diversity but Increased Substitution Rate in Host mtDNA as a Consequence of Wolbachia Endosymbiont Infection

    PubMed Central

    Shoemaker, D. DeWayne; Dyer, Kelly A.; Ahrens, Mike; McAbee, Kevin; Jaenike, John

    2004-01-01

    A substantial fraction of insects and other terrestrial arthropods are infected with parasitic, maternally transmitted endosymbiotic bacteria that manipulate host reproduction. In addition to imposing direct selection on the host to resist these effects, endosymbionts may also have indirect effects on the evolution of the mtDNA with which they are cotransmitted. Patterns of mtDNA diversity and evolution were examined in Drosophila recens, which is infected with the endosymbiont Wolbachia, and its uninfected sister species D. subquinaria. The level of mitochondrial, but not nuclear, DNA diversity is much lower in D. recens than in D. subquinaria, consistent with the hypothesized diversity-purging effects of an evolutionarily recent Wolbachia sweep. The dN/dS ratio in mtDNA is significantly greater in D. recens, suggesting that Muller's ratchet has brought about an increased rate of substitution of slightly deleterious mutations. The data also reveal elevated rates of synonymous substitutions in D. recens, suggesting that these sites may experience weak selection. These findings show that maternally transmitted endosymbionts can severely depress levels of mtDNA diversity within an infected host species, while accelerating the rate of divergence among mtDNA lineages in different species. PMID:15611174

  2. Synthesis of [.sup.13C] and [.sup.2H] substituted methacrylic acid, [.sup.13C] and [.sup.2H] substituted methyl methacrylate and/or related compounds

    DOEpatents

    Alvarez, Marc A.; Martinez, Rodolfo A.; Unkefer, Clifford J.

    2008-01-22

    The present invention is directed to labeled compounds of the formulae ##STR00001## wherein Q is selected from the group consisting of --S--, --S(.dbd.O)--, and --S(.dbd.O).sub.2--, Z is selected from the group consisting of 1-naphthyl, substituted 1-naphthyl, 2-naphthyl, substituted 2-naphthyl, and phenyl groups with the structure ##STR00002## wherein R.sub.1, R.sub.2, R.sub.3, R.sub.4 and R.sub.5 are each independently selected from the group consisting of hydrogen, a C.sub.1-C.sub.4 lower alkyl, a halogen, and an amino group selected from the group consisting of NH.sub.2, NHR and NRR' where R and R' are each independently selected from the group consisting of a C.sub.1-C.sub.4 lower alkyl, an aryl, and an alkoxy group, and X is selected from the group consisting of hydrogen, a C.sub.1-C.sub.4 lower alkyl group, and a fully-deuterated C.sub.1-C.sub.4 lower alkyl group. The present invention is also directed to a process of preparing labeled compounds, e.g., process of preparing [.sup.13C]methacrylic acid by reacting a (CH.sub.3CH.sub.2O--.sup.13C(O)--.sup.13CH.sub.2)-- aryl sulfone precursor with .sup.13CHI to form a (CH.sub.3CH.sub.2O--.sup.13C(O)--.sup.13C(.sup.13CH.sub.3).sub.2)-- aryl sulfone intermediate, and, reacting the (CH.sub.3CH.sub.2O--.sup.13C(O)--.sup.13C(.sup.13CH.sub.3).sub.2)-- aryl sulfone intermediate with sodium hydroxide, followed by acid to form [.sup.13C]methacrylic acid. The present invention is further directed to a process of preparing [.sup.2H.sub.8]methyl methacrylate by reacting a (HOOC--C(C.sup.2H.sub.3).sub.2-- aryl sulfinyl intermediate with CD.sub.3I to form a (.sup.2H.sub.3COOC--C(C.sup.2H.sub.3).sub.2)-- aryl sulfinyl intermediate, and heating the(.sup.2H.sub.3COOC--C(C.sup.2H.sub.3).sub.2)-- aryl sulfinyl intermediate at temperatures and for time sufficient to form [.sup.2H.sub.8]methyl methacrylate.

  3. Synthesis of ω-Oxo Amino Acids and trans-5-Substituted Proline Derivatives Using Cross-Metathesis of Unsaturated Amino Acids.

    PubMed

    Salih, Nabaz; Adams, Harry; Jackson, Richard F W

    2016-09-16

    A range of 7-oxo, 8-oxo, and 9-oxo amino acids, analogues of 8-oxo-2-aminodecanoic acid, one of the key components of the cyclic tetrapeptide apicidin, have been prepared by a three-step process involving copper-catalyzed allylation of serine-, aspartic acid-, and glutamic acid-derived organozinc reagents, followed by cross-metathesis of the resulting terminal alkenes with unsaturated ketones and hydrogenation. The intermediate 7-oxo-5-enones underwent a highly diastereoselective (dr ≥96:4) acid-catalyzed aza-Michael reaction to give trans-2,5-disubstituted pyrrolidines, 5-substituted proline derivatives. The aza-Michael reaction was first observed when the starting enones were allowed to stand in solution in deuterochloroform but can be efficiently promoted by catalytic amounts of dry HCl. PMID:27529354

  4. Reconstruction of the ancestral plastid genome in Geraniaceae reveals a correlation between genome rearrangements, repeats, and nucleotide substitution rates.

    PubMed

    Weng, Mao-Lun; Blazier, John C; Govindu, Madhumita; Jansen, Robert K

    2014-03-01

    Geraniaceae plastid genomes are highly rearranged, and each of the four genera already sequenced in the family has a distinct genome organization. This study reports plastid genome sequences of six additional species, Francoa sonchifolia, Melianthus villosus, and Viviania marifolia from Geraniales, and Pelargonium alternans, California macrophylla, and Hypseocharis bilobata from Geraniaceae. These genome sequences, combined with previously published species, provide sufficient taxon sampling to reconstruct the ancestral plastid genome organization of Geraniaceae and the rearrangements unique to each genus. The ancestral plastid genome of Geraniaceae has a 4 kb inversion and a reduced, Pelargonium-like small single copy region. Our ancestral genome reconstruction suggests that a few minor rearrangements occurred in the stem branch of Geraniaceae followed by independent rearrangements in each genus. The genomic comparison demonstrates that a series of inverted repeat boundary shifts and inversions played a major role in shaping genome organization in the family. The distribution of repeats is strongly associated with breakpoints in the rearranged genomes, and the proportion and the number of large repeats (>20 bp and >60 bp) are significantly correlated with the degree of genome rearrangements. Increases in the degree of plastid genome rearrangements are correlated with the acceleration in nonsynonymous substitution rates (dN) but not with synonymous substitution rates (dS). Possible mechanisms that might contribute to this correlation, including DNA repair system and selection, are discussed. PMID:24336877

  5. Nucleophilic Aromatic Substitution.

    ERIC Educational Resources Information Center

    Avila, Walter B.; And Others

    1990-01-01

    Described is a microscale organic chemistry experiment which demonstrates one feasible route in preparing ortho-substituted benzoic acids and provides an example of nucleophilic aromatic substitution chemistry. Experimental procedures and instructor notes for this activity are provided. (CW)

  6. Inter-relationships between solubilities, distribution coefficients and melting points of some substituted benzoic and phenylacetic acids.

    PubMed

    Armstrong, N A; James, K C; Wong, C K

    1979-09-01

    Ten 4-hydroxy and 4-alkoxy benzoic and phenylalkanoic acids have been investigated. Solubilities in aqueous buffer at pH 1.2 were determined, together with distribution coefficients between the buffer and either octanol or isopropyl myristate. When plotted against the total number of carbon atoms in the side chains, log octanol/water distribution coefficients gave two parallel straight lines, one for the substituted benzoic acids, and the other for the substituted phenylalkanoic acids. The slopes approximated to 0.5, the generally accepted value for methylene. Similar plots could be obtained with isopropyl myristate, provided the hydroxy acid results were ignored, and also when log aqueous solubilities were plotted against carbon number, although there was considerable scatter. The differences between the distribution coefficient results were explained in terms of solute-solvent interactions, and the scatter attributed to variations in the heats of fusion of the solutes. Yalkowsky's equation (1977), linking aqueous solubilities and melting points with distribution coefficients, was applied to the results, and found to be of limited predictive value. PMID:41067

  7. The Lys234Arg Substitution in the Enzyme SHV-72 Is a Determinant for Resistance to Clavulanic Acid Inhibition▿

    PubMed Central

    Mendonça, Nuno; Manageiro, Vera; Robin, Frédéric; Salgado, M. José; Ferreira, Eugénia; Caniça, Manuela; Bonnet, Richard

    2008-01-01

    The new β-lactamase SHV-72 was isolated from clinical Klebsiella pneumoniae INSRA1229, which exhibited the unusual association of resistance to the amoxicillin-clavulanic acid combination (MIC, 64 μg/ml) and susceptibility to cephalosporins, aztreonam, and imipenem. SHV-72 (pI 7.6) harbored the three amino acid substitutions Ile8Phe, Ala146Val, and Lys234Arg. SHV-72 had high catalytic efficiency against penicillins (kcat/Km, 35 to 287 μM−1·s−1) and no activity against oxyimino β-lactams. The concentration of clavulanic acid necessary to inhibit the enzyme activity by 50% was 10-fold higher for SHV-72 than for SHV-1. Molecular-dynamics simulation suggested that the Lys234Arg substitution in SHV-72 stabilized an atypical conformation of the Ser130 side chain, which moved the Oγ atom of Ser130 around 3.5 Å away from the key Oγ atom of the reactive serine (Ser70). This movement may therefore decrease the susceptibility to clavulanic acid by preventing cross-linking between Ser130 and Ser70. PMID:18316518

  8. The keto- and imine-epoxide rearrangements in the formation of substituted tetrahydrofurans and piperidines: Synthesis of ([+-])-teneraic acid; approahces to ([+-])-nemorensic acid and (+)-desoxoprosopinine

    SciTech Connect

    Kucharczyk, R.

    1992-01-01

    Previous work by Wasserman and co-workers showed that 2,7-dioxabicyclo-[2.21]heptanes can be formed via a thermal of Lewis acid-catalyzed rearrangement of 3,4-epoxy ketones. The author utilized this keto-epoxide rearrangement in the approach to the synthesis of nemorensic acid. A novel reaction of Grignard reagents with bicyclic acetals was developed permitting the stereoselective construction o9f highly substituted tetrahydrofurans. Thus, treatment of 1,4-dimethy1-2,7-dioxa-bicyclo[2.21]heptane with vinyl magnesium bromide resulted in Lewis acid-catalyzed acetal opening with subsequent chelation-controlled addition of the vinyl group to afford the tetrasubstituted tetrahydrofuran. Further functional group transformations demonstrated the feasibility of this approach in forming substituted tetrahydrofurans related to nemorensic acid. The carbonyl-epoxide rearrangement described above has been extended to imine epoxides in earlier work by Rusiecki. The resulting bicyclic oxazolidines can be stereoselectively reduced with hydride reagents to form substituted piperidines. The utility was demonstrated of this rearrangement/reduction methodology in the synthesis of [+-]-teneraic acid. The key steps in the synthesis were the formation of the bicyclic oxazolidine via the imine-epoxide rearrangement, and stereoselective hydride reduction to yield the trans piperidine. Oxidation of the t-butyloxycarbonyl-protected piperidine diol to the diacid, followed by deprotection of the amine, completed the synthesis of [+-]-teneraic acid. In seeking to apply the imine-epoxide rearrangement/reduction methodology in enantioselective syntheses, the author investigated an approach to the synthesis of (+)-desoxoprosopinine. This work has resulted in the successful construction of the desired intermediate keto epoxide which, with zinc chloride, rearranges cleanly to the corresponding bicyclic acetal. The corresponding imine-epoxide rearrangement has not been accomplished.

  9. Degradation rates of glycerol polyesters at acidic and basic conditions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polyesters prepared from glycerol with mixtures of adipic and citric acids were evaluated in the laboratory to estimate degradation rates over a range of pH conditions. These renewable polymers provide a market for glycerol that is generated during biodiesel production. The polyesters were prepared...

  10. Unusual coupling reactions of aldehydes and alkynes: a novel preparation of substituted phthalic acid derivatives by automated synthesis.

    PubMed

    Jacobi von Wangelin, Axel; Neumann, Helfried; Gördes, Dirk; Klaus, Stefan; Jiao, Haijun; Spannenberg, Anke; Krüger, Thomas; Wendler, Christian; Thurow, Kerstin; Stoll, Norbert; Beller, Matthias

    2003-05-23

    Based upon a highly versatile multicomponent methodology, a new one-pot synthesis of substituted phthalic acid derivatives from alpha,beta-unsaturated aldehydes was developed. The reaction involves the intermediacy of an acetamidodiene species which undergoes Diels-Alder addition to diethyl acetylenedicarboxylate. The resultant acetamidocyclohexadiene is subject to elimination of acetamide under the reaction conditions to give rise to substituted diethyl phthalates in good yields. This domino condensation-cycloaddition-elimination sequence has been applied to a variety of alpha,beta-unsaturated aldehydes. Furthermore, we demonstrated the exploitation of parallelized and automated synthesis technology for the rapid screening of reaction conditions and compositions. Detailed studies revealed the catalytic role of the employed acetamide and the occurrence of a stereoselective 1,4-syn elimination pathway under standard conditions.

  11. Identification of amino acids in lac repressor protein cross-linked to operator DNA specifically substituted with bromodeoxyuridine.

    PubMed

    Allen, T D; Wick, K L; Matthews, K S

    1991-04-01

    Amino acids in lac repressor protein which form cross-links to lac operator DNA specifically substituted with bromodeoxyuridine (BrdU) have been identified. Five sites of cross-linking in BrdU-substituted operator DNA were found at positions +3, +4, +14, +18, and +19 relative to the initiation site for transcription (Wick, K.L., and Matthews, K.S. (1991) J. Biol. Chem. 266, 6106-6112). Irradiation of complexes of repressor and each of these five singly substituted operator DNAs was executed under large scale conditions to generate sufficient complex for proteolysis, separation of the peptide-DNA, and peptide sequencing. The DNAs substituted with BrdU for thymidine at positions +3, +18, and +19 yielded cross-links to the peptide spanning residues 23-33, with the cross-link identified at His-29. Substitution at position +14 resulted in a cross-link to Tyr-17 within the peptide containing amino acids 13-22. These results are consistent with the structure determined by NMR and molecular dynamics calculations of the NH2-terminal headpiece-symmetric operator complex (Lamerichs, R.M.J.N., Boelens, R., van der Marel, G.A., van Boom, J.H., Kaptein, R., Buck, F., Fera, B., and Rüterjans, H. (1989) Biochemistry 28, 2895-2991; de Vlieg, J., Berendsen, H.J.C., and van Gunsteren, W.F. (1989) Proteins 6, 104-127). This structure indicates proximity of His-29 in the major groove to thymidines at positions +3 and +4. Since base pairs at positions +18 and +19 occupy symmetrical positions to +3 and +4 in the promoter distal region of the operator, it would be anticipated that cross-links similar to the +3 and +4 positions would form at these sites; this prediction is not borne out by the behavior at +4/+18, as no peptide could be identified cross-linked to DNA substituted at +4. Molecular dynamics simulations and the NMR data indicate that Tyr-17 interacts with the thymine at position +8, which is symmetrically related to position +14. Although BrdU-associated strand scission at +8

  12. Study of effect of substitution of the penultimate amino acid residue on expression, structure, and functional properties of Yersinia pseudotuberculosis OmpY porin.

    PubMed

    Solov'eva, T F; Tischenko, N M; Khomenko, V A; Portnyagina, O Y; Kim, N Y; Likhatskaya, G N; Novikova, O D; Isaeva, M P

    2014-07-01

    The purpose of the study was to compare the expression of two Yersinia pseudotuberculosis proteins, wild-type porin OmpY and the mutant porin OmpY designated as OmpY-Q having the uncharged amino acid residue Gln instead of positively charged Arg at the penultimate position in the same heterologous host. According to the literature, a similar substitution (Lys to Gln) of the penultimate amino acid residue in Neisseria meningitidis porin PorA drastically improved the assembly of the protein in the E. coli outer membrane in vivo. Site-directed mutagenesis was used to replace Arg by Gln (R338Q) in OmpY, and the conditions for optimal expression and maturation of OmpY-Q were selected. It was found that the growth rates of E. coli strains producing OmpY and OmpY-Q and the expression levels of the porins were approximately equal. Comparative analysis of recombinant OmpY and OmpY-Q did not show significant differences in structure, antigenic, and functional properties of the porins, or any noticeable effect of the R338Q substitution in OmpY on its assembly in the E. coli outer membrane in vivo. The probable causes of discrepancies between our results and the previous data on porin PorA are discussed considering the known mechanisms of biogenesis of porins at the periplasmic stage.

  13. Analogues of gamma-aminobutyric acid (GABA) and trans-4-aminocrotonic acid (TACA) substituted in the 2 position as GABAC receptor antagonists.

    PubMed

    Chebib, M; Vandenberg, R J; Johnston, G A

    1997-12-01

    1. gamma-Aminobutyric acid (GABA) and trans-4-aminocrotonic acid (TACA) have been shown to activate GABAC receptors. In this study, a range of C2, C3, C4 and N-substituted GABA and TACA analogues were examined for activity at GABAC receptors. 2. The effects of these compounds were examined by use of electrophysiological recording from Xenopus oocytes expressing the human rho 1 subunit of GABAC receptors with the two-electrode voltage-clamp method. 3. trans-4-Amino-2-fluorobut-2-enoic acid was found to be a potent agonist (KD = 2.43 microM). In contrast, trans-4-amino-2-methylbut-2-enoic acid was found to be a moderately potent antagonist (IC50 = 31.0 microM and KB = 45.5 microM). These observations highlight the possibility that subtle structural substitutions may change an agonist into an antagonist. 4. 4-Amino-2-methylbutanoic acid (KD = 189 microM), 4-amino-2-methylenebutanoic acid (KD = 182 microM) and 4-amino-2-chlorobutanoic acid (KD = 285 microM) were weak partial agonists. The intrinsic activities of these compounds were 12.1%, 4.4% and 5.2% of the maximal response of GABA, respectively. These compounds more effectively blocked the effects of the agonist, GABA, giving rise to KB values of 53 microM and 101 microM, respectively. 5. The sulphinic acid analogue of GABA, homohypotaurine, was found to be a potent partial agonist (KD = 4.59 microM, intrinsic activity 69%). 6. It was concluded that substitution of a methyl or a halo group in the C2 position of GABA or TACA is tolerated at GABAC receptors. However, there was dramatic loss of activity when these groups were substituted at the C3, C4 and nitrogen positions of GABA and TACA. 7. Molecular modelling studies on a range of active and inactive compounds indicated that the agonist/competitive antagonist binding site of the GABAC receptor may be smaller than that of the GABAA and GABAB receptors. It is suggested that only compounds that can attain relatively flat conformations may bind to the GABAC receptor

  14. The green-absorbing Drosophila Rh6 visual pigment contains a blue-shifting amino acid substitution that is conserved in vertebrates.

    PubMed

    Salcedo, Ernesto; Farrell, David M; Zheng, Lijun; Phistry, Meridee; Bagg, Eve E; Britt, Steven G

    2009-02-27

    The molecular mechanisms that regulate invertebrate visual pigment absorption are poorly understood. Through sequence analysis and functional investigation of vertebrate visual pigments, numerous amino acid substitutions important for this adaptive process have been identified. Here we describe a serine/alanine (S/A) substitution in long wavelength-absorbing Drosophila visual pigments that occurs at a site corresponding to Ala-292 in bovine rhodopsin. This S/A substitution accounts for a 10-17-nm absorption shift in visual pigments of this class. Additionally, we demonstrate that substitution of a cysteine at the same site, as occurs in the blue-absorbing Rh5 pigment, accounts for a 4-nm shift. Substitutions at this site are the first spectrally significant amino acid changes to be identified for invertebrate pigments sensitive to visible light and are the first evidence of a conserved tuning mechanism in vertebrate and invertebrate pigments of this class. PMID:19126545

  15. Precise investigation of the axial ligand substitution mechanism on a hydrogenphosphato-bridged lantern-type platinum(III) binuclear complex in acidic aqueous solution.

    PubMed

    Iwatsuki, Satoshi; Mizushima, Chiho; Morimoto, Naoyuki; Muranaka, Shinji; Ishihara, Koji; Matsumoto, Kazuko

    2005-10-31

    Detailed equilibrium and kinetic studies on axial water ligand substitution reactions of the "lantern-type" platinum(III) binuclear complex, [Pt(2)(mu-HPO(4))(4)(H(2)O)(2)](2)(-), with halide and pseudo-halide ions (X(-) = Cl(-), Br(-), and SCN(-)) were carried out in acidic aqueous solution at 25 degrees C with I = 1.0 M. The diaqua Pt(III) dimer complex is in acid dissociation equilibrium in aqueous solution with -log K(h1) = 2.69 +/- 0.04. The consecutive formation constants of the aquahalo complex () and the dihalo complex () were determined spectrophotometrically to be log = 2.36 +/- 0.01 and log = 1.47 +/- 0.01 for the reaction with Cl(-) and log = 2.90 +/- 0.04 and log = 2.28 +/- 0.01 for the reaction with Br(-), respectively. In the kinetic measurements carried out under the pseudo-first-order conditions with a large excess concentration of halide ion compared to that of Pt(III) dimer (C(X)()- > C(Pt)), all of the reactions proceeded via a one-step first-order reaction, which is a contrast to the consecutive two-step reaction for the amidato-bridged platinum(III) binuclear complexes. The conditional first-order rate constant (k(obs)) depended on C(X)()- as well as the acidity of the solution. From kinetic analyses, the rate-limiting step was determined to be the first substitution process that forms the monohalo species, which is in rapid equilibrium with the dihalo complex. The reaction with 4-penten-1-ol was also kinetically investigated to examine the reactivity of the lantern complex with olefin compounds.

  16. Amino acid substitution converts WEREWOLF function from an activator to a repressor of Arabidopsis non-hair cell development.

    PubMed

    Tominaga-Wada, Rumi; Nukumizu, Yuka; Wada, Takuji

    2012-02-01

    Root hair cell or non-hair cell fate determination in Arabidopsis thaliana root epidermis is model system for plant cell development. Two types of MYB transcription factors, the R2R3-type MYB, WEREWOLF (WER), and an R3-type MYB, CAPRICE (CPC), are involved in this cell fate determination process. To study the molecular basis of this process, we analyzed the functional relationship of WER and CPC. WER-CPC chimeric constructs were made from WER where all or parts of the MYB R3 region were replaced with the corresponding regions from CPC R3, and the constructs were introduced into the cpc-2 mutant. Although, the WER gene did not rescue the cpc-2 mutant 'small number of root hairs' phenotype, the WER-CPC chimera with two amino acids substitution (WC6) completely rescued the cpc-2 mutant phenotype. Furthermore, the WER-CPC chimera with 37 amino acids substitution (WC5) excessively rescued the cpc-2 mutant and induced 2.5 times more root hairs than wild-type. Consistent with this phenotype, GL2 gene expression was strongly reduced in WC5 in a cpc-2 background. Our results suggest that swapping at least two amino acids is sufficient to convert WER to CPC function. Therefore, these key residues may have strongly contributed to the selection of these important functions over evolution.

  17. Amino acid substitutions in the thymidine kinase gene of induced acyclovir-resistant herpes simplex virus type 1

    NASA Astrophysics Data System (ADS)

    Hussin, Ainulkhir; Nor, Norefrina Shafinaz Md; Ibrahim, Nazlina

    2013-11-01

    Acyclovir (ACV) is an antiviral drug of choice in healthcare setting to treat infections caused by herpes viruses, including, but not limited to genital herpes, cold sores, shingles and chicken pox. Acyclovir resistance has emerged significantly due to extensive use and misuse of this antiviral in human, especially in immunocompromised patients. However, it remains unclear about the amino acid substitutions in thymidine (TK) gene, which specifically confer the resistance-associated mutation in herpes simplex virus. Hence, acyclovir-resistant HSV-1 was selected at high concentration (2.0 - 4.5 μg/mL), and the TK-gene was subjected to sequencing and genotypic characterization. Genotypic sequences comparison was done using HSV-1 17 (GenBank Accesion no. X14112) for resistance-associated mutation determination whereas HSV-1 KOS, HSV-1 473/08 and HSV clinical isolates sequences were used for polymorphism-associated mutation. The result showed that amino acid substitutions at the non-conserved region (UKM-1: Gln34Lys, UKM-2: Arg32Ser & UKM-5: Arg32Cys) and ATP-binding site (UKM-3: Tyr53End & UKM-4: Ile54Leu) of the TK-gene. These discoveries play an important role to extend another dimension to the evolution of acyclovir-resistant HSV-1 and suggest that selection at high ACV concentration induced ACV-resistant HSV-1 evolution. These findings also expand the knowledge on the type of mutations among acyclovir-resistant HSV-1. In conclusion, HSV-1 showed multiple strategies to exhibit acyclovir resistance, including amino acid substitutions in the TK gene.

  18. Amino acid substitutions in naturally occurring variants of ail result in altered invasion activity.

    PubMed Central

    Beer, K B; Miller, V L

    1992-01-01

    Yersinia enterocolitica is the causative agent of a variety of gastrointestinal syndromes ranging from acute enteritis to mesenteric lymphadenitis. In addition, systemic infections resulting in high mortality rates can occur in elderly and immunocompromised patients. More than 50 serotypes of Y. enterocolitica have been identified, but only a few of them commonly cause disease in otherwise healthy hosts. Those serotypes that cause disease have been divided into two groups, American and non-American, based on their geographical distributions, biotypes, and pathogenicity. We have been studying two genes, inv and ail, from Y. enterocolitica that confer in tissue culture assays an invasive phenotype that strongly correlates with virulence. Some differences between the American and non-American serotypes at the ail locus were noted previously and have been investigated further in this report. The ail locus was cloned from seven Y. enterocolitica strains (seven different serotypes). Although the different clones produced similar amounts of Ail, the product of the ail gene from non-American serotypes (AilNA) was less able to promote invasion by Escherichia coli than was the product of the ail gene from American serotypes (AilA). This difference is probably due to one or more of the eight amino acid changes found in the derived amino acid sequence for the mature form of AilNA compared with that of AilA. Seven of these changes are predicted to be in cell surface domains of the protein (a model for the proposed folding of Ail within the outer membrane is presented). These results are discussed in relation to the growing family of outer membrane proteins, which includes Lom from bacteriophage lambda, PagC from salmonella typhimurium, and OmpX from Enterobacter cloacae. Images PMID:1370953

  19. Highly Amino Acid Selective Hydrolysis of Myoglobin at Aspartate Residues as Promoted by Zirconium(IV)-Substituted Polyoxometalates.

    PubMed

    Ly, Hong Giang T; Absillis, Gregory; Janssens, Rik; Proost, Paul; Parac-Vogt, Tatjana N

    2015-06-15

    SDS-PAGE/Edman degradation and HPLC MS/MS showed that zirconium(IV)-substituted Lindqvist-, Keggin-, and Wells-Dawson-type polyoxometalates (POMs) selectively hydrolyze the protein myoglobin at Asp-X peptide bonds under mildly acidic and neutral conditions. This transformation is the first example of highly sequence selective protein hydrolysis by POMs, a novel class of protein-hydrolyzing agents. The selectivity is directed by Asp residues located on the surface of the protein and is further assisted by electrostatic interactions between the negatively charged POMs and positively charged surface patches in the vicinity of the cleavage site.

  20. Arsenic scavenging by aluminum-substituted ferrihydrites in a circumneutral pH river impacted by acid mine drainage.

    PubMed

    Adra, Areej; Morin, Guillaume; Ona-Nguema, Georges; Menguy, Nicolas; Maillot, Fabien; Casiot, Corinne; Bruneel, Odile; Lebrun, Sophie; Juillot, Farid; Brest, Jessica

    2013-11-19

    Ferrihydrite (Fh) is a nanocrystalline ferric oxyhydroxide involved in the retention of pollutants in natural systems and in water-treatment processes. The status and properties of major chemical impurities in natural Fh is however still scarcely documented. Here we investigated the structure of aluminum-rich Fh, and their role in arsenic scavenging in river-bed sediments from a circumneutral river (pH 6-7) impacted by an arsenic-rich acid mine drainage (AMD). Extended X-ray absorption fine structure (EXAFS) spectroscopy at the Fe K-edge shows that Fh is the predominant mineral phase forming after neutralization of the AMD, in association with minor amount of schwertmannite transported from the AMD. TEM-EDXS elemental mapping and SEM-EDXS analyses combined with EXAFS analysis indicates that Al(3+) substitutes for Fe(3+) ions into the Fh structure in the natural sediment samples, with local aluminum concentration within the 25-30 ± 10 mol %Al range. Synthetic aluminous Fh prepared in the present study are found to be less Al-substituted (14-20 ± 5 mol %Al). Finally, EXAFS analysis at the arsenic K-edge indicates that As(V) form similar inner-sphere surface complexes on the natural and synthetic Al-substituted Fh studied. Our results provide direct evidence for the scavenging of arsenic by natural Al-Fh, which emphasize the possible implication of such material for scavenging pollutants in natural or engineered systems.

  1. A Delicate Balance When Substituting a Small Hydrophobe onto Low Molecular Weight Polyethylenimine to Improve Its Nucleic Acid Delivery Efficiency.

    PubMed

    Meneksedag-Erol, Deniz; KC, Remant Bahadur; Tang, Tian; Uludağ, Hasan

    2015-11-11

    High molecular weight (HMW) polyethylenimine (PEI) is one of the most versatile nonviral gene vectors that was extensively investigated over the past two decades. The cytotoxic profile of HMW PEI, however, encouraged a search for safer alternatives. Because of lack of cytotoxicity of low molecular weight (LMW) PEI, enhancing its performance via hydrophobic modifications has been pursued to this end. Since the performance of modified PEIs depends on the nature and extent of substituents, we systematically investigated the effect of hydrophobic modification of LMW (1.2 kDa) PEI with a short propionic acid (PrA). Moderate enhancements in PEI hydrophobicity resulted in enhanced cellular uptake of polyplexes and siRNA-induced silencing efficacy, whereas further increase in PrA substitution abolished the uptake as well as the silencing. We performed all-atom molecular dynamics simulations to elucidate the mechanistic details behind these observations. A new assembly mechanism was observed by the presence of hydrophobic PrA moieties, where PrA migrated to core of the polyplex. This phenomenon caused higher surface hydrophobicity and surface charge density at low substitutions, and it caused deleterious effects on surface hydrophobicity and cationic charge at higher substitutions. It is evident that an optimal balance of hydrophobicity/hydrophilicity is needed to achieve the desired polyplex properties for an efficient siRNA delivery, and our mechanistic findings should provide valuable insights for the design of improved substituents on nonviral carriers.

  2. Effects of Hypoxanthine Substitution in Peptide Nucleic Acids Targeting KRAS2 Oncogenic mRNA Molecules: Theory and Experiment

    PubMed Central

    Sanders, Jeffrey M.; Wampole, Matthew E.; Chen, Chang-Po; Sethi, Dalip; Singh, Amrita; Dupradeau, François-Yves; Wang, Fan; Gray, Brian D.; Thakur, Mathew L.; Wickstrom, Eric

    2013-01-01

    Genetic disorders can arise from single base substitutions in a single gene. A single base substitution for wild type guanine in the twelfth codon of KRAS2 mRNA occurs frequently to initiate lung, pancreatic, and colon cancer. We have observed single base mismatch specificity in radioimaging of mutant KRAS2 mRNA in tumors in mice by in vivo hybridization with radiolabeled peptide nucleic acid (PNA) dodecamers. We hypothesized that multi-mutant specificity could be achieved with a PNA dodecamer incorporating hypoxanthine, which can form Watson-Crick basepairs with adenine, cytosine, thymine, and uracil. Using molecular dynamics simulations and free energy calculations, we show that hypoxanthine substitutions in PNAs are tolerated in KRAS2 RNA-PNA duplexes where wild type guanine is replaced by mutant uracil or adenine in RNA. To validate our predictions, we synthesized PNA dodecamers with hypoxanthine, and then measured the thermal stability of RNA-PNA duplexes. Circular dichroism thermal melting results showed that hypoxanthine-containing PNAs are more stable in duplexes where hypoxanthine-adenine and hypoxanthine-uracil base pairs are formed than single mismatch duplexes or duplexes containing hypoxanthine-guanine opposition. PMID:23972113

  3. A Single Amino Acid Substitution in the Core Protein of West Nile Virus Increases Resistance to Acidotropic Compounds

    PubMed Central

    Martín-Acebes, Miguel A.; Blázquez, Ana-Belén; de Oya, Nereida Jiménez; Escribano-Romero, Estela; Shi, Pei-Yong; Saiz, Juan-Carlos

    2013-01-01

    West Nile virus (WNV) is a worldwide distributed mosquito-borne flavivirus that naturally cycles between birds and mosquitoes, although it can infect multiple vertebrate hosts including horses and humans. This virus is responsible for recurrent epidemics of febrile illness and encephalitis, and has recently become a global concern. WNV requires to transit through intracellular acidic compartments at two different steps to complete its infectious cycle. These include fusion between the viral envelope and the membrane of endosomes during viral entry, and virus maturation in the trans-Golgi network. In this study, we followed a genetic approach to study the connections between viral components and acidic pH. A WNV mutant with increased resistance to the acidotropic compound NH4Cl, which blocks organelle acidification and inhibits WNV infection, was selected. Nucleotide sequencing revealed that this mutant displayed a single amino acid substitution (Lys 3 to Glu) on the highly basic internal capsid or core (C) protein. The functional role of this replacement was confirmed by its introduction into a WNV infectious clone. This single amino acid substitution also increased resistance to other acidification inhibitor (concanamycin A) and induced a reduction of the neurovirulence in mice. Interestingly, a naturally occurring accompanying mutation found on prM protein abolished the resistant phenotype, supporting the idea of a genetic crosstalk between the internal C protein and the external glycoproteins of the virion. The findings here reported unveil a non-previously assessed connection between the C viral protein and the acidic pH necessary for entry and proper exit of flaviviruses. PMID:23874963

  4. A single amino acid substitution in the core protein of West Nile virus increases resistance to acidotropic compounds.

    PubMed

    Martín-Acebes, Miguel A; Blázquez, Ana-Belén; de Oya, Nereida Jiménez; Escribano-Romero, Estela; Shi, Pei-Yong; Saiz, Juan-Carlos

    2013-01-01

    West Nile virus (WNV) is a worldwide distributed mosquito-borne flavivirus that naturally cycles between birds and mosquitoes, although it can infect multiple vertebrate hosts including horses and humans. This virus is responsible for recurrent epidemics of febrile illness and encephalitis, and has recently become a global concern. WNV requires to transit through intracellular acidic compartments at two different steps to complete its infectious cycle. These include fusion between the viral envelope and the membrane of endosomes during viral entry, and virus maturation in the trans-Golgi network. In this study, we followed a genetic approach to study the connections between viral components and acidic pH. A WNV mutant with increased resistance to the acidotropic compound NH4Cl, which blocks organelle acidification and inhibits WNV infection, was selected. Nucleotide sequencing revealed that this mutant displayed a single amino acid substitution (Lys 3 to Glu) on the highly basic internal capsid or core (C) protein. The functional role of this replacement was confirmed by its introduction into a WNV infectious clone. This single amino acid substitution also increased resistance to other acidification inhibitor (concanamycin A) and induced a reduction of the neurovirulence in mice. Interestingly, a naturally occurring accompanying mutation found on prM protein abolished the resistant phenotype, supporting the idea of a genetic crosstalk between the internal C protein and the external glycoproteins of the virion. The findings here reported unveil a non-previously assessed connection between the C viral protein and the acidic pH necessary for entry and proper exit of flaviviruses.

  5. Synthesis and biological activity of some 5-substituted aminomethyl-8-hydroxyquinoline-7-sulphonic acids.

    PubMed

    Yanni, A S; Mohharam, A M

    1990-01-01

    5-Aryl (or alkyl)-8-hydroxyquinoline-7-sulphonic acids have been prepared by the Mannich reaction of 8-hydroxyquinoline-7-sulphonic acid with primary and secondary amines. Their bactericidal activities have been determined.

  6. 40 CFR 721.9460 - Tall oil fatty acids, reaction products with polyamines, alkyl substituted.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Tall oil fatty acids, reaction... SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.9460 Tall oil fatty acids, reaction... reporting. (1) The chemical substance identified generically as tall oil fatty acids, reaction products...

  7. 40 CFR 721.9460 - Tall oil fatty acids, reaction products with polyamines, alkyl substituted.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Tall oil fatty acids, reaction... SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.9460 Tall oil fatty acids, reaction... reporting. (1) The chemical substance identified generically as tall oil fatty acids, reaction products...

  8. 40 CFR 721.9460 - Tall oil fatty acids, reaction products with polyamines, alkyl substituted.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Tall oil fatty acids, reaction... SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.9460 Tall oil fatty acids, reaction... reporting. (1) The chemical substance identified generically as tall oil fatty acids, reaction products...

  9. 40 CFR 721.9460 - Tall oil fatty acids, reaction products with polyamines, alkyl substituted.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Tall oil fatty acids, reaction... SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.9460 Tall oil fatty acids, reaction... reporting. (1) The chemical substance identified generically as tall oil fatty acids, reaction products...

  10. 40 CFR 721.9460 - Tall oil fatty acids, reaction products with polyamines, alkyl substituted.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Tall oil fatty acids, reaction... SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.9460 Tall oil fatty acids, reaction... reporting. (1) The chemical substance identified generically as tall oil fatty acids, reaction products...

  11. 40 CFR 721.10107 - Naphthalenedisulfonic acid, [amino-hydroxy-[(substituted)azo-sulfo-naphthaleneyl]azo]-hydroxy...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false Naphthalenedisulfonic acid, azo... Significant New Uses for Specific Chemical Substances § 721.10107 Naphthalenedisulfonic acid, azo]-hydroxy... chemical substance identified generically as naphthalenedisulfonic acid, azo]-hydroxy- , metal salt (PMN...

  12. 40 CFR 721.10107 - Naphthalenedisulfonic acid, [amino-hydroxy-[(substituted)azo-sulfo-naphthaleneyl]azo]-hydroxy...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false Naphthalenedisulfonic acid, azo... Significant New Uses for Specific Chemical Substances § 721.10107 Naphthalenedisulfonic acid, azo]-hydroxy... chemical substance identified generically as naphthalenedisulfonic acid, azo]-hydroxy- , metal salt (PMN...

  13. 40 CFR 721.10107 - Naphthalenedisulfonic acid, [amino-hydroxy-[(substituted)azo-sulfo-naphthaleneyl]azo]-hydroxy...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Naphthalenedisulfonic acid, azo... Significant New Uses for Specific Chemical Substances § 721.10107 Naphthalenedisulfonic acid, azo]-hydroxy... chemical substance identified generically as naphthalenedisulfonic acid, azo]-hydroxy- , metal salt (PMN...

  14. 40 CFR 721.10107 - Naphthalenedisulfonic acid, [amino-hydroxy-[(substituted)azo-sulfo-naphthaleneyl]azo]-hydroxy...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false Naphthalenedisulfonic acid, azo... Significant New Uses for Specific Chemical Substances § 721.10107 Naphthalenedisulfonic acid, azo]-hydroxy... chemical substance identified generically as naphthalenedisulfonic acid, azo]-hydroxy- , metal salt (PMN...

  15. 40 CFR 721.10107 - Naphthalenedisulfonic acid, [amino-hydroxy-[(substituted)azo-sulfo-naphthaleneyl]azo]-hydroxy...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false Naphthalenedisulfonic acid, azo... Significant New Uses for Specific Chemical Substances § 721.10107 Naphthalenedisulfonic acid, azo]-hydroxy... chemical substance identified generically as naphthalenedisulfonic acid, azo]-hydroxy- , metal salt (PMN...

  16. Influence of the substitution of a grass cover by a mulch on infiltration rate

    NASA Astrophysics Data System (ADS)

    Sastre-Merlín, A.; Martínez-Pérez, S.; Bienes-Allas, R.; Molina-Navarro, E.; Martínez de Baroja, L.

    2012-04-01

    The study was carried out in an urban park of Madrid, in which it was decided to remove part of the prairie, replacing it by a mulch (pine bark). One year after this change in soil cover, infiltration tests were performed using the double ring infiltrometer (Müntz method). In each treatment the number of repetitions was 3. In the infiltration tests carried out the mulch not was withdraw, since we want to study their behavior before a rain or overhead irrigation. After one year, the infiltration rate showed ??much higher values in the prairie (18.9 mm h-1) than in the pine bark (8.4 mm h-1). Removing the prairie has meant a reduction in permeability of about 55%, which demonstrates the important role exerted by the radicular systems on infiltration. The origin is in the ability of roots to create preferred pathways circulation of the water. These pathways are of various types, and perhaps the most important are the root tubes, which are the channels that occur in the soil once the roots decompose. The finer roots create these pathways faster. These root tubes end up crumbling over time, so that is necessary to maintain the constant creation of new pipes in the soil. Under a prairie the number of root tubes that forms annually is enormous. By contrast, in absence of roots, in the surface horizon begins a process of gradual compaction, with reducing of the macroporosity and consequently impacting on the infiltration rate. The first consequence of this reduction in the infiltration rate is a poor flushing of salts from the soil of reclaimed water used in irrigation. This assertion has been corroborated by the analysis of the soil saturated paste, which shows an increasing of the electrical conductivities under the mulch. E.C. (μS cm-1) at the beginning of 2011 irrigation season (March) at different depths. Efficiency of the rains of autumn-winter by to wash soil salts. Depth (cm) PrairiePine bark 20 340 320 35 310 480 60 340 550 Therefore, the results indicate that

  17. Design, synthesis and SAR studies of GABA uptake inhibitors derived from 2-substituted pyrrolidine-2-yl-acetic acids.

    PubMed

    Steffan, Tobias; Renukappa-Gutke, Thejavathi; Höfner, Georg; Wanner, Klaus T

    2015-03-15

    In this paper, we disclose the design and synthesis of a series of 2-substituted pyrrolidine-2-yl-acetic acid as core structures and the N-arylalkyl derivatives thereof as potential GABA transport inhibitors. The 2-position in the side chain of pyrrolidine-2-yl-acetic acid derivatives was substituted with alkyl, hydroxy and amino groups to modulate the activity and selectivity to mGAT1 and mGAT4 proteins. SAR studies of the compounds performed for the four mouse GABA transporter proteins (mGAT1-mGAT4) implied significant potencies and subtype selectivities for 2-hydroxy-2-pyrrolidine-2-yl-acetic acid derivatives. The racemate rac-(u)-13c exhibited the highest potency (pIC50 5.67) at and selectivity for mGAT1 in GABA uptake assays. In fact, the potency of rac-(u)-13c at hGAT-1 (pIC50 6.14) was even higher than its potency at mGAT1. These uptake results for rac-(u)-13c are in line with the binding affinities to the aforesaid proteins mGAT1 (pKi 6.99) and hGAT-1 (pKi 7.18) determined by MS Binding Assay based on NO711 as marker quantified by LC-ESI-MS-MS analysis. Interestingly, the 2-hydroxy-2-pyrrolidine-2-yl-acetic acid rac-(u)-13d containing 2-{[tris(4-methoxyphenyl)]methoxy} ethyl group at the nitrogen atom of the pyrrolidine ring showed high potency at mGAT4 and a comparatively better selectivity for this protein (>15 against mGAT3) than the well known mGAT4 uptake inhibitor (S)-SNAP-5114.

  18. Antibacterial and anticancer activity of a series of novel peptides incorporating cyclic tetra-substituted C(α) amino acids.

    PubMed

    Hicks, Rickey P

    2016-09-15

    Eleven antimicrobial peptides (AMP) based on the incorporation of cyclic tetra substituted C(α) amino acids, as well as other unnatural amino acids were designed, synthesized and screened for in vitro activity against 18 strains of bacteria as well as 12 cancer cell lines. The AMPs discussed herein are derived from the following peptide sequence: Ac-GF(X)G(X)B(X)G(X)F(X)G(X)GB(X)BBBB-amide, X=any one of the following residues, A5c, A6c, Tic or Oic and B=any one of the following residues, Arg, Lys, Orn, Dpr or Dab. A diversity of in vitro inhibitory activity was observed for these AMPs. Several analogs exhibited single digit μM activity against drug resistant bacteria including; multiple drug resistant Mycobacterium tuberculosis, extremely drug resistant Mycobacterium tuberculosis and MRSA. The physicochemical properties of the basic amino acid residues incorporated into these AMPs seem to play a major role in defining antibacterial activity. Overall hydrophobicity seems to play a limited role in defining antibacterial activity. The ESKAPE pathogens were used to compare the activity of these AMPs to another family of synthetic AMPs incorporating the unnatural amino acids Tic and Oic. In most cases similarly substituted members of both families exhibited similar inhibitory activity against the ESKAPE pathogens. In specific cases differences in activity as high as 15 fold were observed between analogs. In addition four of these AMPs exhibited promising IC50 (<7.5μM) values against 12 different and diverse cancer cell lines. Five other AMPs exhibited promising IC50 (<7.5μM) values against selected cancer cell lines. PMID:27387357

  19. Antibacterial and anticancer activity of a series of novel peptides incorporating cyclic tetra-substituted C(α) amino acids.

    PubMed

    Hicks, Rickey P

    2016-09-15

    Eleven antimicrobial peptides (AMP) based on the incorporation of cyclic tetra substituted C(α) amino acids, as well as other unnatural amino acids were designed, synthesized and screened for in vitro activity against 18 strains of bacteria as well as 12 cancer cell lines. The AMPs discussed herein are derived from the following peptide sequence: Ac-GF(X)G(X)B(X)G(X)F(X)G(X)GB(X)BBBB-amide, X=any one of the following residues, A5c, A6c, Tic or Oic and B=any one of the following residues, Arg, Lys, Orn, Dpr or Dab. A diversity of in vitro inhibitory activity was observed for these AMPs. Several analogs exhibited single digit μM activity against drug resistant bacteria including; multiple drug resistant Mycobacterium tuberculosis, extremely drug resistant Mycobacterium tuberculosis and MRSA. The physicochemical properties of the basic amino acid residues incorporated into these AMPs seem to play a major role in defining antibacterial activity. Overall hydrophobicity seems to play a limited role in defining antibacterial activity. The ESKAPE pathogens were used to compare the activity of these AMPs to another family of synthetic AMPs incorporating the unnatural amino acids Tic and Oic. In most cases similarly substituted members of both families exhibited similar inhibitory activity against the ESKAPE pathogens. In specific cases differences in activity as high as 15 fold were observed between analogs. In addition four of these AMPs exhibited promising IC50 (<7.5μM) values against 12 different and diverse cancer cell lines. Five other AMPs exhibited promising IC50 (<7.5μM) values against selected cancer cell lines.

  20. Reduction Rates for Higher Americium Oxidation States in Nitric Acid

    SciTech Connect

    Grimes, Travis Shane; Mincher, Bruce Jay; Schmitt, Nicholas C

    2015-09-30

    The stability of hexavalent americium was measured using multiple americium concentrations and nitric acid concentrations after contact with the strong oxidant sodium bismuthate. Contrary to our hypotheses Am(VI) was not reduced faster at higher americium concentrations, and the reduction was only zero-order at short time scales. Attempts to model the reduction kinetics using zero order kinetic models showed Am(VI) reduction in nitric acid is more complex than the autoreduction processes reported by others in perchloric acid. The classical zero-order reduction of Am(VI) was found here only for short times on the order of a few hours. We did show that the rate of Am(V) production was less than the rate of Am(VI) reduction, indicating that some Am(VI) undergoes two electron-reduction to Am(IV). We also monitored the Am(VI) reduction in contact with the organic diluent dodecane. A direct comparison of these results with those in the absence of the organic diluent showed the reduction rates for Am(VI) were not statistically different for both systems. Additional americium oxidations conducted in the presence of Ce(IV)/Ce(III) ions showed that Am(VI) is reduced without the typical growth of Am(V) observed in the systems sans Ce ion. This was an interesting result which suggests a potential new reduction/oxidation pathway for Am in the presence of Ce; however, these results were very preliminary, and will require additional experiments to understand the mechanism by which this occurs. Overall, these studies have shown that hexavalent americium is fundamentally stable enough in nitric acid to run a separations process. However, the complicated nature of the reduction pathways based on the system components is far from being rigorously understood.

  1. Calcite growth-rate inhibition by fulvic acid and magnesium ion—Possible influence on biogenic calcite formation

    USGS Publications Warehouse

    Reddy, Michael M.

    2012-01-01

    Increases in ocean surface water dissolved carbon dioxide (CO2) concentrations retard biocalcification by reducing calcite supersaturation (Ωc). Reduced calcification rates may influence growth-rate dependent magnesium ion (Mg) incorporation into biogenic calcite modifying the use of calcifying organisms as paleoclimate proxies. Fulvic acid (FA) at biocalcification sites may further reduce calcification rates. Calcite growth-rate inhibition by FA and Mg, two common constituents of seawater and soil water involved in the formation of biogenic calcite, was measured separately and in combination under identical, highly reproducible experimental conditions. Calcite growth rates (pH=8.5 and Ωc=4.5) are reduced by FA (0.5 mg/L) to 47% and by Mg (10−4 M) to 38%, compared to control experiments containing no added growth-rate inhibitor. Humic acid (HA) is twice as effective a calcite growth-rate inhibitor as FA. Calcite growth rate in the presence of both FA (0.5 mg/L) and Mg (10−4 M) is reduced to 5% of the control rate. Mg inhibits calcite growth rates by substitution for calcium ion at the growth site. In contrast, FA inhibits calcite growth rates by binding multiple carboxylate groups on the calcite surface. FA and Mg together have an increased affinity for the calcite growth sites reducing calcite growth rates.

  2. An amino acid substitution in the human intestinal fatty acid binding protein is associated with increased fatty acid binding, increased fat oxidation, and insulin resistance.

    PubMed

    Baier, L J; Sacchettini, J C; Knowler, W C; Eads, J; Paolisso, G; Tataranni, P A; Mochizuki, H; Bennett, P H; Bogardus, C; Prochazka, M

    1995-03-01

    The intestinal fatty acid binding protein locus (FABP2) was investigated as a possible genetic factor in determining insulin action in the Pima Indian population. A polymorphism at codon 54 of FABP2 was identified that results in an alanine-encoding allele (frequency 0.71) and a threonine-encoding allele (frequency 0.29). Pimas who were homozygous or heterozygous for the threonine-encoding allele were found to have a higher mean fasting plasma insulin concentration, a lower mean insulin-stimulated glucose uptake rate, a higher mean insulin response to oral glucose and a mixed meal, and a higher mean fat oxidation rate compared with Pimas who were homozygous for the alanine-encoding allele. Since the FABP2 threonine-encoding allele was found to be associated with insulin resistance and increased fat oxidation in vivo, we further analyzed the FABP2 gene products for potential functional differences. Titration microcalorimetry studies with purified recombinant protein showed that the threonine-containing protein had a twofold greater affinity for long-chain fatty acids than the alanine-containing protein. We conclude that the threonine-containing protein may increase absorption and/or processing of dietary fatty acids by the intestine and thereby increase fat oxidation, which has been shown to reduce insulin action. PMID:7883976

  3. 2-substituted thiazolidine-4(R)-carboxylic acids as prodrugs of L-cysteine. Protection of mice against acetaminophen hepatotoxicity

    SciTech Connect

    Nagasawa, H.T.; Goon, D.J.; Muldoon, W.P.; Zera, R.T.

    1984-05-01

    A number of 2-alkyl- and 2-aryl-substituted thiazolidine-4(R)-carboxylic acids were evaluated for their protective effect against hepatotoxic deaths produced in mice by LD/sub 90/ doses of acetaminophen. 2(RS)-Methyl-, 2(RS)-n-propyl-, and 2(RS)-n- pentylthiazolidine -4(R)-carboxylic acids (compounds 1b,d,e, respectively) were nearly equipotent in their protective effect based on the number of surviving animals at 48 h as well as by histological criteria. 2(RS)-Ethyl-, 2(RS)-phenyl-, and 2(RS)-(4-pyridyl)thiazolidine-4(R)-carboxylic acids (compounds 1c,f,g) were less protective. The enantiomer of 1b, viz., 2(RS)- methylthiazolidine -4(S)-carboxylic acid (2b), was totally ineffective in this regard. Thiazolidine-4(R)-carboxylic acid (1a), but not its enantiomer, 2a, was a good substrate for a solubilized preparation of rat liver mitochondrial proline oxidase (K/sub m/ 1.1 x 10(-4) M; V/sub max/ . 5.4 mumol min-1 (mg of protein)-1). Compound 1b was not a substrate for proline oxidase but dissociated to L-cysteine in this system. At physiological pH and temperature, the hydrogens on the methyl group of 1b underwent deuterium exchange with solvent D/sub 2/O (k1 . 2.5 X 10(-5) s), suggesting that opening of the thiazolidine ring must have taken place. Indeed, 1b labeled with /sup 14/C in the 2 and methyl positions was rapidly metabolized by the rat to produce /sup 14/CO/sub 2/, 80% of the dose being excreted in this form in the expired air after 24 h. It is suggested that these 2-substituted thiazolidine-4(R)-carboxylic acids are prodrugs of L-cysteine that liberate this sulfhydryl amino acid in vivo by nonenzymatic ring opening, followed by solvolysis.

  4. 40 CFR 721.10045 - Diazotized substituted heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel...

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (generic). 721.10045... derivative, nickel complex, alkaline salt (generic). (a) Chemical substance and significant new uses subject... heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (PMN P-02-737)...

  5. 40 CFR 721.10045 - Diazotized substituted heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel...

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (generic). 721.10045... derivative, nickel complex, alkaline salt (generic). (a) Chemical substance and significant new uses subject... heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (PMN P-02-737)...

  6. 40 CFR 721.10045 - Diazotized substituted heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel...

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (generic). 721.10045... derivative, nickel complex, alkaline salt (generic). (a) Chemical substance and significant new uses subject... heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (PMN P-02-737)...

  7. 40 CFR 721.10045 - Diazotized substituted heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel...

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (generic). 721.10045... derivative, nickel complex, alkaline salt (generic). (a) Chemical substance and significant new uses subject... heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (PMN P-02-737)...

  8. 40 CFR 721.10045 - Diazotized substituted heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel...

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (generic). 721.10045... derivative, nickel complex, alkaline salt (generic). (a) Chemical substance and significant new uses subject... heteromonocycle coupled with naphthalene sulfonic acid derivative, nickel complex, alkaline salt (PMN P-02-737)...

  9. A More Challenging Interpretative Nitration Experiment Employing Substituted Benzoic Acids and Acetanilides

    ERIC Educational Resources Information Center

    Treadwell, Edward M.; Lin, Tung-Yin

    2008-01-01

    An experiment is described involving the nitration of ortho or meta monosubstituted benzoic acids (XC[subscript 6]H[subscript 4]CO[subscript 2]H, X = Halogen, Me, OH, or OMe) and monochlorinated acetanilides with nitric acid to determine the regioselectivity of addition by [superscript 1]H NMR spectroscopy and molecular modeling. Students were…

  10. Chiral phosphoric acid catalyzed highly enantioselective Friedel-Crafts alkylation reaction of C3-substituted indoles to β,γ-unsaturated α-ketimino esters.

    PubMed

    Bi, Bo; Lou, Qin-Xin; Ding, Yu-Yang; Chen, Sheng-Wei; Zhang, Sha-Sha; Hu, Wen-Hui; Zhao, Jun-Ling

    2015-02-01

    A highly enantioselective C2 Friedel-Crafts alkylation reaction of 3-substituted indoles to β,γ-unsaturated α-ketimino esters has been developed. This reaction was efficiently catalyzed by a chiral phosphoric acid catalyst. The corresponding C2-substituted indole derivatives, bearing an α-ketimino ester motif, were obtained in moderate to high yields (up to 93%) and with high enantioselectivities (up to >99% ee). PMID:25594307

  11. Susceptibilities of human immunodeficiency virus type 1 enzyme and viral variants expressing multiple resistance-engendering amino acid substitutions to reserve transcriptase inhibitors.

    PubMed Central

    Byrnes, V W; Emini, E A; Schleif, W A; Condra, J H; Schneider, C L; Long, W J; Wolfgang, J A; Graham, D J; Gotlib, L; Schlabach, A J

    1994-01-01

    To evaluate the potential that multiply resistant human immunodeficiency virus type 1 variants may arise during combination nucleoside and nonnucleoside reverse transcriptase inhibitor therapy, we constructed a series of mutant reverse transcriptase enzymes and viruses that coexpressed various combinations of resistance-associated amino acid substitutions. Substitutions at residues 100 (Leu-->Ile) and 181 (Tyr-->Cys), which mediate resistance to the nonnucleosides, suppressed resistance to 3'-azido-3'-deoxythymidine (AZT) when coexpressed with AZT-specific substitutions. However, a number of viral variants that exhibited significantly reduced susceptibilities to both classes of inhibitors were constructed. PMID:7522428

  12. Effect of single amino acid substitution on oxidative modifications of the Parkinson's disease-related protein, DJ-1.

    PubMed

    Madian, Ashraf G; Hindupur, Jagadish; Hulleman, John D; Diaz-Maldonado, Naomi; Mishra, Vartika R; Guigard, Emmanuel; Kay, Cyril M; Rochet, Jean-Christophe; Regnier, Fred E

    2012-02-01

    Mutations in the gene encoding DJ-1 have been identified in patients with familial Parkinson's disease (PD) and are thought to inactivate a neuroprotective function. Oxidation of the sulfhydryl group to a sulfinic acid on cysteine residue C106 of DJ-1 yields the "2O " form, a variant of the protein with enhanced neuroprotective function. We hypothesized that some familial mutations disrupt DJ-1 activity by interfering with conversion of the protein to the 2O form. To address this hypothesis, we developed a novel quantitative mass spectrometry approach to measure relative changes in oxidation at specific sites in mutant DJ-1 as compared with the wild-type protein. Treatment of recombinant wild-type DJ-1 with a 10-fold molar excess of H(2)O(2) resulted in a robust oxidation of C106 to the sulfinic acid, whereas this modification was not detected in a sample of the familial PD mutant M26I exposed to identical conditions. Methionine oxidized isoforms of wild-type DJ-1 were depleted, presumably as a result of misfolding and aggregation, under conditions that normally promote conversion of the protein to the 2O form. These data suggest that the M26I familial substitution and methionine oxidation characteristic of sporadic PD may disrupt DJ-1 function by disfavoring a site-specific modification required for optimal neuroprotective activity. Our findings indicate that a single amino acid substitution can markedly alter a protein's ability to undergo oxidative modification, and they imply that stimulating the conversion of DJ-1 to the 2O form may be therapeutically beneficial in familial or sporadic PD. PMID:22104028

  13. DNA-LCEB: a high-capacity and mutation-resistant DNA data-hiding approach by employing encryption, error correcting codes, and hybrid twofold and fourfold codon-based strategy for synonymous substitution in amino acids.

    PubMed

    Hafeez, Ibbad; Khan, Asifullah; Qadir, Abdul

    2014-11-01

    Data-hiding in deoxyribonucleic acid (DNA) sequences can be used to develop an organic memory and to track parent genes in an offspring as well as in genetically modified organism. However, the main concerns regarding data-hiding in DNA sequences are the survival of organism and successful extraction of watermark from DNA. This implies that the organism should live and reproduce without any functional disorder even in the presence of the embedded data. Consequently, performing synonymous substitution in amino acids for watermarking becomes a primary option. In this regard, a hybrid watermark embedding strategy that employs synonymous substitution in both twofold and fourfold codons of amino acids is proposed. This work thus presents a high-capacity and mutation-resistant watermarking technique, DNA-LCEB, for hiding secret information in DNA of living organisms. By employing the different types of synonymous codons of amino acids, the data storage capacity has been significantly increased. It is further observed that the proposed DNA-LCEB employing a combination of synonymous substitution, lossless compression, encryption, and Bose-Chaudary-Hocquenghem coding is secure and performs better in terms of both capacity and robustness compared to existing DNA data-hiding schemes. The proposed DNA-LCEB is tested against different mutations, including silent, miss-sense, and non-sense mutations, and provides substantial improvement in terms of mutation detection/correction rate and bits per nucleotide. A web application for DNA-LCEB is available at http://111.68.99.218/DNA-LCEB.

  14. Systematic methodology for the development of biocatalytic hydrogen-borrowing cascades: application to the synthesis of chiral α-substituted carboxylic acids from α-substituted α,β-unsaturated aldehydes.

    PubMed

    Knaus, Tanja; Mutti, Francesco G; Humphreys, Luke D; Turner, Nicholas J; Scrutton, Nigel S

    2015-01-01

    Ene-reductases (ERs) are flavin dependent enzymes that catalyze the asymmetric reduction of activated carbon-carbon double bonds. In particular, α,β-unsaturated carbonyl compounds (e.g. enals and enones) as well as nitroalkenes are rapidly reduced. Conversely, α,β-unsaturated esters are poorly accepted substrates whereas free carboxylic acids are not converted at all. The only exceptions are α,β-unsaturated diacids, diesters as well as esters bearing an electron-withdrawing group in α- or β-position. Here, we present an alternative approach that has a general applicability for directly obtaining diverse chiral α-substituted carboxylic acids. This approach combines two enzyme classes, namely ERs and aldehyde dehydrogenases (Ald-DHs), in a concurrent reductive-oxidative biocatalytic cascade. This strategy has several advantages as the starting material is an α-substituted α,β-unsaturated aldehyde, a class of compounds extremely reactive for the reduction of the alkene moiety. Furthermore no external hydride source from a sacrificial substrate (e.g. glucose, formate) is required since the hydride for the first reductive step is liberated in the second oxidative step. Such a process is defined as a hydrogen-borrowing cascade. This methodology has wide applicability as it was successfully applied to the synthesis of chiral substituted hydrocinnamic acids, aliphatic acids, heterocycles and even acetylated amino acids with elevated yield, chemo- and stereo-selectivity. A systematic methodology for optimizing the hydrogen-borrowing two-enzyme synthesis of α-chiral substituted carboxylic acids was developed. This systematic methodology has general applicability for the development of diverse hydrogen-borrowing processes that possess the highest atom efficiency and the lowest environmental impact.

  15. Self-assembled nanoparticles based on chondroitin sulfate-deoxycholic acid conjugates for docetaxel delivery: Effect of degree of substitution of deoxycholic acid.

    PubMed

    Liu, Mengrui; Du, Hongliang; Zhai, Guangxi

    2016-10-01

    Hydrophobically-modified polymers based on chondroitin sulfate with different degree of substitution (DS) of deoxycholic acid (DOCA) were developed for docetaxel delivery. Chondroitin sulfate-deoxycholic acid (CSAD) bioconjugates were synthesized via the linker of adipic dihydrazide by amide bond. They were characterized with spherical shape, mean diameter of around 165.2nm and negative zeta potential (-14.87 to -20.53mV). An increase of DOCA DS reduced size of nanoparticles, while increasing drug loading efficiency. Drug release in vitro showed a triphasic sustained pattern and higher accumulative drug release percentage was observed with increased DS of DOCA on polymer. Self-assemblies with higher DS also had enhanced internalization of nanoparticles and stronger cytotoxicity at the cellular level. The self-assemble nanoparticles demonstrate to be excellent targeting drug delivery systems and the desired therapeutics can be achieved via the alteration of DS. PMID:27343846

  16. In Vitro characterization of low modulus linoleic acid coated strontium-substituted hydroxyapatite containing PMMA bone cement.

    PubMed

    Lam, W M; Pan, H B; Fong, M K; Cheung, W S; Wong, K L; Li, Z Y; Luk, K D K; Chan, W K; Wong, C T; Yang, C; Lu, W W

    2011-01-01

    Poly (methyl methacrylate) (PMMA) bone cement is widely used in vertebral body augmentation procedures such as vertebroplasty and balloon kyphoplasty. Filling high modulus PMMA increases the modulus of filled verterbra, increasing the risk of fracture in the adjacent vertebra. On the other hand, in porous PMMA bone cements, wear particle generation and deterioration of mechanical performance are the major drawbacks. This study adopts a new approach by utilizing linoleic acid coated strontium substituted hydroxyapatite nanoparticle (Sr-5 HA) and linoleic acid as plasticizer reducing bone cement's modulus with minimal impact on its strength. We determined the compressive strength (UCS) and modulus (Ec), hydrophobicity, injectability, in vitro bioactivity and biocompatibility of this bone cement at different filler and linoleic acid loading. At 20 wt % Sr5-HA incorporation, UCS and Ec were reduced from 63 ± 2 MPa, 2142 ± 129 MPa to 58 ± 2 MPa, 1785 ± 64 MPa, respectively. UCS and Ec were further reduced to 49 ± 2 MPa and 774 ± 70 MPa respectively when 15 v/v of linoleic acid was incorporated. After 7 days of incubation, pre-osteoblast cells (MC3T3-E1) attached on 20 wt % Sr5-HA and 20 wt % Sr5-HA with 15 v/v of linoleic acid group were higher (3.73 ± 0.01 x 10⁴, 2.27 ± 0.02 x 10⁴) than their PMMA counterpart (1.83 ± 0.04 x 10⁴). Incorporation of Sr5-HA with linoleic acid in monomer phase is more effective in reducing the bone cement's stiffness than Sr5-HA alone. Combination of low stiffness and high mechanical strength gives the novel bone cement the potential for use in vertebroplasty cement applications. PMID:21053263

  17. Laboratory measurements of H-D substitution rates in solid methanol-dn (n=0-2) at 10 K

    NASA Astrophysics Data System (ADS)

    Nagaoka, Akihiro; Watanabe, Naoki; Kouchi, Akira

    The deuterium fractionation of interstellar methanol is investigated experimentally using the ASURA (Apparatus for SUrface Reactions in Astrophysics) system. Recent observations toward the low-mass protostars IRAS16293 found the very high D/H ratios in formaldehyde and methanol up to 0.2 and 0.4, respectively (Loinard et al. 2000; Parise et al. 2004; Aikawa et al. 2005). To date, several models have been proposed to explain D-fractionation mechanism. Pure gas-phase models are difficult to reproduce the D-fractionation, particularly, for multideuterated species, while the results of some gas-grain models can achieve the observed fractionation levels fairly well (Stantcheva & Herbst 2003). However, the gas-grain models require many assumptions regarding the grain surface reactions. Then, the experiments on the surface reaction have been highly desirable. In this context, we performed the experiments on the formation of deuterated formaldehyde and methanol on cold (10 K) interstellar grain analogues and revealed that a key route for the D-fractionation is not successive addition of H and D to CO as previously considered (e.g., Charnley, Tielens, & Rodgers 1997) but H-D substitution in solid CH3OH on icy grains (Nagaoka, Watanabe, & Kouchi 2005). We report the results of further experiments on the deuteration of CH3OH using a cold (30 K) atomic D beam. The relative rates of H-D substitution reactions; CH3OH → CH2DOH, CH2DOH → CHD2OH, CHD2OH → CD3OH, were measured. Experiments were performed using the ASURA system described previously (Watanabe et al. 2004; Nagaoka, Watanabe, & Kouchi 2005). The experimental procedure is as follows. An aluminum substrate was placed in the centre of an ultra-high vacuum chamber (10-10 Torr) and cooled to 10 K by a helium refrigerator. The solid samples of normal and deuterated methanol (CH3OH, CH2DOH, CHD2OH) were vapor-deposited on the substrate. The D atoms produced by dissociation of D2 molecules by microwave discharge were

  18. Functional impact of polar and acidic substitutions in the lactose repressor hydrophobic monomer.monomer interface with a buried lysine.

    PubMed

    Zhan, Hongli; Sun, Zhifei; Matthews, Kathleen Shive

    2009-02-17

    Despite predicted energetic penalties, the charged K84 side chains of tetrameric lactose repressor protein (LacI) are found buried within the highly hydrophobic monomer.monomer interface that includes side chains of V94 and V96. Once inducer binding has occurred, these K84 side chains move to interact with the more solvent-exposed side chains of D88 and E100'. Previous studies demonstrated that hydrophobic substitutions for K84 increased protein stability and significantly impaired the allosteric response. These results indicated that enhanced hydrophobic interactions at the monomer.monomer interface remove the energetic driving force of the buried charges, decreasing the likelihood of a robust conformational change and stabilizing the structure. We hypothesized that creating a salt bridge network with the lysine side chains by including nearby negatively charged residues might result in a similar outcome. To that end, acidic residues, D and E, and their neutral amides, N and Q, were substituted for the valines at positions 94 and 96. These variants exhibited one or more of the following functional changes: weakened inducer binding, impaired allosteric response, and diminished protein stability. For V96D and V96E, ion pair formation with K84 appears optimal, and the loss of inducer response exceeds that of the hydrophobic K84A and -L variants. However, impacts on functional properties indicate that stabilizing the buried positive charge with polar or ion pair interactions is not functionally equivalent to structural stabilization via hydrophobic enhancement. PMID:19166325

  19. Uptake of benzoic acid and chloro-substituted benzoic acids by alcaligenes denitrificans BRI 3010 and BRI 6011

    SciTech Connect

    Miguez, C.B.; Ingram, J.M.; MacLeod, R.A.

    1995-12-01

    The mechanism of uptake of benzoic and 2,4-dichlorobenzoic acid (2,4-DCBA) by Alcaligenes denitrificans BRI 3010 and BRI 6011 and Pseudomonas sp. strain B13, three organisms capable of degrading isomers of chlorinated benzoic acids, was investigated. In all three organisms, uptake of benzoic acid was inducible. For benzoic acid uptake into BRI 3010, monophasic saturation kinetics with apparent K{sub m} and V{sub max} values of 1.4 {mu}M and 3.2 nmol/min/mg of cell dry weight, respectively, were obtained. For BRI 6011, biphasic saturation kinetics were observed, suggesting presence of two uptake systems for benzoic acid with distinct K{sub m} (0.72 and 5.3 {mu}M) and V{sub max} (3.3 and 4.6 nmol/min/mg of cell dry weight) values. BRI 3010 and BRI 6011 accumulated benzoic acid against a concentration gradient by a factor of 8 and 10, respectively. A wide range of structural analogs, at 50-fold excess concentrations, inhibited benzoic acid uptake by BRI 3010 and BRI 6011, whereas with B13, only 3-chlorobenzoic acid was an effective inhibitor. For BRI 3010 and BRI 6011, the inhibition by the structural analogs was not of a competitive nature. Uptake of benzoic acid by BRI 3010 and BRI 6011 was inhibited by KCN, by the protonophore 3,5,3`, 4`-tetrachlorosalicylanilide (TCS), and, for BRI 6011, by anaerobiosis unless nitrate was present, thus indicating that energy was required for the uptake process. Uptake of 2,4-DCBA by BRI 6011 was constitutive and saturation uptake kinetics were not observed. Uptake of 2,4-DCBA by BRI 6011 was inhibited by KCN, TCS, and anaerobiosis even if nitrate was present, but the compound was not accumulated intracellularly against a concentration gradient. Uptake of 2,4-DCBA by BRI 6011 appears to occur by passive diffusion into the cell down its concentration gradient, which is maintained by the intracellular metabolism of the compound. This process could play an important role in the degradation of xenobiotic compounds by microorganisms.

  20. [Evaluation of folate substitution in women with epilepsy. Determination of erythrocyte folic acid concentrations].

    PubMed

    Bauer, J; Bös, M; Rück, J; Stoffel-Wagner, B

    2011-04-01

    Insufficient maternal folate concentrations appear to be a fetal risk factor for neural tube defects (NTD). Erythrocyte folate concentrations are widely accepted as an indicator of tissue folate storage. We retrospectively evaluated erythrocyte folate concentrations to examine if a recommended daily dosage of 5 mg folic acid is sufficient to balance the impact of antiepileptic drugs (AED) on folate metabolism in women with epilepsy. Data of 48 women (mean age 30.3 years) with idiopathic epilepsy with generalized seizures (n=12) or symptomatic epilepsy with focal seizures (n=36) were available, 43 women submitted to further analysis and 30 women received AED monotherapy. Duration of folic acid supplementation varied between 0.5 and 12 months. The daily dosage of folic acid ranged from 0.4 to 15 mg and 32 women received 5 mg/day. Erythrocyte folate concentrations ranged from 282 to 1596 ng/ml (mean 780 ng/ml). In 29 out of the 32 women (90.6%) on 5 mg folic acid per day, red cell folate was ≥400 ng/ml. In previous studies the risk for NTD was estimated to be 0.8‰ if red cell folate was ≥400 ng/ml. Our results suggest that 5 mg/day folic acid as preconception supplementation in women with epilepsy is effective to balance the impact of AEDs on folate metabolism in women with epilepsy.

  1. Amino acid substitution in Cryptococcus neoformans lanosterol 14-α-demethylase involved in fluconazole resistance in clinical isolates.

    PubMed

    Bosco-Borgeat, María E; Mazza, Mariana; Taverna, Constanza G; Córdoba, Susana; Murisengo, Omar A; Vivot, Walter; Davel, Graciela

    2016-01-01

    The molecular basis of fluconazole resistance in Cryptococcus neoformans has been poorly studied. A common azole resistance mechanism in Candida species is the acquisition of point mutations in the ERG11 gene encoding the enzyme lanosterol 14-α-demethylase, target of the azole class of drugs. In C. neoformans only two mutations were described in this gene. In order to evaluate other mutations that could be implicated in fluconazole resistance in C. neoformans we studied the genomic sequence of the ERG11 gene in 11 clinical isolates with minimal inhibitory concentration (MIC) values to fluconazole of ≥16μg/ml. The sequencing revealed the G1855A mutation in 3 isolates, resulting in the enzyme amino acid substitution G484S. These strains were isolated from two fluconazole-treated patients. This mutation would not intervene in the susceptibility to itraconazole and voriconazole. PMID:27311753

  2. X-ray photoelectron spectroscopy study of para-substituted benzoic acids chemisorbed to aluminum oxide thin films

    SciTech Connect

    Kreil, Justin; Ellingsworth, Edward; Szulczewski, Greg

    2013-11-15

    A series of para-substituted, halogenated (F, Cl, Br, and I) benzoic acid monolayers were prepared on the native oxide of aluminum surfaces by solution self-assembly and spin-coating techniques. The monolayers were characterized by x-ray photoelectron spectroscopy (XPS) and water contact angles. Several general trends are apparent. First, the polarity of the solvent is critical to monolayer formation. Protic polar solvents produced low coverage monolayers; in contrast, nonpolar solvents produced higher coverage monolayers. Second, solution deposition yields a higher surface coverage than spin coating. Third, the thickness of the monolayers determined from XPS suggests the plane of the aromatic ring is perpendicular to the surface with the carboxylate functional group most likely binding in a bidentate chelating geometry. Fourth, the saturation coverage (∼2.7 × 10{sup 14} molecules cm{sup −2}) is independent of the para-substituent.

  3. Morphology of the diastereomeric salt of the alkaloid ephedrine and a chlorine substituted cyclic phosphoric acid (CLINAM)

    NASA Astrophysics Data System (ADS)

    Strom, C. S.; Leusen, F. J. J.; Geertman, R. M.; Ariaans, G. J. A.

    1997-01-01

    The morphology of the diastereomeric salt of the alkaloid ephedrine and a chlorine substituted cyclic phosphoric acid is studied theoretically by means of a first-principles application of Hartman's PBC theory. A rigorous graph-theoretic derivation of the F slices of CLINAM and 2,4-DICLINAM has yielded all possible growth layers and their orientations. The Coulomb and Van der Waals contributions to the energy quantities characterizing CLINAM are calculated, using the Ewald formulation adjusted to lamina shapes, exactly and free from adjustable parameters. Several schemes of computing partial charges, in combination with energy minimization techniques are used for computing the atomic point charges. The structural morphology follows from the total attachment energies. The theoretical growth habit depends sensitively on the choice of the employed atomic charge scheme. The theoretical morphology of CLINAM crystals is discussed in the light of experimental results.

  4. Divergent reactivity in palladium-catalyzed annulation with diarylamines and α,β-unsaturated acids: direct access to substituted 2-quinolinones and indoles.

    PubMed

    Kancherla, Rajesh; Naveen, Togati; Maiti, Debabrata

    2015-06-01

    A palladium-catalyzed CH activation strategy has been successfully employed for exclusive synthesis of a variety of 3-substituted indoles. A [3+3] annulation for synthesizing substituted 2-quinolinones was recently developed by reaction of α,β-unsaturated carboxylic acids with diarylamines under acidic conditions. In the present work, an analogous [3+2] annulation is achieved from the same set of starting materials under basic conditions to generate 1,3-disubstituted indoles exclusively. Mechanistic studies revealed an ortho-palladation-π-coordination-β-migratory insertion-β-hydride elimination reaction sequence to be operative under the reaction conditions. PMID:25941155

  5. Anisotropic scattering rate in Fe-substituted Bi2Sr2Ca(Cu1-xFex)2O8+δ

    SciTech Connect

    Naamneh, M.; Lubashevsky, Y.; Lahoud, E.; Gu, G.; Kanigel, A.

    2015-05-27

    We measured the electronic structure of Fe substituted Bi2212 using Angle Resolved Photoemission Spectroscopy (ARPES). We find that the substitution does not change the momentum dependence of the superconducting gap but induces a very anisotropic enhancement of the scattering rate. A comparison of the effect of Fe substitution to that of Zn substitution suggests that the Fe reduces Tc so effectively because it supresses very strongly the coherence weight around the anti-nodes.

  6. Identification of Low- and High-Impact Hemagglutinin Amino Acid Substitutions That Drive Antigenic Drift of Influenza A(H1N1) Viruses

    PubMed Central

    Harvey, William T.; Benton, Donald J.; Gregory, Victoria; Hall, James P. J.; Daniels, Rodney S.; Bedford, Trevor; Haydon, Daniel T.; Hay, Alan J.; McCauley, John W.; Reeve, Richard

    2016-01-01

    Determining phenotype from genetic data is a fundamental challenge. Identification of emerging antigenic variants among circulating influenza viruses is critical to the vaccine virus selection process, with vaccine effectiveness maximized when constituents are antigenically similar to circulating viruses. Hemagglutination inhibition (HI) assay data are commonly used to assess influenza antigenicity. Here, sequence and 3-D structural information of hemagglutinin (HA) glycoproteins were analyzed together with corresponding HI assay data for former seasonal influenza A(H1N1) virus isolates (1997–2009) and reference viruses. The models developed identify and quantify the impact of eighteen amino acid substitutions on the antigenicity of HA, two of which were responsible for major transitions in antigenic phenotype. We used reverse genetics to demonstrate the causal effect on antigenicity for a subset of these substitutions. Information on the impact of substitutions allowed us to predict antigenic phenotypes of emerging viruses directly from HA gene sequence data and accuracy was doubled by including all substitutions causing antigenic changes over a model incorporating only the substitutions with the largest impact. The ability to quantify the phenotypic impact of specific amino acid substitutions should help refine emerging techniques that predict the evolution of virus populations from one year to the next, leading to stronger theoretical foundations for selection of candidate vaccine viruses. These techniques have great potential to be extended to other antigenically variable pathogens. PMID:27057693

  7. Identification of Low- and High-Impact Hemagglutinin Amino Acid Substitutions That Drive Antigenic Drift of Influenza A(H1N1) Viruses.

    PubMed

    Harvey, William T; Benton, Donald J; Gregory, Victoria; Hall, James P J; Daniels, Rodney S; Bedford, Trevor; Haydon, Daniel T; Hay, Alan J; McCauley, John W; Reeve, Richard

    2016-04-01

    Determining phenotype from genetic data is a fundamental challenge. Identification of emerging antigenic variants among circulating influenza viruses is critical to the vaccine virus selection process, with vaccine effectiveness maximized when constituents are antigenically similar to circulating viruses. Hemagglutination inhibition (HI) assay data are commonly used to assess influenza antigenicity. Here, sequence and 3-D structural information of hemagglutinin (HA) glycoproteins were analyzed together with corresponding HI assay data for former seasonal influenza A(H1N1) virus isolates (1997-2009) and reference viruses. The models developed identify and quantify the impact of eighteen amino acid substitutions on the antigenicity of HA, two of which were responsible for major transitions in antigenic phenotype. We used reverse genetics to demonstrate the causal effect on antigenicity for a subset of these substitutions. Information on the impact of substitutions allowed us to predict antigenic phenotypes of emerging viruses directly from HA gene sequence data and accuracy was doubled by including all substitutions causing antigenic changes over a model incorporating only the substitutions with the largest impact. The ability to quantify the phenotypic impact of specific amino acid substitutions should help refine emerging techniques that predict the evolution of virus populations from one year to the next, leading to stronger theoretical foundations for selection of candidate vaccine viruses. These techniques have great potential to be extended to other antigenically variable pathogens. PMID:27057693

  8. Identification of Low- and High-Impact Hemagglutinin Amino Acid Substitutions That Drive Antigenic Drift of Influenza A(H1N1) Viruses.

    PubMed

    Harvey, William T; Benton, Donald J; Gregory, Victoria; Hall, James P J; Daniels, Rodney S; Bedford, Trevor; Haydon, Daniel T; Hay, Alan J; McCauley, John W; Reeve, Richard

    2016-04-01

    Determining phenotype from genetic data is a fundamental challenge. Identification of emerging antigenic variants among circulating influenza viruses is critical to the vaccine virus selection process, with vaccine effectiveness maximized when constituents are antigenically similar to circulating viruses. Hemagglutination inhibition (HI) assay data are commonly used to assess influenza antigenicity. Here, sequence and 3-D structural information of hemagglutinin (HA) glycoproteins were analyzed together with corresponding HI assay data for former seasonal influenza A(H1N1) virus isolates (1997-2009) and reference viruses. The models developed identify and quantify the impact of eighteen amino acid substitutions on the antigenicity of HA, two of which were responsible for major transitions in antigenic phenotype. We used reverse genetics to demonstrate the causal effect on antigenicity for a subset of these substitutions. Information on the impact of substitutions allowed us to predict antigenic phenotypes of emerging viruses directly from HA gene sequence data and accuracy was doubled by including all substitutions causing antigenic changes over a model incorporating only the substitutions with the largest impact. The ability to quantify the phenotypic impact of specific amino acid substitutions should help refine emerging techniques that predict the evolution of virus populations from one year to the next, leading to stronger theoretical foundations for selection of candidate vaccine viruses. These techniques have great potential to be extended to other antigenically variable pathogens.

  9. Nicotinic Acid Adenine Dinucleotide Phosphate Analogs Substituted on the Nicotinic Acid and Adenine Ribosides. Effects on Receptor-Mediated Ca2+ release

    PubMed Central

    Trabbic, Christopher J.; Zhang, Fan; Walseth, Timothy F.; Slama, James T.

    2015-01-01

    Nicotinic acid adenine dinucleotide phosphate (NAADP) is a Ca2+ releasing intracellular second messenger in both mammals and echinoderms. We report that large functionalized substituents introduced at the nicotinic acid 5-position are recognized by the sea urchin receptor, albeit with a 20–500 fold loss in agonist potency. 5-(3-Azidopropyl)-NAADP was shown to release Ca2+ with an EC50 of 31 µM and to compete with NAADP for receptor binding with an IC50 of 56 nM. Attachment of charged groups to the nicotinic acid of NAADP is associated with loss of activity, suggesting that the nicotinate riboside moiety is recognized as a neutral zwitterion. Substituents (Br- and N3-) can be introduced at the 8-adenosyl position of NAADP while preserving high potency and agonist efficacy and an NAADP derivative substituted at both the 5-position of the nicotinic acid and at the 8-adenosyl position was also recognized although the agonist potency was significantly reduced. PMID:25826221

  10. Greater efficiency of photosynthetic carbon fixation due to single amino-acid substitution

    PubMed Central

    Paulus, Judith Katharina; Schlieper, Daniel; Groth, Georg

    2013-01-01

    The C4-photosynthetic carbon cycle is an elaborated addition to the classical C3-photosynthetic pathway, which improves solar conversion efficiency. The key enzyme in this pathway, phosphoenolpyruvate carboxylase, has evolved from an ancestral non-photosynthetic C3 phosphoenolpyruvate carboxylase. During evolution, C4 phosphoenolpyruvate carboxylase has increased its kinetic efficiency and reduced its sensitivity towards the feedback inhibitors malate and aspartate. An open question is the molecular basis of the shift in inhibitor tolerance. Here we show that a single-point mutation is sufficient to account for the drastic differences between the inhibitor tolerances of C3 and C4 phosphoenolpyruvate carboxylases. We solved high-resolution X-ray crystal structures of a C3 phosphoenolpyruvate carboxylase and a closely related C4 phosphoenolpyruvate carboxylase. The comparison of both structures revealed that Arg884 supports tight inhibitor binding in the C3-type enzyme. In the C4 phosphoenolpyruvate carboxylase isoform, this arginine is replaced by glycine. The substitution reduces inhibitor affinity and enables the enzyme to participate in the C4 photosynthesis pathway. PMID:23443546

  11. Single amino acid substitutions on the needle tip protein IpaD increased Shigella virulence.

    PubMed

    Meghraoui, Alaeddine; Schiavolin, Lionel; Allaoui, Abdelmounaaïm

    2014-07-01

    Infection of colonic epithelial cells by Shigella is associated with the type III secretion system, which serves as a molecular syringe to inject effectors into host cells. This system includes an extracellular needle used as a conduit for secreted proteins. Two of these proteins, IpaB and IpaD, dock at the needle tip to control secretion and are also involved in the insertion of a translocation pore into host cell membrane allowing effector delivery. To better understand the function of IpaD, we substituted thirteen residues conserved among homologous proteins in other bacterial species. Generated variants were tested for their ability to surface expose IpaB and IpaD, to control secretion, to insert the translocation pore, and to invade host cells. In addition to a first group of seven ipaD variants that behaved similarly to the wild-type strain, we identified a second group with mutations V314D and I319D that deregulated secretion of all effectors, but remained fully invasive. Moreover, we identified a third group with mutations Y153A, T161D, Q165L and Y276A, that exhibited increased levels of translocators secretion, pore formation, and cell entry. Altogether, our results offer a better understanding of the role of IpaD in the control of Shigella virulence.

  12. Pleiotropic effects of hemagglutinin amino acid substitutions of H5 influenza escape mutants

    SciTech Connect

    Rudneva, Irina A.; Timofeeva, Tatiana A.; Ignatieva, Anna V.; Shilov, Aleksandr A.; Krylov, Petr S.; Ilyushina, Natalia A.; Kaverin, Nikolai V.

    2013-12-15

    In the present study we assessed pleiotropic characteristics of the antibody-selected mutations. We examined pH optimum of fusion, temperatures of HA heat inactivation, and in vitro and in vivo replication kinetics of the previously obtained influenza H5 escape mutants. Our results showed that HA1 N142K mutation significantly lowered the pH of fusion optimum. Mutations of the escape mutants located in the HA lateral loop significantly affected H5 HA thermostability (P<0.05). HA changes at positions 131, 144, 145, and 156 and substitutions at positions 131, 142, 145, and 156 affected the replicative ability of H5 escape mutants in vitro and in vivo, respectively. Overall, a co-variation between antigenic specificity and different HA phenotypic properties has been demonstrated. We believe that the monitoring of pleiotropic effects of the HA mutations found in H5 escape mutants is essential for accurate prediction of mutants with pandemic potential. - Highlights: • HA1 N142K mutation significantly lowered the pH of fusion optimum. • Mutations located in the HA lateral loop significantly affected H5 HA thermostability. • HA changes at positions 131, 142, 144, 145, and 156 affected the replicative ability of H5 mutants. • Acquisition of glycosylation site could lead to the emergence of multiple pleiotropic effects.

  13. Canine distemper virus neutralization activity is low in human serum and it is sensitive to an amino acid substitution in the hemagglutinin protein.

    PubMed

    Zhang, Xinsheng; Wallace, Olivia L; Domi, Arban; Wright, Kevin J; Driscoll, Jonathan; Anzala, Omu; Sanders, Eduard J; Kamali, Anatoli; Karita, Etienne; Allen, Susan; Fast, Pat; Gilmour, Jill; Price, Matt A; Parks, Christopher L

    2015-08-01

    Serum was analyzed from 146 healthy adult volunteers in eastern Africa to evaluate measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb) prevalence and potency. MV plaque reduction neutralization test (PRNT) results indicated that all sera were positive for MV nAbs. Furthermore, the 50% neutralizing dose (ND50) for the majority of sera corresponded to antibody titers induced by MV vaccination. CDV nAbs titers were low and generally were detected in sera with high MV nAb titers. A mutant CDV was generated that was less sensitive to neutralization by human serum. The mutant virus genome had 10 nucleotide substitutions, which coded for single amino acid substitutions in the fusion (F) and hemagglutinin (H) glycoproteins and two substitutions in the large polymerase (L) protein. The H substitution occurred in a conserved region involved in receptor interactions among morbilliviruses, implying that this region is a target for cross-reactive neutralizing antibodies. PMID:25880113

  14. Canine distemper virus neutralization activity is low in human serum and it is sensitive to an amino acid substitution in the hemagglutinin protein.

    PubMed

    Zhang, Xinsheng; Wallace, Olivia L; Domi, Arban; Wright, Kevin J; Driscoll, Jonathan; Anzala, Omu; Sanders, Eduard J; Kamali, Anatoli; Karita, Etienne; Allen, Susan; Fast, Pat; Gilmour, Jill; Price, Matt A; Parks, Christopher L

    2015-08-01

    Serum was analyzed from 146 healthy adult volunteers in eastern Africa to evaluate measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb) prevalence and potency. MV plaque reduction neutralization test (PRNT) results indicated that all sera were positive for MV nAbs. Furthermore, the 50% neutralizing dose (ND50) for the majority of sera corresponded to antibody titers induced by MV vaccination. CDV nAbs titers were low and generally were detected in sera with high MV nAb titers. A mutant CDV was generated that was less sensitive to neutralization by human serum. The mutant virus genome had 10 nucleotide substitutions, which coded for single amino acid substitutions in the fusion (F) and hemagglutinin (H) glycoproteins and two substitutions in the large polymerase (L) protein. The H substitution occurred in a conserved region involved in receptor interactions among morbilliviruses, implying that this region is a target for cross-reactive neutralizing antibodies.

  15. Functional role of positively selected amino acid substitutions in mammalian rhodopsin evolution

    PubMed Central

    Fernández-Sampedro, Miguel A.; Invergo, Brandon M.; Ramon, Eva; Bertranpetit, Jaume; Garriga, Pere

    2016-01-01

    Visual rhodopsins are membrane proteins that function as light photoreceptors in the vertebrate retina. Specific amino acids have been positively selected in visual pigments during mammal evolution, which, as products of adaptive selection, would be at the base of important functional innovations. We have analyzed the top candidates for positive selection at the specific amino acids and the corresponding reverse changes (F13M, Q225R and A346S) in order to unravel the structural and functional consequences of these important sites in rhodopsin evolution. We have constructed, expressed and immunopurified the corresponding mutated pigments and analyzed their molecular phenotypes. We find that position 13 is very important for the folding of the receptor and also for proper protein glycosylation. Position 225 appears to be important for the function of the protein affecting the G-protein activation process, and position 346 would also regulate functionality of the receptor by enhancing G-protein activation and presumably affecting protein phosphorylation by rhodopsin kinase. Our results represent a link between the evolutionary analysis, which pinpoints the specific amino acid positions in the adaptive process, and the structural and functional analysis, closer to the phenotype, making biochemical sense of specific selected genetic sequences in rhodopsin evolution. PMID:26865329

  16. Nitrogen mineralization rates of the acidic, xeric soils of the New Jersey Pinelands: field rates

    SciTech Connect

    Poovarodom, S.; Tate, R.L. III; Bloom, R.A.

    1988-04-01

    Using the buried-bag procedure, the authors quantified nitrogen mineralization rates in the xeric, acidic Lakehurst, and Atsion sands of the New Jersey Pine Barrens. Average annual nitrogen yields in the upper 15 cm for the Lakehurst and the Atsion sands were 38.4 and 53.0 kg N/ha, corresponding to 4.5 and 2.5% of the total nitrogen, respectively. Net nitrogen mineralization in both soils exhibited distinct seasonal patterns with maxima in summer and minimum rates in the winter. Nitrification accounted for only 5% of the total N mineralized in both soils. This is consistent with the finding of low populations of autotrophic nitrifiers in these soils.

  17. Kinetic studies of ligand substitution rates for the Ru(NH/sub 3/)/sub 5/(H/sub 2/O)/sup 2 +/ ion in Nafion films

    SciTech Connect

    Lieber, C.M.; Schmidt, M.H.; Lewis, N.S.

    1986-10-01

    Substitution rates have been measured for reaction of a number of pyridines with the Nafion-bound Ru(NH/sub 3/)/sub 5/(H/sub 2/O)/sup 2 +/ complex. Reaction activities have been determined by electrochemical techniques, which also allow for determination of site thermodynamics and heterogeneity during the course of the reaction. Diffusion-coefficient effects are investigated by variation in polymer film thickness, and partition coefficients have been determined under equilibrium conditions by optical absorbance techniques. The partition-coefficient corrected rate law is found to be first order in Nafion-bound (Ru/sup II/) and first order in ligand concentration in the polymer phase. The partition-coefficient corrected bimolecular rate constants for a variety of pyridine ligands are found to vary by a factor of 5, which contrasts with the relatively constant substitution rates observed in aqueous solution. Also, sterically hindered ligands, such as 2-propylpyridine, exhibit surprisingly high substitution rate constants on the Nafion-bound Ru/sup II/ ion. These rate data indicate that pronounced molecular reactivity changes can occur upon electrode modification and have implications with respect to the design of chemically modified electrodes for use in electrocatalysis.

  18. WHIM syndrome caused by a single amino acid substitution in the carboxy-tail of chemokine receptor CXCR4

    PubMed Central

    Liu, Qian; Chen, Haoqian; Ojode, Teresa; Gao, Xiangxi; Anaya-O'Brien, Sandra; Turner, Nicholas A.; Ulrick, Jean; DeCastro, Rosamma; Kelly, Corin; Cardones, Adela R.; Gold, Stuart H.; Hwang, Eugene I.; Wechsler, Daniel S.; Malech, Harry L.; Murphy, Philip M.

    2012-01-01

    WHIM syndrome is a rare, autosomal dominant, immunodeficiency disorder so-named because it is characterized by warts, hypogammaglobulinemia, infections, and myelokathexis (defective neutrophil egress from the BM). Gain-of-function mutations that truncate the C-terminus of the chemokine receptor CXCR4 by 10-19 amino acids cause WHIM syndrome. We have identified a family with autosomal dominant inheritance of WHIM syndrome that is caused by a missense mutation in CXCR4, E343K (1027G → A). This mutation is also located in the C-terminal domain, a region responsible for negative regulation of the receptor. Accordingly, like CXCR4R334X, the most common truncation mutation in WHIM syndrome, CXCR4E343K mediated approximately 2-fold increased signaling in calcium flux and chemotaxis assays relative to wild-type CXCR4; however, CXCR4E343K had a reduced effect on blocking normal receptor down-regulation from the cell surface. Therefore, in addition to truncating mutations in the C-terminal domain of CXCR4, WHIM syndrome may be caused by a single charge-changing amino acid substitution in this domain, E343K, that results in increased receptor signaling. PMID:22596258

  19. Variable clinical manifestations of a glycine to glutamic acid substitution of the COL3A1 gene at residue 736

    SciTech Connect

    Pope, F.M.; Narcisi, P.; Richards, A.J.

    1994-09-01

    Glycine substitutions at the 3{prime} end of the COL3A1 gene generally produce a characteristic clinical phenotype including acrogeria and severe vascular fragility. Here we report a three generation British family in which the propositus presented with aneurysms of the groins. He, his mother, sister and elder daughter all had the external clinical phenotype of vascular EDS IV whilst another daughter and nephew were clinically normal. Cultured skin fibroblasts from the propositus and his clinically affected relatives poorly secreted normal and overmodified collagen III species. Normal components of secreted proteins predominated whilst overmodified molecules were prominent in intracellular material. Surprisingly the normal children also secreted less collagen type III than expected (though more than their clinically abnormal relatives). cDNA from bases 2671 to 3714 were amplified as four overlapping PCR fragments and analysed by DGGE. The region between 2671 and 3015 was heterozygous. Sequencing showed a mutation of glycine to glutamic acid at residue 736. This mutation created an extra Apa 1 restriction site which was suitable for family studies. These showed inheritance of the mutant gene by both vascular and non-vascular clinical phenotypes. This family therefore illustrates that replacement of glycine to glutamic acid at position 736 produces variable clinical and biochemical phenotypes ranging from easily recognizable vascular EDS IV with very poor collagen secretion to an EDS III-like picture and with less severe protein disturbance. The reasons for these differences are at present unexplained.

  20. Acceleration of protein folding by four orders of magnitude through a single amino acid substitution

    PubMed Central

    Roderer, Daniel J. A.; Schärer, Martin A.; Rubini, Marina; Glockshuber, Rudi

    2015-01-01

    Cis prolyl peptide bonds are conserved structural elements in numerous protein families, although their formation is energetically unfavorable, intrinsically slow and often rate-limiting for folding. Here we investigate the reasons underlying the conservation of the cis proline that is diagnostic for the fold of thioredoxin-like thiol-disulfide oxidoreductases. We show that replacement of the conserved cis proline in thioredoxin by alanine can accelerate spontaneous folding to the native, thermodynamically most stable state by more than four orders of magnitude. However, the resulting trans alanine bond leads to small structural rearrangements around the active site that impair the function of thioredoxin as catalyst of electron transfer reactions by more than 100-fold. Our data provide evidence for the absence of a strong evolutionary pressure to achieve intrinsically fast folding rates, which is most likely a consequence of proline isomerases and molecular chaperones that guarantee high in vivo folding rates and yields. PMID:26121966

  1. The nucleotide sequence of HLA-B{sup *}2704 reveals a new amino acid substitution in exon 4 which is also present in HLA-B{sup *}2706

    SciTech Connect

    Rudwaleit, M.; Bowness, P.; Wordsworth, P.

    1996-12-31

    The HLA-B27 subtype HLA-B{sup *}2704 is virtually absent in Caucasians but common in Orientals, where it is associated with ankylosing spondylitis. The amino acid sequence of HLA-B{sup *}2704 has been established by peptide mapping and was shown to differ by two amino acids from HLA-B{sup *}2705, HLA-B{sup *}2704 is characterized by a serine for aspartic acid substitution at position 77 and glutamic acid for valine at position 152. To date, however, no nucleotide sequence confirming these changes at the DNA level has been published. 13 refs., 2 figs.

  2. Destabilization of pea lectin by substitution of a single amino acid in a surface loop.

    PubMed

    Hoedemaeker, F J; van Eijsden, R R; Díaz, C L; de Pater, B S; Kijne, J W

    1993-09-01

    Legume lectins are considered to be antinutritional factors (ANF) in the animal feeding industry. Inactivation of ANF is an important element in processing of food. In our study on the stability of Pisum sativum L. lectin (PSL), a conserved hydrophobic amino acid (Val103) in a surface loop was replaced with alanine. The mutant lectin, PSL V103A, showed a decrease in unfolding temperature (Tm) by some 10 degrees C in comparison with wild-type (wt) PSL, and the denaturation energy (delta H) is only about 55% of that of wt PSL. Replacement of an adjacent amino acid (Phe104) with alanine did not result in a significant difference in stability in comparison with wt PSL. Both mutations did not change the sugar-binding properties of the lectin, as compared with wt PSL and with PSL from pea seeds, at ambient temperatures. The double mutant, PSL V103A/F104A, was produced in Escherichia coli, but could not be isolated in an active (i.e. sugar-binding) form. Interestingly, the mutation in PSL V103A reversibly affected sugar-binding at 37 degrees C, as judged from haemagglutination assays. These results open the possibility of production of lectins that are active in planta at ambient temperatures, but are inactive and possibly non-toxic at 37 degrees C in the intestines of mammals. PMID:8400124

  3. Substitution of a single amino acid (aspartic acid for histidine) converts the functional activity of human complement C4B to C4A

    SciTech Connect

    Carroll, M.C.; Fathallah, D.M.; Bergamaschini, L.; Alicot, E.M. ); Isenman, D.E. )

    1990-09-01

    The C4B isotype of the fourth component of human complement (C4) displays 3- to 4-fold greater hemolytic activity than does its other isotype C4A. This correlates with differences in their covalent binding efficiencies to erythrocytes coated with antibody and complement C1. C4A binds to a greater extent when C1 is on IgG immune aggregates. The differences in covalent binding properties correlate only with amino acid changes between residues 1101 and 1106 (pro-C4 numbering)-namely, Pro-1101, Cys-1102, Leu-1105, and Asp-1106 in C4A and Leu-1101, Ser-1102, Ile-1105, and His-1106 in C4B, which are located in the C4d region of the {alpha} chain. To more precisely identify the residues that are important for the functional differences, C4A-C4B hybrid proteins were constructed by using recombinant DNA techniques. Comparison of these by hemolytic assay and binding to IgG aggregates showed that the single substitution of aspartic acid for histidine at position 1106 largely accounted for the change in functional activity and nature of the chemical bond formed. Surprisingly, substitution of a neutral residue, alanine, for histidine at position 1106 resulted in an increase in binding to immune aggregates without subsequent reduction in the hemolytic activity. This result strongly suggests that position 1106 is not catalytic as previously proposed but interacts sterically/electrostatically with potential acceptor sites and serves to select binding sites on potential acceptor molecules.

  4. Lewis acid-promoted reactions of zirconacyclopentadienes with isocyanates. A one-pot three-component synthesis of multiply-substituted iminocyclopentadienes from one isocyanate and two alkynes.

    PubMed

    Lu, Jiang; Mao, Guoliang; Zhang, Wenxiong; Xi, Zhenfeng

    2005-10-14

    Multiply-substituted iminocyclopentadienes were formed from Lewis acid-promoted reactions of zirconacyclopentadienes and isocyanates via a one-pot three-component coupling process; the C=O double bond of the RN=C=O moiety in the isocyanate was cleaved, and the isocyanates behaved formally as a one-carbon unit with Lewis acid-dependent and substituent-dependent reactions being realized.

  5. Dipeptide Nanotubes Containing Unnatural Fluorine-Substituted β(2,3)-Diarylamino Acid and L-Alanine as Candidates for Biomedical Applications.

    PubMed

    Bonetti, Andrea; Pellegrino, Sara; Das, Priyadip; Yuran, Sivan; Bucci, Raffaella; Ferri, Nicola; Meneghetti, Fiorella; Castellano, Carlo; Reches, Meital; Gelmi, Maria Luisa

    2015-09-18

    The synthesis and the structural characterization of dipeptides composed of unnatural fluorine-substituted β(2,3)-diarylamino acid and L-alanine are reported. Depending on the stereochemistry of the β amino acid, these dipeptides are able to self-assemble into proteolytic stable nanotubes. These architectures were able to enter the cell and locate in the cytoplasmic/perinuclear region and represent interesting candidates for biomedical applications.

  6. Enhanced sensitivity to neutralizing antibodies in a variant of equine infectious anemia virus is linked to amino acid substitutions in the surface unit envelope glycoprotein.

    PubMed Central

    Cook, R F; Berger, S L; Rushlow, K E; McManus, J M; Cook, S J; Harrold, S; Raabe, M L; Montelaro, R C; Issel, C J

    1995-01-01

    Serial passage of the prototype (PR) cell-adapted Wyoming strain of equine infectious anemia virus (EIAV) in fetal donkey dermal (FDD) rather than fetal horse (designated fetal equine kidney [FEK]) cell cultures resulted in the generation of a variant virus strain which produced accelerated cytopathic effects in FDD cells and was 100- to 1,000-fold more sensitive to neutralizing antibodies than its parent. This neutralization-sensitive variant was designated the FDD strain. Although there were differences in glycosylation between the PR and FDD strains, passage of the FDD virus in FEK cells did not reduce its sensitivity to neutralizing antibody. Nucleotide sequencing of the region encoding the surface unit (SU) protein from the FDD strain revealed nine amino acid substitutions compared with the PR strain. Two of these substitutions resulted in changes in the polarity of charge, four caused the introduction of a charged residue, and three had no net change in charge. Nucleotide sequence analysis was extended to the region of the FDD virus genome encoding the extracellular domain of the transmembrane envelope glycoprotein (TM). Unlike the situation with the FDD virus coding region, there were minor variations in nucleotide sequence between individual molecular clones containing this region of the TM gene. Although each clone contained three nucleotide substitutions compared with the PR strain, only one of these was common to all, and this did not affect the amino acid content. Of the remaining two nucleotide substitutions, only one resulted in an amino acid change, and in each case, this change appeared to be conservative. To determine if amino acid substitutions in the SU protein of FDD cell-grown viruses were responsible for the enhanced sensitivity to neutralizing antibodies, chimeric viruses were constructed by using an infectious molecular clone of EIAV. These chimeric viruses contained all of the amino acid substitutions found in the FDD virus strain and were

  7. Oxidation of substituted phenols in the environment: A QSAR analysis of rate constants for reaction with singlet oxygen. [Quantitative Structure-Activity Relationship

    SciTech Connect

    Tratnyek, P.G.; Holgne, J. , Duebendorf )

    1991-09-01

    Substituted phenols can be oxidized by singlet oxygen ({sup 1}O{sub 2}), which is formed in sunlit surface waters, and it has been suggested that this reaction may contribute to the environmental fate of phenolic substances. In aqueous solution, the observed rate of phenol disappearance is due to reaction of both the phenolate anion and the undissociated phenol. In order to quantify the effect of substituents on the rates of these reactions, second-order rate constants have been measured for both species for 22 substituted phenols by use of a model system containing the sensitizer rose bengal. Correlation analysis based on half-wave oxidation potentials, E{sub 1/2}, and on {sigma} constants reveals significant quantitative structure-activity relationships (QSARs) for both the undissociated phenols and the phenolate anions. Ortho- and multisubstituted phenols have been included in the correlations. These QSARs are consistent with the rate-limiting formation of a precursor complex with a small amount of charge-transfer character and can be used to predict additional rate constants for a wide range of environmentally significant substituted phenols.

  8. Molecular distribution of amino acid substitutions on neuraminidase from the 2009 (H1N1) human influenza pandemic virus.

    PubMed

    Quiliano, Miguelmiguel; Valdivia-Olarte, Hugo; Olivares, Carlos; Requena, David; Gutiérrez, Andrés H; Reyes-Loyola, Paola; Tolentino-Lopez, Luis E; Sheen, Patricia; Briz, Verónica; Muñoz-Fernández, Maria A; Correa-Basurto, José; Zimic, Mirko

    2013-01-01

    The pandemic influenza AH1N1 (2009) caused an outbreak of human infection that spread to the world. Neuraminidase (NA) is an antigenic surface glycoprotein, which is essential to the influenza infection process, and is the target of anti-flu drugs oseltamivir and zanamivir. Currently, NA inhibitors are the pillar pharmacological strategy against seasonal and global influenza. Although mutations observed after NA-inhibitor treatment are characterized by changes in conserved amino acids of the enzyme catalytic site, it is possible that specific amino acid substitutions (AASs) distant from the active site such as H274Y, could confer oseltamivir or zanamivir resistance. To better understand the molecular distribution pattern of NA AASs, we analyzed NA AASs from all available reported pandemic AH1N1 NA sequences, including those reported from America, Africa, Asia, Europe, Oceania, and specifically from Mexico. The molecular distributions of the AASs were obtained at the secondary structure domain level for both the active and catalytic sites, and compared between geographic regions. Our results showed that NA AASs from America, Asia, Europe, Oceania and Mexico followed similar molecular distribution patterns. The compiled data of this study showed that highly conserved amino acids from the NA active site and catalytic site are indeed being affected by mutations. The reported NA AASs follow a similar molecular distribution pattern worldwide. Although most AASs are distributed distantly from the active site, this study shows the emergence of mutations affecting the previously conserved active and catalytic site. A significant number of unique AASs were reported simultaneously on different continents.

  9. Molecular distribution of amino acid substitutions on neuraminidase from the 2009 (H1N1) human influenza pandemic virus

    PubMed Central

    Quiliano, MiguelMiguel; Valdivia-Olarte, Hugo; Olivares, Carlos; Requena, David; Gutiérrez, Andrés H; Reyes-Loyola, Paola; Tolentino-Lopez, Luis E; Sheen, Patricia; Briz, Verónica; Muñoz-Fernández, Maria A; Correa-Basurto, José; Zimic, Mirko

    2013-01-01

    The pandemic influenza AH1N1 (2009) caused an outbreak of human infection that spread to the world. Neuraminidase (NA) is an antigenic surface glycoprotein, which is essential to the influenza infection process, and is the target of anti-flu drugs oseltamivir and zanamivir. Currently, NA inhibitors are the pillar pharmacological strategy against seasonal and global influenza. Although mutations observed after NA-inhibitor treatment are characterized by changes in conserved amino acids of the enzyme catalytic site, it is possible that specific amino acid substitutions (AASs) distant from the active site such as H274Y, could confer oseltamivir or zanamivir resistance. To better understand the molecular distribution pattern of NA AASs, we analyzed NA AASs from all available reported pandemic AH1N1 NA sequences, including those reported from America, Africa, Asia, Europe, Oceania, and specifically from Mexico. The molecular distributions of the AASs were obtained at the secondary structure domain level for both the active and catalytic sites, and compared between geographic regions. Our results showed that NA AASs from America, Asia, Europe, Oceania and Mexico followed similar molecular distribution patterns. The compiled data of this study showed that highly conserved amino acids from the NA active site and catalytic site are indeed being affected by mutations. The reported NA AASs follow a similar molecular distribution pattern worldwide. Although most AASs are distributed distantly from the active site, this study shows the emergence of mutations affecting the previously conserved active and catalytic site. A significant number of unique AASs were reported simultaneously on different continents. PMID:23930018

  10. Double D-π-A dye linked by 2,2'-bipyridine dicarboxylic acid: influence of para- and meta-substituted carboxyl anchoring group.

    PubMed

    Ganesan, Paramaguru; Chandiran, Aravind Kumar; Gao, Peng; Rajalingam, Renganathan; Grätzel, Michael; Nazeeruddin, Mohammad Khaja

    2015-04-01

    Starting from 2,2'-bipyridine dicarboxylic acid, two new (D-π-A)2 sensitizers, including m-DA with the carboxyl anchoring group substituted meta to the donor-bridge moiety and p-DA with a para-substituted anchoring group, were synthesized in order to evaluate the impact of the position of the anchoring group on the optical, electrochemical, and photovoltaic properties of dye-sensitized solar cells. p-DA exhibits red-shifted absorption behavior compared to m-DA, owing to the more efficiently extended π-conjugation with para substitution. Both m-DA and p-DA are adsorbed on the mesoporous TiO2 surface by using both of their carboxylic acid groups in a bianchoring mode, which is confirmed through attenuated total reflectance FTIR analysis. Red-shifted absorption of p-DA assists the achievement of a red-shifted incident photon-to-electron conversion efficiency and a higher short-circuit current density than m-DA. The photogenerated electron lifetime in TiO2 is also found to be higher for para substituted p-DA than the meta-substituted m-DA, which results in a higher open-circuit voltage. All of the results suggest that dicarboxyl-2,2'-bipyridine can be used as an acceptor for metal-free organic sensitizers. However, the anchoring segments should be adjusted to the favorable position of the corresponding donor-bridge moieties for better conjugation.

  11. Controlling the Photoreactivity of the Photoactive Yellow Protein Chromophore by Substituting at the p-Coumaric Acid Group

    PubMed Central

    2011-01-01

    We have performed ab initio CASSCF, CASPT2, and EOM-CCSD calculations on doubly deprotonated p-coumaric acid (pCA2−), the chromophore precursor of the photoactive yellow protein. The results of the calculations demonstrate that pCA2− can undergo only photoisomerization of the double bond. In contrast, the chromophore derivative with the acid replaced by a ketone (p-hydroxybenzylidene acetone, pCK−) undergoes both single- and double-bond photoisomerization, with the single-bond relaxation channel more favorable than the double-bond channel. The substitution alters the nature of the first excited states and the associated potential energy landscape. The calculations show that the electronic nature of the first two (π,π*) excited states are interchanged in vacuo due to the substitution. In pCK−, the first excited state is a charge-transfer (CT π,π*) state, in which the negative charge has migrated from the phenolate ring onto the alkene tail of the chromophore, whereas the locally excited (LE π,π*) state, in which the excitation involves the orbitals on the phenol ring, lies higher in energy and is the fourth excited state. In pCA2−, the CT state is higher in energy due the presence of a negative charge on the tail of the chromophore, and the first excited state is the LE state. In isolated pCA2−, there is a 68 kJ/mol barrier for double-bond photoisomerization on the potential energy surface of this LE state. In water, however, hydrogen bonding with water molecules reduces this barrier to 9 kJ/mol. The barrier separates the local trans minimum near the Franck−Condon region from the global minimum on the excited-state potential energy surface. The lowest energy conical intersection was located near this minimum. In contrast to pCK−, single-bond isomerization is highly unfavorable both in the LE and CT states of pCA2−. These results demonstrate that pCA2− can only decay efficiently in water and exclusively by double-bond photoisomerization

  12. Antimicrobial effect of para-alkoxyphenylcarbamic acid esters containing substituted N-phenylpiperazine moiety

    PubMed Central

    Malík, Ivan; Bukovský, Marián; Andriamainty, Fils; Gališinová, Jana

    2013-01-01

    In current research, nine basic esters of para-alkoxyphenylcarbamic acid with incorporated 4-(4-fluoro-/3-trifluoromethylphenyl)piperazin-1-yl fragment, 6i–6m and 8f–8i, were screened for their in vitro antimicrobial activity against Candida albicans, Staphylococcus aureus and Escherichia coli, respectively. Taking into account the minimum inhibitory concentration assay (MIC), as the most active against given yeast was evaluated 8i (MIC = 0.20 mg/mL), the most lipophilic structure containing para-butoxy and trifluoromethyl substituents. Investigating the efficiency of the compounds bearing only a single atom of fluorine and appropriate para-alkoxy side chain against Candida albicans, the cut-off effect was observed. From evaluated homological series, the maximum of the effectiveness was noticed for the stucture 6 k (MIC = 0.39 mg/mL), containing para-propoxy group attached to phenylcarbamoyloxy fragment, beyond which the compounds ceased to be active. On the contrary, all the tested molecules were against Staphylococcus aureus and Escherichia coli (MICs > 1.00 mg/mL) practically inactive. PMID:24294237

  13. L-dehydroascorbic acid can substitute l-ascorbic acid as dietary vitamin C source in guinea pigs.

    PubMed

    Frikke-Schmidt, Henriette; Tveden-Nyborg, Pernille; Lykkesfeldt, Jens

    2016-04-01

    Vitamin C deficiency globally affects several hundred million people and has been associated with increased morbidity and mortality in numerous studies. In this study, bioavailability of the oxidized form of vitamin C (l-dehydroascorbic acid or DHA)-commonly found in vitamin C containing food products prone to oxidation-was studied. Our aim was to compare tissue accumulation of vitamin C in guinea pigs receiving different oral doses of either ascorbate or DHA. In all tissues tested (plasma, liver, spleen, lung, adrenal glands, kidney, muscle, heart, and brain), only sporadic differences in vitamin C accumulation from ascorbate or DHA were observed except for the lowest dose of DHA (0.25mg/ml in the drinking water), where approximately half of the tissues had slightly yet significantly less vitamin C accumulation than from the ascorbate source. As these results contradicted data from rats, we continued to explore the ability to recycle DHA in blood, liver and intestine in guinea pigs, rats and mice. These investigations revealed that guinea pigs have similar recycling capacity in red blood cells as observed in humans, while rats and mice do not have near the same ability to reduce DHA in erythrocytes. In liver and intestinal homogenates, guinea pigs also showed a significantly higher ability to recycle DHA compared to rats and mice. These data demonstrate that DHA in guinea pigs-as in humans-is almost as effective as ascorbate as vitamin C source when it comes to taking up and storing vitamin C and further suggest that the guinea pig is superior to other rodents in modeling human vitamin C homeostasis.

  14. l-dehydroascorbic acid can substitute l-ascorbic acid as dietary vitamin C source in guinea pigs

    PubMed Central

    Frikke-Schmidt, Henriette; Tveden-Nyborg, Pernille; Lykkesfeldt, Jens

    2015-01-01

    Vitamin C deficiency globally affects several hundred million people and has been associated with increased morbidity and mortality in numerous studies. In this study, bioavailability of the oxidized form of vitamin C (l-dehydroascorbic acid or DHA)—commonly found in vitamin C containing food products prone to oxidation—was studied. Our aim was to compare tissue accumulation of vitamin C in guinea pigs receiving different oral doses of either ascorbate or DHA. In all tissues tested (plasma, liver, spleen, lung, adrenal glands, kidney, muscle, heart, and brain), only sporadic differences in vitamin C accumulation from ascorbate or DHA were observed except for the lowest dose of DHA (0.25 mg/ml in the drinking water), where approximately half of the tissues had slightly yet significantly less vitamin C accumulation than from the ascorbate source. As these results contradicted data from rats, we continued to explore the ability to recycle DHA in blood, liver and intestine in guinea pigs, rats and mice. These investigations revealed that guinea pigs have similar recycling capacity in red blood cells as observed in humans, while rats and mice do not have near the same ability to reduce DHA in erythrocytes. In liver and intestinal homogenates, guinea pigs also showed a significantly higher ability to recycle DHA compared to rats and mice. These data demonstrate that DHA in guinea pigs—as in humans—is almost as effective as ascorbate as vitamin C source when it comes to taking up and storing vitamin C and further suggest that the guinea pig is superior to other rodents in modeling human vitamin C homeostasis. PMID:26609560

  15. A Single Amino Acid Substitution in an ORANGE Protein Promotes Carotenoid Overaccumulation in Arabidopsis.

    PubMed

    Yuan, Hui; Owsiany, Katherine; Sheeja, T E; Zhou, Xiangjun; Rodriguez, Caroline; Li, Yongxi; Welsch, Ralf; Chayut, Noam; Yang, Yong; Thannhauser, Theodore W; Parthasarathy, Mandayam V; Xu, Qiang; Deng, Xiuxin; Fei, Zhangjun; Schaffer, Ari; Katzir, Nurit; Burger, Joseph; Tadmor, Yaakov; Li, Li

    2015-09-01

    Carotenoids are crucial for plant growth and human health. The finding of ORANGE (OR) protein as a pivotal regulator of carotenogenesis offers a unique opportunity to comprehensively understand the regulatory mechanisms of carotenoid accumulation and develop crops with enhanced nutritional quality. Here, we demonstrated that alteration of a single amino acid in a wild-type OR greatly enhanced its ability to promote carotenoid accumulation. Whereas overexpression of OR from Arabidopsis (Arabidopsis thaliana; AtOR) or from the agronomically important crop sorghum (Sorghum bicolor; SbOR) increased carotenoid levels up to 2-fold, expression of AtOR(His) (R90H) or SbOR(His) (R104H) variants dramatically enhanced carotenoid accumulation by up to 7-fold in the Arabidopsis calli. Moreover, we found that AtOR(Ala) (R90A) functioned similarly to AtOR(His) to promote carotenoid overproduction. Neither AtOR nor AtOR(His) greatly affected carotenogenic gene expression. AtOR(His) exhibited similar interactions with phytoene synthase (PSY) as AtOR in posttranscriptionally regulating PSY protein abundance. AtOR(His) triggered biogenesis of membranous chromoplasts in the Arabidopsis calli, which shared structures similar to chromoplasts found in the curd of the orange cauliflower (Brassica oleracea) mutant. By contrast, AtOR did not cause plastid-type changes in comparison with the controls, but produced plastids containing larger and electron-dense plastoglobuli. The unique ability of AtOR(His) in mediating chromoplast biogenesis is responsible for its induced carotenoid overproduction. Our study demonstrates OR(His/Ala) as powerful tools for carotenoid enrichment in plants, and provides insights into the mechanisms underlying OR(His)-regulated carotenoid accumulation.

  16. Comparisons of Pollen Substitute Diets for Honey bees: Consumprion Rates by Colonies and Effects on Brood and Adult Populations

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Commercially available pollen substitute diets for honey bees (Apis mellifera L.) were evaluated for consumption and colony growth (brood and adult populations) and compared with pollen cake and high fructose corn syrup (HFCS). Two trials were conducted; the first for 4 months during the fall and wi...

  17. Comparisons of pollen substitute diets for honey bees: consumption rates by colonies and effects on brood and adult populations.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Commercially available pollen substitute diets for honey bees (Apis mellifera L.) were evaluated for consumption and colony growth (brood and adult populations) and compared with pollen cake and high fructose corn syrup (HFCS). Two trials were conducted; the first for 3 months during the fall and w...

  18. Preparation of a nitro-substituted tris(indolyl)methane modified silica in deep eutectic solvents for solid-phase extraction of organic acids.

    PubMed

    Wang, Na; Wang, Jiamin; Liao, Yuan; Shao, Shijun

    2016-05-01

    A new sorbent for solid-phase extraction was synthesized by chemical immobilization of nitro-substituted tris(indolyl)methane on silica in new and green deep eutectic solvents. Elemental analysis results indicated that deep eutectic solvents could be an alternative to the traditional solvents in preparing nitro-substituted tris(indolyl)methane modified silica. Coupled with high performance liquid chromatography, the extraction performance of the sorbent was evaluated by using four organic acids as model analytes. The rebinding experiments results showed that the nitro-substituted tris(indolyl)methane modified silica sorbent had a good adsorption capacity towards the selected organic acids. Under the appropriate experimental conditions, good precision and wide linear ranges with coefficient of determination (R(2)) of higher than 0.9957 were obtained, and the limits of detection were in the range of 0.50-2.0μgL(-1) for the organic acids tested. The developed solid-phase extraction-high performance liquid chromatography-diode array detection (SPE-HPLC-DAD) method was successfully applied for the determination of organic acids in two drinking samples with recoveries ranging from 76.7% to 110.0% and 67.7% to 104.0% for all the selected organic acids, respectively.

  19. Hydrogenation of imines catalyzed by trisphosphine-substituted molybdenum and tungsten nitrosyl hydrides and co-catalytic acid.

    PubMed

    Chakraborty, Subrata; Blacque, Olivier; Fox, Thomas; Berke, Heinz

    2014-10-01

    Hydride complexes Mo,W(CO)(NO)H(mer-etp(i)p) (iPr2PCH2CH2)2PPh=etp(i)p) (2 a,b(syn), syn and anti of NO and Ph(etp(i)p) orientions) were prepared and probed in imine hydrogenations together with co-catalytic [H(Et2O)2][B(C6F5)4] (140 °C, 60 bar H2). 2 a,b(syn) were obtained via reduction of syn/anti-Mo,W(NO)Cl3(mer-etp(i)p) and syn,anti-Mo,W(NO)(CO)Cl(mer-etp(i)p). [H(Et2O)2][B(C6F5)4] in THF converted the hydrides into THF complexes syn-[Mo,W(NO)(CO)(etp(i)p)(THF)][B(C6F5)4]. Combinations of the p-substituents of aryl imines p-R(1)C6H4CH=N-p-C6H4R(2) (R(1),R(2)=H,F,Cl,OMe,α-Np) were hydrogenated to amines (maximum initial TOFs of 1960 h(-1) (2 a(syn)) and 740 h(-1) (2 b(syn)) for N-(4-methoxybenzylidene)aniline). An 'ionic hydrogenation' mechanism based on linear Hammett plots (ρ=-10.5, p-substitution on the C-side and ρ=0.86, p-substitution on the N-side), iminium intermediates, linear P(H2) dependence, and DKIE=1.38 is proposed. Heterolytic splitting of H2 followed by 'proton before hydride' transfers are the steps in the ionic mechanism where H2 ligand addition is rate limiting.

  20. Synthesis of [.sup.13C] and [.sup.2H] substituted methacrylic acid, [.sup.13C] and [.sup.2H] substituted methyl methacrylate and/or related compounds

    SciTech Connect

    Alvarez, Marc A.; Martinez, Rodolfo A.; Unkefer, Clifford J.

    2010-02-16

    The present invention is directed to labeled compounds of the formulae ##STR00001## wherein Q is selected from the group consisting of --S(.dbd.O)--, and --S(.dbd.O).sub.2--, Z is selected from the group consisting of 1-naphthyl, substituted 1-naphthyl, 2-naphthyl, substituted 2-naphthyl, and phenyl groups with the structure ##STR00002## wherein R.sub.1, R.sub.2, R.sub.3, R.sub.4 and R.sub.5 are each independently selected from the group consisting of hydrogen, a C.sub.1-C.sub.4 lower alkyl, a halogen, and an amino group selected from the group consisting of NH.sub.2, NHR and NRR' where R and R' are each independently selected from the group consisting of a C.sub.1-C.sub.4 lower alkyl, an aryl, and an alkoxy group, and X is selected from the group consisting of hydrogen, a C.sub.1-C.sub.4 lower alkyl group, and a fully-deuterated C.sub.1-C.sub.4 lower alkyl group.

  1. Identification of Amino Acid Substitutions with Compensational Effects in the Attachment Protein of Canine Distemper Virus

    PubMed Central

    Sattler, Ursula; Khosravi, Mojtaba; Avila, Mislay; Pilo, Paola; Langedijk, Johannes P.; Ader-Ebert, Nadine; Alves, Lisa A.; Plattet, Philippe

    2014-01-01

    ABSTRACT The hemagglutinin (H) gene of canine distemper virus (CDV) encodes the receptor-binding protein. This protein, together with the fusion (F) protein, is pivotal for infectivity since it contributes to the fusion of the viral envelope with the host cell membrane. Of the two receptors currently known for CDV (nectin-4 and the signaling lymphocyte activation molecule [SLAM]), SLAM is considered the most relevant for host susceptibility. To investigate how evolution might have impacted the host-CDV interaction, we examined the functional properties of a series of missense single nucleotide polymorphisms (SNPs) naturally accumulating within the H-gene sequences during the transition between two distinct but related strains. The two strains, a wild-type strain and a consensus strain, were part of a single continental outbreak in European wildlife and occurred in distinct geographical areas 2 years apart. The deduced amino acid sequence of the two H genes differed at 5 residues. A panel of mutants carrying all the combinations of the SNPs was obtained by site-directed mutagenesis. The selected mutant, wild type, and consensus H proteins were functionally evaluated according to their surface expression, SLAM binding, fusion protein interaction, and cell fusion efficiencies. The results highlight that the most detrimental functional effects are associated with specific sets of SNPs. Strikingly, an efficient compensational system driven by additional SNPs appears to come into play, virtually neutralizing the negative functional effects. This system seems to contribute to the maintenance of the tightly regulated function of the H-gene-encoded attachment protein. IMPORTANCE To investigate how evolution might have impacted the host-canine distemper virus (CDV) interaction, we examined the functional properties of naturally occurring single nucleotide polymorphisms (SNPs) in the hemagglutinin gene of two related but distinct strains of CDV. The hemagglutinin gene encodes

  2. Identification of amino acid substitutions associated with neutralization phenotype in the human immunodeficiency virus type-1 subtype C gp120

    PubMed Central

    Kirchherr, Jennifer L; Hamilton, Jennifer; Lu, Xiaozhi; Gnanakaran, S; Muldoon, Mark; Daniels, Marcus; Kasongo, Webster; Chalwe, Victor; Mulenga, Chanda; Mwananyanda, Lawrence; Musonda, Rosemary M; Yuan, Xing; Montefiori, David C; Korber, Bette T; Haynes, Barton F; Gao, Feng

    2010-01-01

    Neutralizing antibodies (Nabs) are thought to play an important role in prevention and control of HIV-1 infection and should be targeted by an AIDS vaccine. It is critical to understand how HIV-1 induces Nabs by analyzing viral sequences in both tested viruses and sera. Neutralization susceptibility to antibodies in autologous and heterologous plasma was determined for multiple Envs (3–6) from each of 15 subtype C infected-individuals. Heterologous neutralization was divided into two distinct groups: plasma with strong, cross-reactive neutralization (N=9) and plasma with weak neutralization (N=6). Plasma with cross-reactive heterologous Nabs also more potently neutralized contemporaneous autologous viruses. Analysis of Env sequences in plasma from both groups revealed a three-amino acid substitution pattern in the V4 region that was associated with greater neutralization potency and breadth. Identification of such potential neutralization signatures may have important implications for the development of HIV-1 vaccines capable of inducing Nabs to subtype C HIV-1. PMID:21036380

  3. Improvement of the reverse tetracycline transactivator by single amino acid substitutions that reduce leaky target gene expression to undetectable levels

    PubMed Central

    Roney, Ian J.; Rudner, Adam D.; Couture, Jean-François; Kærn, Mads

    2016-01-01

    Conditional gene expression systems that enable inducible and reversible transcriptional control are essential research tools and have broad applications in biomedicine and biotechnology. The reverse tetracycline transcriptional activator is a canonical system for engineered gene expression control that enables graded and gratuitous modulation of target gene transcription in eukaryotes from yeast to human cell lines and transgenic animals. However, the system has a tendency to activate transcription even in the absence of tetracycline and this leaky target gene expression impedes its use. Here, we identify single amino-acid substitutions that greatly enhance the dynamic range of the system in yeast by reducing leaky transcription to undetectable levels while retaining high expression capacity in the presence of inducer. While the mutations increase the inducer concentration required for full induction, additional sensitivity-enhancing mutations can compensate for this effect and confer a high degree of robustness to the system. The novel transactivator variants will be useful in applications where tight and tunable regulation of gene expression is paramount. PMID:27323850

  4. Two single amino acid substitutions in the intervening region of Newcastle disease virus HN protein attenuate viral replication and pathogenicity

    PubMed Central

    Liu, Bin; Ji, Yanhong; Lin, Zhongqing; Fu, Yuguang; Muhammad Dafallah, Rihab; Zhu, Qiyun

    2015-01-01

    Among the proteins encoded by Newcastle disease virus (NDV), the attachment protein (HN) is an important determinant of virulence and pathogenicity. HN has been molecularly characterized at the protein level; however, the relationship between the molecular character of HN and the animal pathotype it causes has not been well explored. Here, we revisited the intervening region (IR) of the HN stalk and extended the known biological functions of HN. Three distinct substitutions (A89Q, P93A, and L94A) in the IR of genotype VII NDV (G7 strain) HN protein were analyzed. The A89Q and L94A mutations weakened the fusion promotion activity of HN to 44% and 41% of that of wild type, respectively, whereas P93A decreased the neuraminidase activity to 21% of the parental level. At the virus level, P93A and L94A-bearing viruses displayed impaired receptor recognition ability, neuraminidase activity, and fusion-promoting activity, all of which led to virus attenuation. In addition, the L94A-mutated virus showed a dramatic decline in replication and was attenuated in cells and in chickens. Our data demonstrate that the HN biological activities and functions modulated by these specific amino acids in the IR are associated with NDV replication and pathogenicity. PMID:26267791

  5. One-Tube-Only Standardized Site-Directed Mutagenesis: An Alternative Approach to Generate Amino Acid Substitution Collections

    PubMed Central

    Mingo, Janire; Erramuzpe, Asier; Luna, Sandra; Aurtenetxe, Olaia; Amo, Laura; Diez, Ibai; Schepens, Jan T. G.; Hendriks, Wiljan J. A. J.; Cortés, Jesús M.; Pulido, Rafael

    2016-01-01

    Site-directed mutagenesis (SDM) is a powerful tool to create defined collections of protein variants for experimental and clinical purposes, but effectiveness is compromised when a large number of mutations is required. We present here a one-tube-only standardized SDM approach that generates comprehensive collections of amino acid substitution variants, including scanning- and single site-multiple mutations. The approach combines unified mutagenic primer design with the mixing of multiple distinct primer pairs and/or plasmid templates to increase the yield of a single inverse-PCR mutagenesis reaction. Also, a user-friendly program for automatic design of standardized primers for Ala-scanning mutagenesis is made available. Experimental results were compared with a modeling approach together with stochastic simulation data. For single site-multiple mutagenesis purposes and for simultaneous mutagenesis in different plasmid backgrounds, combination of primer sets and/or plasmid templates in a single reaction tube yielded the distinct mutations in a stochastic fashion. For scanning mutagenesis, we found that a combination of overlapping primer sets in a single PCR reaction allowed the yield of different individual mutations, although this yield did not necessarily follow a stochastic trend. Double mutants were generated when the overlap of primer pairs was below 60%. Our results illustrate that one-tube-only SDM effectively reduces the number of reactions required in large-scale mutagenesis strategies, facilitating the generation of comprehensive collections of protein variants suitable for functional analysis. PMID:27548698

  6. One-Tube-Only Standardized Site-Directed Mutagenesis: An Alternative Approach to Generate Amino Acid Substitution Collections.

    PubMed

    Mingo, Janire; Erramuzpe, Asier; Luna, Sandra; Aurtenetxe, Olaia; Amo, Laura; Diez, Ibai; Schepens, Jan T G; Hendriks, Wiljan J A J; Cortés, Jesús M; Pulido, Rafael

    2016-01-01

    Site-directed mutagenesis (SDM) is a powerful tool to create defined collections of protein variants for experimental and clinical purposes, but effectiveness is compromised when a large number of mutations is required. We present here a one-tube-only standardized SDM approach that generates comprehensive collections of amino acid substitution variants, including scanning- and single site-multiple mutations. The approach combines unified mutagenic primer design with the mixing of multiple distinct primer pairs and/or plasmid templates to increase the yield of a single inverse-PCR mutagenesis reaction. Also, a user-friendly program for automatic design of standardized primers for Ala-scanning mutagenesis is made available. Experimental results were compared with a modeling approach together with stochastic simulation data. For single site-multiple mutagenesis purposes and for simultaneous mutagenesis in different plasmid backgrounds, combination of primer sets and/or plasmid templates in a single reaction tube yielded the distinct mutations in a stochastic fashion. For scanning mutagenesis, we found that a combination of overlapping primer sets in a single PCR reaction allowed the yield of different individual mutations, although this yield did not necessarily follow a stochastic trend. Double mutants were generated when the overlap of primer pairs was below 60%. Our results illustrate that one-tube-only SDM effectively reduces the number of reactions required in large-scale mutagenesis strategies, facilitating the generation of comprehensive collections of protein variants suitable for functional analysis. PMID:27548698

  7. Designing inhibitors of cytochrome c/cardiolipin peroxidase complexes: mitochondria-targeted imidazole-substituted fatty acids.

    PubMed

    Jiang, Jianfei; Bakan, Ahmet; Kapralov, Alexandr A; Silva, K Ishara; Huang, Zhentai; Amoscato, Andrew A; Peterson, James; Garapati, Venkata Krishna; Saxena, Sunil; Bayir, Hülya; Atkinson, Jeffrey; Bahar, Ivet; Kagan, Valerian E

    2014-06-01

    Mitochondria have emerged as the major regulatory platform responsible for the coordination of numerous metabolic reactions as well as cell death processes, whereby the execution of intrinsic apoptosis includes the production of reactive oxygen species fueling oxidation of cardiolipin (CL) catalyzed by cytochrome (Cyt) c. As this oxidation occurs within the peroxidase complex of Cyt c with CL, the latter represents a promising target for the discovery and design of drugs with antiapoptotic mechanisms of action. In this work, we designed and synthesized a new group of mitochondria-targeted imidazole-substituted analogs of stearic acid TPP-n-ISAs with various positions of the attached imidazole group on the fatty acid (n = 6, 8, 10, 13, and 14). By using a combination of absorption spectroscopy and EPR protocols (continuous wave electron paramagnetic resonance and electron spin echo envelope modulation) we demonstrated that TPP-n-ISAs indeed were able to potently suppress CL-induced structural rearrangements in Cyt c, paving the way to its peroxidase competence. TPP-n-ISA analogs preserved the low-spin hexa-coordinated heme-iron state in Cyt c/CL complexes whereby TPP-6-ISA displayed a significantly more effective preservation pattern than TPP-14-ISA. Elucidation of these intermolecular stabilization mechanisms of Cyt c identified TPP-6-ISA as an effective inhibitor of the peroxidase function of Cyt c/CL complexes with a significant antiapoptotic potential realized in mouse embryonic cells exposed to ionizing irradiation. These experimental findings were detailed and supported by all-atom molecular dynamics simulations. Based on the experimental data and computation predictions, we identified TPP-6-ISA as a candidate drug with optimized antiapoptotic potency.

  8. Designing Inhibitors of Cytochrome c/Cardiolipin Peroxidase Complexes: Mitochondria-Targeted Imidazole-Substituted Fatty Acids

    PubMed Central

    Jiang, Jianfei; Bakan, Ahmet; Kapralov, Alexandr A.; Silva, K. Ishara; Huang, Zhentai; Amoscato, Andrew A.; Peterson, James; Garapati, Venkata Krishna; Saxena, Sunil; Bayir, Hülya; Atkinson, Jeffrey; Bahar, Ivet; Kagan, Valerian E.

    2014-01-01

    Mitochondria have emerged as the major regulatory platform responsible for coordination of numerous metabolic reactions as well as cell death processes, whereby the execution of intrinsic apoptosis includes the production of reactive oxygen species fueling oxidation of cardiolipin (CL) catalyzed by cytochrome (cyt) c. As this oxidation occurs within the peroxidase complex of cyt c with CL, the latter represents a promising target for the discovery and design of drugs with anti-apoptotic mechanism of action. In this work, we designed and synthesized a new group of mitochondria-targeted imidazole-substituted analogues of stearic acid TPP-n-ISA with different positions of the attached imidazole group on the fatty acid (n=6, 8, 10, 13 and 14). By using a combination of absorption spectroscopy and EPR protocols (continuous wave electron paramagnetic resonance, and electron spin echo envelope modulation) we demonstrated that TPP-n-ISA indeed were able to potently suppress CL induced structural re-arrangements in cyt c paving the way to its peroxidase competence. TPP-n-ISA analogues preserved the low spin hexa-coordinated heme iron state in cyt c/CL complexes whereby TPP-6-ISA displayed a significantly more effective preservation pattern than TPP-14-ISA. Elucidation of these intermolecular stabilization mechanisms of cyt c identified TPP-6-ISA as an effective inhibitor of the peroxidase function of cyt c/CL complexes with a significant anti-apoptotic potential realized in mouse embryonic cells exposed to ionizing irradiation. These experimental findings were detailed and supported by all atom molecular dynamics simulations. Based on the experimental data and computations predictions, we identified TPP-6-ISA as a candidate drug with optimized anti-apoptotic potency. PMID:24631490

  9. Requirement for Asn298 on D1 protein for oxygen evolution: analyses by exhaustive amino acid substitution in the green alga Chlamydomonas reinhardtii.

    PubMed

    Kuroda, Hiroshi; Kodama, Natsumi; Sun, Xiao-Yu; Ozawa, Shin-ichiro; Takahashi, Yuichiro

    2014-07-01

    PSII generates strong oxidants used for water oxidation. The secondary electron donor, Y(Z), is Tyr161 on PSII reaction center D1 protein and mediates electron transfer from the oxygen-evolving Mn(4)CaO(5) cluster to the primary electron donor, P680. The latest PSII crystal structure revealed the presence of a hydrogen bond network around Y(Z), which is anticipated to play important roles in the electron and proton transfer reactions. Y(Z) forms a hydrogen bond with His190 which in turn forms a hydrogen bond with Asn298 on D1 protein. Although functional roles of Y(Z) and His190 have already been characterized, little is known about the functional role of Asn298. Here we have generated 19 mutants from a green alga Chlamydomonas reinhardtii, in which the Asn298 has been substituted by each of the other 19 amino acid residues. All mutants showed significantly impaired or no photosynthetic growth. Seven mutants capable of photosynthetic growth showed oxygen-evolving activity although at a significantly reduced rate. Interestingly the oxygen-evolving activity of these mutants was markedly photosensitive. The 19 mutants accumulated PSII at variable levels and showed a light-induced electron transfer reaction from 1,5-diphenylcarbazide (DPC) to 2,6-dichlorophenolindophenol (DCIP), suggesting that Asn298 is important for the function and photoprotection of the Mn(4)CaO(5) cluster. PMID:24853102

  10. The same substitution, glutamic acid----lysine at position 501, occurs in three alloalbumins of Asiatic origin: albumins Vancouver, Birmingham, and Adana.

    PubMed Central

    Huss, K; Madison, J; Ishioka, N; Takahashi, N; Arai, K; Putnam, F W

    1988-01-01

    A strategy is described for identifying structural changes in genetic variants of human serum albumin (alloalbumins). By use of this strategy we have determined an amino acid substitution in three alloalbumins of Asiatic origin. The same amino acid exchange, glutamic acid----lysine at position 501, occurs in albumins Vancouver and Birmingham, both from families that migrated from northern India, and also in albumin Adana from Turkey. This exchange corresponds to a single base mutation in the codon GAG to AAG and accords with the slow mobility of the three albumins at pH 8.6. Each of the three alloalbumins had been reported to be a new variant, yet they have the same substitution. These results emphasize the need for structural study of genetic variants that have been differentiated only by nonspecific physical criteria such as dye binding and electrophoretic mobility. We know of no other description of the substitution involved in an alloalbumin originating from the Indian subcontinent. However, the same change of glutamic acid----lysine at position 501 may be present in several other named variants reported for populations in north India and the surrounding regions. Images PMID:2901102

  11. Acid-base chemical reaction model for nucleation rates in the polluted atmospheric boundary layer.

    PubMed

    Chen, Modi; Titcombe, Mari; Jiang, Jingkun; Jen, Coty; Kuang, Chongai; Fischer, Marc L; Eisele, Fred L; Siepmann, J Ilja; Hanson, David R; Zhao, Jun; McMurry, Peter H

    2012-11-13

    Climate models show that particles formed by nucleation can affect cloud cover and, therefore, the earth's radiation budget. Measurements worldwide show that nucleation rates in the atmospheric boundary layer are positively correlated with concentrations of sulfuric acid vapor. However, current nucleation theories do not correctly predict either the observed nucleation rates or their functional dependence on sulfuric acid concentrations. This paper develops an alternative approach for modeling nucleation rates, based on a sequence of acid-base reactions. The model uses empirical estimates of sulfuric acid evaporation rates obtained from new measurements of neutral molecular clusters. The model predicts that nucleation rates equal the sulfuric acid vapor collision rate times a prefactor that is less than unity and that depends on the concentrations of basic gaseous compounds and preexisting particles. Predicted nucleation rates and their dependence on sulfuric acid vapor concentrations are in reasonable agreement with measurements from Mexico City and Atlanta. PMID:23091030

  12. Acid-base chemical reaction model for nucleation rates in the polluted atmospheric boundary layer.

    PubMed

    Chen, Modi; Titcombe, Mari; Jiang, Jingkun; Jen, Coty; Kuang, Chongai; Fischer, Marc L; Eisele, Fred L; Siepmann, J Ilja; Hanson, David R; Zhao, Jun; McMurry, Peter H

    2012-11-13

    Climate models show that particles formed by nucleation can affect cloud cover and, therefore, the earth's radiation budget. Measurements worldwide show that nucleation rates in the atmospheric boundary layer are positively correlated with concentrations of sulfuric acid vapor. However, current nucleation theories do not correctly predict either the observed nucleation rates or their functional dependence on sulfuric acid concentrations. This paper develops an alternative approach for modeling nucleation rates, based on a sequence of acid-base reactions. The model uses empirical estimates of sulfuric acid evaporation rates obtained from new measurements of neutral molecular clusters. The model predicts that nucleation rates equal the sulfuric acid vapor collision rate times a prefactor that is less than unity and that depends on the concentrations of basic gaseous compounds and preexisting particles. Predicted nucleation rates and their dependence on sulfuric acid vapor concentrations are in reasonable agreement with measurements from Mexico City and Atlanta.

  13. Additivity of the Stabilization Effect of Single Amino Acid Substitutions in Triple Mutants of Recombinant Formate Dehydrogenase from the Soybean Glycine max.

    PubMed

    Alekseeva, A A; Kargov, I S; Kleimenov, S Yu; Savin, S S; Tishkov, V I

    2015-01-01

    Recently, we demonstrated that the amino acid substitutions Ala267Met and Ala267Met/Ile272Val (Alekseeva et al., Biochemistry, 2012), Phe290Asp, Phe290Asn and Phe290Ser (Alekseeva et al., Prot. Eng. Des. Select, 2012) in recombinant formate dehydrogenase from soya Glycine max (SoyFDH) lead to a significant (up to 30-100 times) increase in the thermal stability of the enzyme. The substitutions Phe290Asp, Phe290Asn and Phe290Ser were introduced into double mutant SoyFDH Ala267Met/Ile272Val by site-directed mutagenesis. Combinations of three substitutions did not lead to a noticeable change in the catalytic properties of the mutant enzymes. The stability of the resultant triple mutants was studied through thermal inactivation kinetics and differential scanning calorimetry. The thermal stability of the new mutant SoyFDHs was shown to be much higher than that of their precursors. The stability of the best mutant SoyFDH Ala267Met/Ile272Val/Phe290Asp turned out to be comparable to that of the most stable wild-type formate dehydrogenases from other sources. The results obtained with both methods indicate a great synergistic contribution of individual amino acid substitutions to the common stabilization effect.

  14. Pentahaloethane-based chlorofluorocarbon substitutes and halothane: correlation of in vivo hepatic protein trifluoroacetylation and urinary trifluoroacetic acid excretion with calculated enthalpies of activation.

    PubMed

    Harris, J W; Jones, J P; Martin, J L; LaRosa, A C; Olson, M J; Pohl, L R; Anders, M W

    1992-01-01

    The hydrochlorofluorocarbons (HCFCs) 2,2-dichloro-1,1,1-trifluoroethane (HCFC-123) and 2-chloro-1,1,1,2-tetrafluoroethane (HCFC-124) and the hydrofluorocarbon (HFC) pentafluoroethane (HFC-125) are being developed as substitutes for chlorofluorocarbons that deplete stratospheric ozone. The structural similarity of these HCFCs and HFCs to halothane, which is hepatotoxic under certain circumstances, indicates that the metabolism and cellular interactions of HCFCs and HFCs must be explored. In a previous study [Harris et al. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 1407], similar patterns of trifluoroacetylated proteins (TFA-proteins) were detected by immunoblotting with anti-TFA-protein antibodies in livers of rats exposed to halothane or HCFC-123. The present study extends these results and demonstrates that in vivo TFA-protein formation resulting from a 6-h exposure to a 1% atmosphere of these compounds follows the trend: halothane approximately HCFC-123 much greater than HFC-124, greater than HFC-125. The calculated enthalpies of activation of halothane, HCFC-123, HCFC-124, and HFC-125 paralleled the observed rate of trifluoroacetic acid excretion in HCFC- or HFC-exposed rats. Exposure of rats to a range of HCFC-123 concentrations indicated that TFA-protein formation was saturated at an exposure concentration between 0.01% and 0.1% HCFC-123. Deuteration of HCFC-123 decreased TFA-protein formation in vivo. Urinary trifluoroacetic acid excretion by treated rats correlated with the levels of TFA-proteins found after each of these treatments. No TFA-proteins were detected in hepatic fractions from rats given 1,1,1,2-tetrafluoroethane (HFC-134a), which is not metabolized to a trifluoroacetyl halide.(ABSTRACT TRUNCATED AT 250 WORDS)

  15. Effect of Oral Sebacic Acid on Postprandial Glycemia, Insulinemia, and Glucose Rate of Appearance in Type 2 Diabetes

    PubMed Central

    Iaconelli, Amerigo; Gastaldelli, Amalia; Chiellini, Chiara; Gniuli, Donatella; Favuzzi, Angela; Binnert, Christophe; Macé, Katherine; Mingrone, Geltrude

    2010-01-01

    OBJECTIVE Dicarboxylic acids are natural products with the potential of being an alternate dietary source of energy. We aimed to evaluate the effect of sebacic acid (a 10-carbon dicarboxylic acid; C10) ingestion on postprandial glycemia and glucose rate of appearance (Ra) in healthy and type 2 diabetic subjects. Furthermore, the effect of C10 on insulin-mediated glucose uptake and on GLUT4 expression was assessed in L6 muscle cells in vitro. RESEARCH DESIGN AND METHODS Subjects ingested a mixed meal (50% carbohydrates, 15% proteins, and 35% lipids) containing 0 g (control) or 10 g C10 in addition to the meal or 23 g C10 as a substitute of fats. RESULTS In type 2 diabetic subjects, the incremental glucose area under the curve (AUC) decreased by 42% (P < 0.05) and 70% (P < 0.05) in the 10 g C10 and 23 g C10 groups, respectively. At the largest amounts used, C10 reduced the glucose AUC in healthy volunteers also. When fats were substituted with 23 g C10, AUC of Ra was significantly reduced on the order of 18% (P < 0.05) in both healthy and diabetic subjects. The insulin-dependent glucose uptake by L6 cells was increased in the presence of C10 (38.7 ± 10.3 vs. 11.4 ± 5.4%; P = 0.026). This increase was associated with a 1.7-fold raise of GLUT4. CONCLUSIONS Sebacic acid significantly reduced hyperglycemia after a meal in type 2 diabetic subjects. This beneficial effect was associated with a reduction in glucose Ra, probably due to lowered hepatic glucose output and increased peripheral glucose disposal. PMID:20724647

  16. A Computational Study of Cytotoxicity of Substituted Amides of Pyrazine- 2-carboxylic acids Using QSAR and DFT Based Molecular Surface Electrostatic Potential

    PubMed Central

    Hosseini, Sharieh; Monajjemi, Majid; Rajaeian, Elahe; Haghgu, Mohammad; Salari, Aliakbar; Gholami, Mohammad Reza

    2013-01-01

    Pyrazine derivatives are important class of compounds with diverse biological and cytotoxic activities and clinical applications. In this study, B3 p 86 / 6 – 31 + + G * was used to compute and map the molecular surface electrostatic potentials of a group of substituted amides of pyrazine-2-carboxylic acids to identify common features related to their subsequent cytotoxicities. Several statistical properties including potentials extrema (Vs ,min,Vs ,max), the average of positive electrostatic potential on the surface (Vs+), the average of V(r) over the surface (Vs) and the Lowest Unoccupied Molecular Orbital (LUMO) and system cytotoxicities were computed. Statistically, the most significant correlation is a five -parameter equation with correlation coefficient, R² values of 0.922 and R²adj = 0.879. The obtained models allowed us to reveal cytotoxic activity of substituted amides of Pyrazine2- carboxcylic acid. PMID:24523754

  17. A review of molecular-clock calibrations and substitution rates in liverworts, mosses, and hornworts, and a timeframe for a taxonomically cleaned-up genus Nothoceros.

    PubMed

    Villarreal, Juan Carlos; Renner, Susanne S

    2014-09-01

    Absolute times from calibrated DNA phylogenies can be used to infer lineage diversification, the origin of new ecological niches, or the role of long distance dispersal in shaping current distribution patterns. Molecular-clock dating of non-vascular plants, however, has lagged behind flowering plant and animal dating. Here, we review dating studies that have focused on bryophytes with several goals in mind, (i) to facilitate cross-validation by comparing rates and times obtained so far; (ii) to summarize rates that have yielded plausible results and that could be used in future studies; and (iii) to calibrate a species-level phylogeny for Nothoceros, a model for plastid genome evolution in hornworts. Including the present work, there have been 18 molecular clock studies of liverworts, mosses, or hornworts, the majority with fossil calibrations, a few with geological calibrations or dated with previously published plastid substitution rates. Over half the studies cross-validated inferred divergence times by using alternative calibration approaches. Plastid substitution rates inferred for "bryophytes" are in line with those found in angiosperm studies, implying that bryophyte clock models can be calibrated either with published substitution rates or with fossils, with the two approaches testing and cross-validating each other. Our phylogeny of Nothoceros is based on 44 accessions representing all suspected species and a matrix of six markers of nuclear, plastid, and mitochondrial DNA. The results show that Nothoceros comprises 10 species, nine in the Americas and one in New Zealand (N. giganteus), with the divergence between the New Zealand species and its Chilean sister species dated to the Miocene and therefore due to long-distance dispersal. Based on the new tree, we formally transfer two species of Megaceros into Nothoceros, resulting in the new combinations N. minarum (Nees) J.C. Villarreal and N. schizophyllus (Gottsche ex Steph.) J.C. Villarreal, and we also

  18. Hydroxyapatite-calcium sulfate-hyaluronic acid composite encapsulated with collagenase as bone substitute for alveolar bone regeneration.

    PubMed

    Subramaniam, Sadhasivam; Fang, Yen-Hsin; Sivasubramanian, Savitha; Lin, Feng-Huei; Lin, Chun-pin

    2016-01-01

    Periodontitis is a very severe inflammatory condition of the periodontium that progressively damages the soft tissue and destroys the alveolar bone that supports the teeth. The bone loss is naturally irreversible because of limited reparability of the teeth. Advancement in tissue engineering provides an effective regeneration of osseous defects with suitable dental implants or tissue-engineered constructs. This study reports a hydroxyapatite, calcium sulfate hemihydrate and hyaluronic acid laden collagenase (HAP/CS/HA-Col) as a bone substitute for the alveolar bone regeneration. The composite material was mechanically tested and the biocompatibility was evaluated by WST-1 assay. The in vivo bone formation was assessed in rat with alveolar bone defects and the bone augmentation by the HAP/CS/HA-Col composite was confirmed by micro-CT images and histological examination. The mechanical strength of 6.69 MPa with excellent biocompatibility was obtained for the HAP/CS/HA-Col composite. The collagenase release profile had facilitated the acceleration of bone remodeling process and it was confirmed by the findings of micro-CT and H&E staining. The bone defects implanted with HAP/CS/HA composite containing 2 mg/mL type I collagenase have shown improved new bone formation with matured bone morphology in comparison with the HAP/CS/HA composite that lacks the collagenase and the porous hydroxyapatite (p-HAP) granules. The said findings demonstrated that the collagenase inclusion in HAP/CS/HA composite is a feasible approach for the alveolar bone regeneration and the same design can also be applied to other defective tissues.

  19. Hydroxyapatite-calcium sulfate-hyaluronic acid composite encapsulated with collagenase as bone substitute for alveolar bone regeneration.

    PubMed

    Subramaniam, Sadhasivam; Fang, Yen-Hsin; Sivasubramanian, Savitha; Lin, Feng-Huei; Lin, Chun-pin

    2016-01-01

    Periodontitis is a very severe inflammatory condition of the periodontium that progressively damages the soft tissue and destroys the alveolar bone that supports the teeth. The bone loss is naturally irreversible because of limited reparability of the teeth. Advancement in tissue engineering provides an effective regeneration of osseous defects with suitable dental implants or tissue-engineered constructs. This study reports a hydroxyapatite, calcium sulfate hemihydrate and hyaluronic acid laden collagenase (HAP/CS/HA-Col) as a bone substitute for the alveolar bone regeneration. The composite material was mechanically tested and the biocompatibility was evaluated by WST-1 assay. The in vivo bone formation was assessed in rat with alveolar bone defects and the bone augmentation by the HAP/CS/HA-Col composite was confirmed by micro-CT images and histological examination. The mechanical strength of 6.69 MPa with excellent biocompatibility was obtained for the HAP/CS/HA-Col composite. The collagenase release profile had facilitated the acceleration of bone remodeling process and it was confirmed by the findings of micro-CT and H&E staining. The bone defects implanted with HAP/CS/HA composite containing 2 mg/mL type I collagenase have shown improved new bone formation with matured bone morphology in comparison with the HAP/CS/HA composite that lacks the collagenase and the porous hydroxyapatite (p-HAP) granules. The said findings demonstrated that the collagenase inclusion in HAP/CS/HA composite is a feasible approach for the alveolar bone regeneration and the same design can also be applied to other defective tissues. PMID:26454048

  20. Carbapenem resistance in cystic fibrosis strains of Pseudomonas aeruginosa as a result of amino acid substitutions in porin OprD.

    PubMed

    Richardot, Charlotte; Plésiat, Patrick; Fournier, Damien; Monlezun, Laura; Broutin, Isabelle; Llanes, Catherine

    2015-05-01

    The aim of this work was to investigate the impact of single amino acid substitutions occurring in specific porin OprD on carbapenem resistance of cystic fibrosis (CF) strains of Pseudomonas aeruginosa. A PAO1ΔoprD mutant was complemented with the oprD genes from five carbapenem-resistant CF strains exhibiting very low amounts of mutated OprD porins in their outer membrane despite wild-type levels of oprD transcripts. Compared with wild-type porin from strain PAO1, single amino acid substitutions S403P (in periplasmic loop 8), Y242H, S278P and L345P (in β-sheets 10, 12 and 14, respectively) were found to result in reduced amounts of OprD in the outer membrane, increased carbapenem resistance, and slower growth in minimal medium containing gluconate, an OprD substrate, as the sole source of carbon and energy. This indicates that in CF strains of P. aeruginosa, loss of porin OprD may not only result from mutations downregulating the expression of or disrupting the oprD gene, but also from mutations generating deleterious amino acid substitutions in the porin structure.

  1. The role of local and remote amino acid substitutions for optimizing fluorescence in bacteriophytochromes: A case study on iRFP

    PubMed Central

    Buhrke, David; Velazquez Escobar, Francisco; Sauthof, Luisa; Wilkening, Svea; Herder, Nico; Tavraz, Neslihan N.; Willoweit, Mario; Keidel, Anke; Utesch, Tillmann; Mroginski, Maria-Andrea; Schmitt, Franz-Josef; Hildebrandt, Peter; Friedrich, Thomas

    2016-01-01

    Bacteriophytochromes are promising tools for tissue microscopy and imaging due to their fluorescence in the near-infrared region. These applications require optimization of the originally low fluorescence quantum yields via genetic engineering. Factors that favour fluorescence over other non-radiative excited state decay channels are yet poorly understood. In this work we employed resonance Raman and fluorescence spectroscopy to analyse the consequences of multiple amino acid substitutions on fluorescence of the iRFP713 benchmark protein. Two groups of mutations distinguishing iRFP from its precursor, the PAS-GAF domain of the bacteriophytochrome P2 from Rhodopseudomonas palustris, have qualitatively different effects on the biliverdin cofactor, which exists in a fluorescent (state II) and a non-fluorescent conformer (state I). Substitution of three critical amino acids in the chromophore binding pocket increases the intrinsic fluorescence quantum yield of state II from 1.7 to 5.0% due to slight structural changes of the tetrapyrrole chromophore. Whereas these changes are accompanied by an enrichment of state II from ~40 to ~50%, a major shift to ~88% is achieved by remote amino acid substitutions. Additionally, an increase of the intrinsic fluorescence quantum yield of this conformer by ~34% is achieved. The present results have important implications for future design strategies of biofluorophores. PMID:27329837

  2. The role of local and remote amino acid substitutions for optimizing fluorescence in bacteriophytochromes: A case study on iRFP.

    PubMed

    Buhrke, David; Velazquez Escobar, Francisco; Sauthof, Luisa; Wilkening, Svea; Herder, Nico; Tavraz, Neslihan N; Willoweit, Mario; Keidel, Anke; Utesch, Tillmann; Mroginski, Maria-Andrea; Schmitt, Franz-Josef; Hildebrandt, Peter; Friedrich, Thomas

    2016-01-01

    Bacteriophytochromes are promising tools for tissue microscopy and imaging due to their fluorescence in the near-infrared region. These applications require optimization of the originally low fluorescence quantum yields via genetic engineering. Factors that favour fluorescence over other non-radiative excited state decay channels are yet poorly understood. In this work we employed resonance Raman and fluorescence spectroscopy to analyse the consequences of multiple amino acid substitutions on fluorescence of the iRFP713 benchmark protein. Two groups of mutations distinguishing iRFP from its precursor, the PAS-GAF domain of the bacteriophytochrome P2 from Rhodopseudomonas palustris, have qualitatively different effects on the biliverdin cofactor, which exists in a fluorescent (state II) and a non-fluorescent conformer (state I). Substitution of three critical amino acids in the chromophore binding pocket increases the intrinsic fluorescence quantum yield of state II from 1.7 to 5.0% due to slight structural changes of the tetrapyrrole chromophore. Whereas these changes are accompanied by an enrichment of state II from ~40 to ~50%, a major shift to ~88% is achieved by remote amino acid substitutions. Additionally, an increase of the intrinsic fluorescence quantum yield of this conformer by ~34% is achieved. The present results have important implications for future design strategies of biofluorophores. PMID:27329837

  3. Amino acid substitutions in malate dehydrogenases of piezophilic bacteria isolated from intestinal contents of deep-sea fishes retrieved from the abyssal zone.

    PubMed

    Saito, Rie; Kato, Chiaki; Nakayama, Akihiko

    2006-02-01

    To examine the occurrence in other deep-sea bacteria of two amino acid substitutions (Ala-180 and His-229) in malate dehydrogenase (MDH) found previously in the deep-sea piezophilic Moritella sp. strain 2D2, we cloned and sequenced MDH genes of deep-sea piezophilic Moritella and Shewanella strains isolated from intestinal contents of deep-sea fishes, as well as other Moritella species from deep-sea water and sediments: M. marina, M. japonica, and M. yayanosii. The piezophilic Moritella strains had a Val residue or an Ala residue at position 180 and all the Moritella strains except for one had a His residue at position 229. However, four piezophilic-strain-specific substitutions at positions 103, 111, 229, and 283 were found to be completely conserved in the MDH of the intestinal Moritella strains of deep-sea fishes, indicating the substitutions may be habitat-specific. The piezophilic Shewanella strains had a Val residue and a Gln residue at positions 180 and 229, respectively. However, the MDHs of the Shewanella strains had five piezophilic-strain-specific substitutions at positions 61, 65, 107, 161, and 202. Therefore, the enzymatic strategies for responding to deep-sea high pressure environments of the MDHs between the genera Moritella and Shewanella are potentially different. Moreover, homology modeling shows these substitutions found in the MDHs of both genera except for position 229 in the subunit interface are located on the exposed region of the MDH molecules, indicating the substitutions may be related to the hydration state of the molecules. PMID:16598154

  4. Sensory evaluation ratings and melting characteristics show that okra gum is an acceptable milk-fat ingredient substitute in chocolate frozen dairy dessert.

    PubMed

    Romanchik-Cerpovicz, Joelle E; Costantino, Amanda C; Gunn, Laura H

    2006-04-01

    Reducing dietary fat intake may lower the risk of developing coronary heart disease. This study examined the feasibility of substituting okra gum for 25%, 50%, 75%, or 100% milk fat in frozen chocolate dairy dessert. Fifty-six consumers evaluated the frozen dairy desserts using a hedonic scale. Consumers rated color, smell, texture, flavor, aftertaste, and overall acceptability characteristics of all products as acceptable. All ratings were similar among the products except for the aftertaste rating, which was significantly lower for chocolate frozen dairy dessert containing 100% milk-fat replacement with okra gum compared with the control (0% milk-fat replacement) (P<0.05). Whereas melting points of all products were similar, melting rates slowed significantly as milk-fat replacement with okra gum increased, suggesting that okra gum may increase the stability of frozen dairy desserts (P<0.05). Overall, this study shows that okra gum is an acceptable milk-fat ingredient substitute in chocolate frozen dairy dessert.

  5. Sulfonic acid functionalized nano-γ-Al2O3: a new, efficient, and reusable catalyst for synthesis of 3-substituted-2H-1,4-benzothiazines.

    PubMed

    Li, Wei Lin; Tian, Shuan Bao; Zhu, Feng

    2013-01-01

    A simple and efficient synthetic protocol has been developed for the synthesis of 3-substituted-2H-1,4-benzothiazines by using a novel sulfonic acid functionalized nano-γ-Al2O3 catalyst, devoid of corrosive acidic, and basic reagents. The developed method has the advantages of good to excellent yields, short reaction times, operational simplicity, and a recyclable catalyst. The catalyst can be prepared by a simple procedure from inexpensive and readily available nano-γ-Al2O3 and has been shown to be recoverable and reusable up to six cycles without any loss of activity.

  6. Asymmetric synthesis of highly substituted β-lactones through oxidative carbene catalysis with LiCl as cooperative Lewis acid.

    PubMed

    Bera, Srikrishna; Samanta, Ramesh C; Daniliuc, Constantin G; Studer, Armido

    2014-09-01

    The reaction of enals with β-diketones, β-ketoesters, and malonates bearing a β-oxyalkyl substituent at the α-position by oxidative NHC catalysis to provide highly substituted β-lactones is described. Reactions occur with excellent diastereo- and enantioselectivity. The organo cascade comprises two CC bond formations and one CO bond formation. Up to four contiguous stereogenic centers including two fully substituted stereocenters are formed in the cascade.

  7. Comparison of three chiral stationary phases with respect to their enantio- and diastereoselectivity for cyclic beta-substituted alpha-amino acids.

    PubMed

    Schlauch, Michael; Kos, Olha; Frahm, August W

    2002-01-15

    Three chiral stationary phases were examined for the enantio- and diastereoseparation of cycloaliphatic beta-substituted alpha-quaternary alpha-amino acids. Resolution of diastereomeric analytes is feasible with a chiral crown ether based column, whereas the separation of enantiomers, except for one pair of amino acids, could not be achieved. The two chiral stationary phases with the glycopeptide antibiotic teicoplanin and with the copper(II)-D-penicillamine complex, respectively, are, however, both very potent in the separation of the enantiomeric, as well as of the diastereomeric amino acids. A baseline separation of all four stereoisomeric forms in one chromatographic run was possible with the exception of one type of amino acid. The results of the method development are presented in this paper.

  8. Canine distemper virus neutralization activity is low in human serum and it is sensitive to an amino acid substitution in the hemagglutinin protein

    SciTech Connect

    Zhang, Xinsheng; Wallace, Olivia L.; Domi, Arban; Wright, Kevin J.; Driscoll, Jonathan; Anzala, Omu; Sanders, Eduard J.; Kamali, Anatoli; Allen, Susan; Fast, Pat; Gilmour, Jill; Price, Matt A.; Parks, Christopher L.

    2015-08-15

    Serum was analyzed from 146 healthy adult volunteers in eastern Africa to evaluate measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb) prevalence and potency. MV plaque reduction neutralization test (PRNT) results indicated that all sera were positive for MV nAbs. Furthermore, the 50% neutralizing dose (ND50) for the majority of sera corresponded to antibody titers induced by MV vaccination. CDV nAbs titers were low and generally were detected in sera with high MV nAb titers. A mutant CDV was generated that was less sensitive to neutralization by human serum. The mutant virus genome had 10 nucleotide substitutions, which coded for single amino acid substitutions in the fusion (F) and hemagglutinin (H) glycoproteins and two substitutions in the large polymerase (L) protein. The H substitution occurred in a conserved region involved in receptor interactions among morbilliviruses, implying that this region is a target for cross-reactive neutralizing antibodies. - Highlights: • Screened 146 serum samples for measles virus (MV) and canine distemper virus (CDV) neutralizing antibody (nAb). • MV nAb is prevalent in the sera. • CDV neutralizing activity is generally low or absent and when detected it is present in sera with high MV nAb titers. • A neutralization-resistant CDV mutant was isolated using human serum selection. • A mutation was identified in the receptor-binding region of CDV hemagglutinin protein that confers the neutralization resistance.

  9. Determination of small halogenated carboxylic acid residues in drug substances by high performance liquid chromatography-diode array detection following derivatization with nitro-substituted phenylhydrazines.

    PubMed

    Hou, Desheng; Fan, Jingjing; Han, Lingfei; Ruan, Xiaoling; Feng, Feng; Liu, Wenyuan; Zheng, Feng

    2016-03-18

    A method for the determination of small halogenated carboxylic acid (HCA) residues in drug substances is urgently needed because of the potential of HCAs for genotoxicity and carcinogenicity in humans. We have now developed a simple method, involving derivatization followed by high performance liquid chromatography-diode array detection (HPLC-DAD), for the determination of six likely residual HCAs (monochloroacetic acid, monobromoacetic acid, dichloroacetic acid, 2-chloropropionic acid, 2-bromopropionic acid and 3-chloropropionic acid) in drug substances. Different nitro-substituted phenylhydrazines (NPHs) derivatization reagents were systematically compared and evaluated. 2-Nitrophenylhydrazine hydrochloride (2-NPH·HCl) was selected as the most suitable choice since its derivatives absorb strongly at 392 nm, a region of the spectrum where most drug substances and impurities absorb very weakly. During the derivatization process, the commonly used catalyst, pyridine, caused rapid dechlorination or chlorine substitution of α-halogenated derivatives. To avoid these unwanted side reactions, a reliable derivatization method that did not use pyridine was developed. Reaction with 2-NPH·HCl using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride as coupling agent in acetonitrile-water (70:30) at room temperature for 2h gave complete reaction and avoided degradation products. The derivatives were analyzed, without any pretreatment, using gradient HPLC with detection in the near visible region. Organic acids commonly found in drug substances and other impurities did not interfere with the analysis. Good linearity (r>0.999) and low limits of quantitation (0.05-0.12 μg mL(-1)) were obtained. The mean recoveries were in the range of 80-115% with RSD <5.81% except for 3-CPA in ibuprofen which was 78.5%. The intra- and inter-day precisions were expressed as RSD <1.98% and <4.39%, respectively. Finally, the proposed method was successfully used for the residue

  10. Determination of small halogenated carboxylic acid residues in drug substances by high performance liquid chromatography-diode array detection following derivatization with nitro-substituted phenylhydrazines.

    PubMed

    Hou, Desheng; Fan, Jingjing; Han, Lingfei; Ruan, Xiaoling; Feng, Feng; Liu, Wenyuan; Zheng, Feng

    2016-03-18

    A method for the determination of small halogenated carboxylic acid (HCA) residues in drug substances is urgently needed because of the potential of HCAs for genotoxicity and carcinogenicity in humans. We have now developed a simple method, involving derivatization followed by high performance liquid chromatography-diode array detection (HPLC-DAD), for the determination of six likely residual HCAs (monochloroacetic acid, monobromoacetic acid, dichloroacetic acid, 2-chloropropionic acid, 2-bromopropionic acid and 3-chloropropionic acid) in drug substances. Different nitro-substituted phenylhydrazines (NPHs) derivatization reagents were systematically compared and evaluated. 2-Nitrophenylhydrazine hydrochloride (2-NPH·HCl) was selected as the most suitable choice since its derivatives absorb strongly at 392 nm, a region of the spectrum where most drug substances and impurities absorb very weakly. During the derivatization process, the commonly used catalyst, pyridine, caused rapid dechlorination or chlorine substitution of α-halogenated derivatives. To avoid these unwanted side reactions, a reliable derivatization method that did not use pyridine was developed. Reaction with 2-NPH·HCl using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride as coupling agent in acetonitrile-water (70:30) at room temperature for 2h gave complete reaction and avoided degradation products. The derivatives were analyzed, without any pretreatment, using gradient HPLC with detection in the near visible region. Organic acids commonly found in drug substances and other impurities did not interfere with the analysis. Good linearity (r>0.999) and low limits of quantitation (0.05-0.12 μg mL(-1)) were obtained. The mean recoveries were in the range of 80-115% with RSD <5.81% except for 3-CPA in ibuprofen which was 78.5%. The intra- and inter-day precisions were expressed as RSD <1.98% and <4.39%, respectively. Finally, the proposed method was successfully used for the residue

  11. Water evaporation rates across hydrophobic acid monolayers at equilibrium spreading pressure.

    PubMed

    Tsuji, Minami; Nakahara, Hiromichi; Moroi, Yoshikiyo; Shibata, Osamu

    2008-02-15

    The effect of alkanoic acid [CH(3)(CH(2))(n-2)COOH; HCn] and perfluoroalkanoic acid [CF(3)(CF(2))(n-2)COOH; FCn] monolayers on the water evaporation rate was investigated by thermogravimetry tracing the decrease in amount of water with time. The evaporation rate from the surface covered by a monolayer was measured as a function of temperature and hydrophobic chain length of the acids, where the monolayer was under an equilibrium spreading pressure. From thermal behavior of the crystallized acids, their solid states are C-type in crystalline state over the temperature range from 298.2 to 323.2 K. The dry air was flowed through a furnace tube of a thermogravimetry apparatus at the flow rate of 80 mL min(-1), where the evaporation rate becomes almost constant irrespective of the flow rate. The temperature dependence of the evaporation rate was analyzed kinetically to evaluate the activation energy and thermodynamics values for the activated complex, which demonstrated that these values were almost the same for both alkanoic acids and perfluoroalkanoic acids, although the effect of perfluoroalkanoic acids on the evaporation rate was smaller than that of corresponding hydrogenated fatty acids. The difference in the evaporation rate between FCn and HCn was examined by atomic force microscopy (AFM), Brewster angle microscopy (BAM), surface potential (DeltaV) at equilibrium spreading pressure, and Langmuir curve (pi-A isotherm), and their results were consistent and supported the difference. PMID:18048050

  12. Relationship between the electrochemical activity of Raney nickel and the rate of hydrogenation of maleic acid

    SciTech Connect

    Pervii, E.N.; Sofronkov, A.N.; Fedyshina, N.M.

    1986-02-10

    The purpose of this investigation was to determine the conditions in which a direct correlation exists between the rate of hydrogenation of maleic acid and the electrochemical activity of catalysts of hydrogen ionization. The rate of maleic acid hydrogenation in presence of Raney nickel catalyst was studied by a combination of volumetric and potentiometric methods.

  13. Influence of chlorine substitution on intramolecular hydrogen bond energy and ESIPT barrier: Experimental and theoretical measurements on the photophysics of 3,5-dichlorosalicylic acid

    NASA Astrophysics Data System (ADS)

    Paul, Bijan Kumar; Samanta, Anuva; Guchhait, Nikhil

    2010-08-01

    The effect of chlorine atom on the intramolecular hydrogen bond strength and excited state proton transfer barrier in pharmaceutically important chloro-substituted derivative of salicylic acid viz., 3,5-dichlorosalicylic acid (3,5DCSA) has been explored through steady-state absorption, emission and time-resolved fluorescence spectroscopy. Stokes shifted emission band with negligible solvent polarity dependency corresponds to the spectroscopic signature of excited state intramolecular proton transfer (ESIPT) reaction. The spectral signature was compared with its parent molecule salicylic acid (SA) and 5-chlorosalicylic acid (5ClSA). Quantum chemical calculations by ab initio Hartree-Fock (HF) and Density Functional Theory (DFT) methods have been fruitfully employed to correlate experimental findings. Calculated S0 and S1 states potential energy surfaces across the proton transfer co-ordinate substantiates the experimental evidence for the occurrence of ESIPT process and negates the ground state intramolecular proton transfer (GSIPT) reaction. Weakening of intramolecular hydrogen bond (IMHB) energy and subsequent enhancement of barrier to ESIPT reaction in 3,5DCSA as compared to SA and 5ClSA appears to be a reflection of conjugate impact of electron withdrawing inductive and electron donating resonance effects of chlorine substitutions depending on its location on the aromatic benzene nucleus.

  14. Substitution of DNA-Contacting Amino Acids with Functional Variants in the Gata-1 Zinc Finger: A Structurally and Phylogenetically Guided Mutagenesis

    PubMed Central

    Vonderfecht, Tyson R.; Schroyer, Daniel L.; Schenck, Brandy L.; McDonough, Virginia M.; Pikaart, Michael J.

    2008-01-01

    DNA binding functionality among transcription factor proteins is afforded by a number of structural motifs, such as the helix-turn-helix, helix-loop-helix, and zinc finger domains. The common thread among these diverse structures is their sequence-specific binding to essential promoter or other genetic regulatory sequences with high selectivity and affinity. One such motif, present in a wide range of organisms from bacteria to vertebrates, is the Gata-type zinc finger. This family of DNA-binding proteins is characterized by the presence of one or two (Cys)4 metal binding sites which recognize the protein’s eponymous binding site, GATA. Unlike other conserved DNA binding domains, Gata proteins appear to be restricted to binding consensus GATA sequences, or near variations, in DNA. Since the architecture of the Gata finger seems built around recognizing this particular sequence, we set out to define the allowable range of amino acid substitutions along the DNA-binding surface of a Gata finger that could continue to support sequence specific DNA binding activity. Accordingly, we set up a one-hybrid screen in yeast based on the chicken Gata-1 C-terminal zinc finger. Mutant libraries were generated at five amino acids identified in the Gata-DNA structure as likely to mediate sequence-specific contacts between the Gata finger and DNA. These libraries were designed to give as exhaustive amino acid coverage as possible such that almost all alternative amino acids were screened at each of the five probed positions. Screening and characterization of these libraries revealed several functional amino acid substitutions at two leucines which contact the DNA at the 3’ and 5’ flanks of the GATA binding site, but no functional substituents for amino acids near the core of the binding site. This pattern is consistent with amino acid sequences of known DNA-binding Gata fingers. PMID:18328814

  15. Measurement of the rates of oxindole-3-acetic acid turnover, and indole-3-acetic acid oxidation in Zea mays seedlings

    NASA Technical Reports Server (NTRS)

    Nonhebel, H. M.; Bandurski, R. S. (Principal Investigator)

    1986-01-01

    Oxindole-3-acetic acid is the principal catabolite of indole-3-acetic acid in Zea mays seedlings. In this paper measurements of the turnover of oxindole-3-acetic acid are presented and used to calculate the rate of indole-3-acetic acid oxidation. [3H]Oxindole-3-acetic acid was applied to the endosperm of Zea mays seedlings and allowed to equilibrate for 24 h before the start of the experiment. The subsequent decrease in its specific activity was used to calculate the turnover rate. The average half-life of oxindole-3-acetic acid in the shoots was found to be 30 h while that in the kernels had an average half-life of 35h. Using previously published values of the pool sizes of oxindole-3-acetic acid in shoots and kernels from seedlings of the same age and variety, and grown under the same conditions, the rate of indole-3-acetic acid oxidation was calculated to be 1.1 pmol plant-1 h-1 in the shoots and 7.1 pmol plant-1 h-1 in the kernels.

  16. The effect of linoleic acid on the whole body synthesis rates of polyunsaturated fatty acids from α-linolenic acid and linoleic acid in free-living rats.

    PubMed

    Domenichiello, Anthony F; Kitson, Alex P; Chen, Chuck T; Trépanier, Marc-Olivier; Stavro, P Mark; Bazinet, Richard P

    2016-04-01

    Docosahexaenoic acid (DHA) is thought to be important for brain function. The main dietary source of DHA is fish, however, DHA can also be synthesized from precursor omega-3 polyunsaturated fatty acids (n-3 PUFA), the most abundantly consumed being α-linolenic acid (ALA). The enzymes required to synthesize DHA from ALA are also used to synthesize longer chain omega-6 (n-6) PUFA from linoleic acid (LNA). The large increase in LNA consumption that has occurred over the last century has led to concern that LNA and other n-6 PUFA outcompete n-3 PUFA for enzymes involved in DHA synthesis, and therefore, decrease overall DHA synthesis. To assess this, rats were fed diets containing LNA at 53 (high LNA diet), 11 (medium LNA diet) or 1.5% (low LNA diet) of the fatty acids with ALA being constant across all diets (approximately 4% of the fatty acids). Rats were maintained on these diets from weaning for 8 weeks, at which point they were subjected to a steady-state infusion of labeled ALA and LNA to measure DHA and arachidonic acid (ARA) synthesis rates. DHA and ARA synthesis rates were generally highest in rats fed the medium and high LNA diets, while the plasma half-life of DHA was longer in rats fed the low LNA diet. Therefore, increasing dietary LNA, in rats, did not impair DHA synthesis; however, low dietary LNA led to a decrease in DHA synthesis with tissue concentrations of DHA possibly being maintained by a longer DHA half-life.

  17. Dietary Medium Chain Fatty Acid Supplementation Leads to Reduced VLDL Lipolysis and Uptake Rates in Comparison to Linoleic Acid Supplementation

    PubMed Central

    van Schalkwijk, Daniël B.; Pasman, Wilrike J.; Hendriks, Henk F. J.; Verheij, Elwin R.; Rubingh, Carina M.; van Bochove, Kees; Vaes, Wouter H. J.; Adiels, Martin; Freidig, Andreas P.; de Graaf, Albert A.

    2014-01-01

    Dietary medium chain fatty acids (MCFA) and linoleic acid follow different metabolic routes, and linoleic acid activates PPAR receptors. Both these mechanisms may modify lipoprotein and fatty acid metabolism after dietary intervention. Our objective was to investigate how dietary MCFA and linoleic acid supplementation and body fat distribution affect the fasting lipoprotein subclass profile, lipoprotein kinetics, and postprandial fatty acid kinetics. In a randomized double blind cross-over trial, 12 male subjects (age 51±7 years; BMI 28.5±0.8 kg/m2), were divided into 2 groups according to waist-hip ratio. They were supplemented with 60 grams/day MCFA (mainly C8:0, C10:0) or linoleic acid for three weeks, with a wash-out period of six weeks in between. Lipoprotein subclasses were measured using HPLC. Lipoprotein and fatty acid metabolism were studied using a combination of several stable isotope tracers. Lipoprotein and tracer data were analyzed using computational modeling. Lipoprotein subclass concentrations in the VLDL and LDL range were significantly higher after MCFA than after linoleic acid intervention. In addition, LDL subclass concentrations were higher in lower body obese individuals. Differences in VLDL metabolism were found to occur in lipoprotein lipolysis and uptake, not production; MCFAs were elongated intensively, in contrast to linoleic acid. Dietary MCFA supplementation led to a less favorable lipoprotein profile than linoleic acid supplementation. These differences were not due to elevated VLDL production, but rather to lower lipolysis and uptake rates. PMID:25049048

  18. Pleistocene evolutionary history of the Clouded Apollo (Parnassius mnemosyne): genetic signatures of climate cycles and a 'time-dependent' mitochondrial substitution rate.

    PubMed

    Gratton, P; Konopiński, M K; Sbordoni, V

    2008-10-01

    Genetic data are currently providing a large amount of new information on past distribution of species and are contributing to a new vision of Pleistocene ice ages. Nonetheless, an increasing number of studies on the 'time dependency' of mutation rates suggest that date assessments for evolutionary events of the Pleistocene might be overestimated. We analysed mitochondrial (mt) DNA (COI) sequence variation in 225 Parnassius mnemosyne individuals sampled across central and eastern Europe in order to assess (i) the existence of genetic signatures of Pleistocene climate shifts; and (ii) the timescale of demographic and evolutionary events. Our analyses reveal a phylogeographical pattern markedly influenced by the Pleistocene/Holocene climate shifts. Eastern Alpine and Balkan populations display comparatively high mtDNA diversity, suggesting multiple glacial refugia. On the other hand, three widely distributed and spatially segregated lineages occupy most of northern and eastern Europe, indicating postglacial recolonization from different refugial areas. We show that a conventional 'phylogenetic' substitution rate cannot account for the present distribution of genetic variation in this species, and we combine phylogeographical pattern and palaeoecological information in order to determine a suitable intraspecific rate through a Bayesian coalescent approach. We argue that our calibrated 'time-dependent' rate (0.096 substitutions/ million years), offers the most convincing time frame for the evolutionary events inferred from sequence data. When scaled by the new rate, estimates of divergence between Balkan and Alpine lineages point to c. 19 000 years before present (last glacial maximum), and parameters of demographic expansion for northern lineages are consistent with postglacial warming (5-11 000 years before present).

  19. The intraspecific variability of mitochondrial genes of Agaricus bisporus reveals an extensive group I intron mobility combined with low nucleotide substitution rates.

    PubMed

    Jalalzadeh, Banafsheh; Saré, Idy Carras; Férandon, Cyril; Callac, Philippe; Farsi, Mohammad; Savoie, Jean-Michel; Barroso, Gérard

    2015-02-01

    Intraspecific mitochondrial variability was studied in ten strains of A. bisporus var. bisporus, in a strain representative of A. bisporus var. eurotetrasporus and in a strain of the closely related species Agaricus devoniensis. In A. bisporus, the cox1 gene is the richest in group I introns harboring homing endonuclease genes (heg). This study led to identify group I introns as the main source of cox1 gene polymorphism. Among the studied introns, two groups were distinguished according to the heg they contained. One group harbored heg maintained putatively functional. The other group was composed of eroded heg sequences that appeared to evolve toward their elimination. Low nucleotide substitution rates were found in both types of intronic sequences. This feature was also shared by all types of studied mitochondrial sequences, not only intronic but also genic and intergenic ones, when compared with nuclear sequences. Hence, the intraspecific evolution of A. bisporus mitochondrial genome appears characterized by both an important mobility (presence/absence) of large group I introns and by low nt substitution rates. This stringent conservation of mitochondrial sequences, when compared with their nuclear counterparts, appears irrespective of their apparent functionality and contrasts to what is widely accepted in fungal sequence evolution. This strengthens the usefulness of mtDNA sequences to get clues on intraspecific evolution.

  20. Palladium-catalyzed C–N and C–O bond formation of N-substituted 4-bromo-7-azaindoles with amides, amines, amino acid esters and phenols

    PubMed Central

    Surasani, Rajendra; Rao, A V Dhanunjaya; Chandrasekhar, K B

    2012-01-01

    Summary Simple and efficient procedures for palladium-catalyzed cross-coupling reactions of N-substituted 4-bromo-7-azaindole (1H-pyrrole[2,3-b]pyridine), with amides, amines, amino acid esters and phenols through C–N and C–O bond formation have been developed. The C–N cross-coupling reaction of amides, amines and amino acid esters takes place rapidly by using the combination of Xantphos, Cs2CO3, dioxane and palladium catalyst precursors Pd(OAc)2/Pd2(dba)3. The combination of Pd(OAc)2, Xantphos, K2CO3 and dioxane was found to be crucial for the C–O cross-coupling reaction. This is the first report on coupling of amides, amino acid esters and phenols with N-protected 4-bromo-7-azaindole derivatives. PMID:23209536

  1. A single amino-acid substitution toggles chloride dependence of the alpha-amylase paralog amyrel in Drosophila melanogaster and Drosophila virilis species.

    PubMed

    Claisse, Gaëlle; Feller, Georges; Bonneau, Magalie; Da Lage, Jean-Luc

    2016-08-01

    In animals, most α-amylases are chloride-dependent enzymes. A chloride ion is required for allosteric activation and is coordinated by one asparagine and two arginine side chains. Whereas the asparagine and one arginine are strictly conserved, the main chloride binding arginine is replaced by a glutamine in some rare instances, resulting in the loss of chloride binding and activation. Amyrel is a distant paralogue of α-amylase in Diptera, which was not characterized biochemically to date. Amyrel shows both substitutions depending on the species. In Drosophila melanogaster, an arginine is present in the sequence but in Drosophila virilis, a glutamine occurs at this position. We have investigated basic enzymological parameters and the dependence to chloride of Amyrel of both species, produced in yeast, and in mutants substituting arginine to glutamine or glutamine to arginine. We found that the amylolytic activity of Amyrel is about thirty times weaker than the classical Drosophila α-amylase, and that the substitution of the arginine by a glutamine in D. melanogaster suppressed the chloride-dependence but was detrimental to activity. In contrast, changing the glutamine into an arginine rendered D. virilis Amyrel chloride-dependent, and interestingly, significantly increased its catalytic efficiency. These results show that the chloride ion is not mandatory for Amyrel but stimulates the reaction rate. The possible phylogenetic origin of the arginine/glutamine substitution is also discussed. PMID:27312592

  2. Bovine rhodopsin: amino acid substitutions Asp-83----Asn and Glu-134----Gln prevent activation of cyclic GMP phosphodiesterase.

    PubMed

    Gurevich, V V; Zozulya, S A; Zerf, E P; Pokrovskaya, I D; Obukhova, T A; Garnovskaya, M N; Dumler, I L; Rychkova, M P

    1990-01-01

    Two bovine rhodopsin mutants with substitutions of negatively charged residues within transmembrane domains II and III by uncharged ones (Asp-83----Asn and Glu-134----Gln) were constructed. Both mutants stimulated transducin GTPase with slightly lowered efficiency, but were completely unable to activate cyclic GMP phosphodiesterase. PMID:1966786

  3. Effect of specific amino acid substitutions in the putative fusion peptide of structural glycoprotein E2 on Classical Swine Fever Virus replication

    SciTech Connect

    Fernández-Sainz, I.J.; Largo, E.; Gladue, D.P.; Fletcher, P.; O’Donnell, V.; Holinka, L.G.; Carey, L.B.; Lu, X.; Nieva, J.L.; Borca, M.V.

    2014-05-15

    E2, along with E{sup rns} and E1, is an envelope glycoprotein of Classical Swine Fever Virus (CSFV). E2 is involved in several virus functions: cell attachment, host range susceptibility and virulence in natural hosts. Here we evaluate the role of a specific E2 region, {sup 818}CPIGWTGVIEC{sup 828}, containing a putative fusion peptide (FP) sequence. Reverse genetics utilizing a full-length infectious clone of the highly virulent CSFV strain Brescia (BICv) was used to evaluate how individual amino acid substitutions within this region of E2 may affect replication of BICv. A synthetic peptide representing the complete E2 FP amino acid sequence adopted a β-type extended conformation in membrane mimetics, penetrated into model membranes, and perturbed lipid bilayer integrity in vitro. Similar peptides harboring amino acid substitutions adopted comparable conformations but exhibited different membrane activities. Therefore, a preliminary characterization of the putative FP {sup 818}CPIGWTGVIEC{sup 828} indicates a membrane fusion activity and a critical role in virus replication. - Highlights: • A putative fusion peptide (FP) region in CSFV E2 protein was shown to be critical for virus growth. • Synthetic FPs were shown to efficiently penetrate into lipid membranes using an in vitro model. • Individual residues in the FP affecting virus replication were identified by reverse genetics. • The same FP residues are also responsible for mediating membrane fusion.

  4. Epistasis effects of multiple ancestral-consensus amino acid substitutions on the thermal stability of glycerol kinase from Cellulomonas sp. NT3060.

    PubMed

    Fukuda, Yasuhisa; Abe, Asuka; Tamura, Takashi; Kishimoto, Takahide; Sogabe, Atsushi; Akanuma, Satoshi; Yokobori, Shin-Ichi; Yamagishi, Akihiko; Imada, Katsumi; Inagaki, Kenji

    2016-05-01

    Thermostable variants of the Cellulomonas sp. NT3060 glycerol kinase have been constructed by through the introduction of ancestral-consensus mutations. We produced seven mutants, each having an ancestral-consensus amino acid residue that might be present in the common ancestors of both bacteria and of archaea, and that appeared most frequently at the position of 17 glycerol kinase sequences in the multiple sequence alignment. The thermal stabilities of the resulting mutants were assessed by determining their melting temperatures (Tm), which was defined as the temperature at which 50% of the initial catalytic activity is lost after 15 min of incubation, as well as when the half-life of the catalytic activity occurs at a temperature of 60°C (t1/2). Three mutants showed increased stabilities compared to the wild-type protein. We then produced five more mutants with multiple amino acid substitutions. Some of the resulting mutants showed thermal stabilities much greater than those expected given the stabilities of the respective mutants with single mutations. Therefore, the effects of mutations are not always simply additive and some amino acid substitutions, which do not affect or only slightly improve stability when individually introduced into the protein, show substantial stabilizing effects in combination with other mutations.

  5. Evolution of a new function in an esterase: simple amino acid substitutions enable the activity present in the larger paralog, BioH.

    PubMed

    Flores, Humberto; Lin, Steven; Contreras-Ferrat, Gabriel; Cronan, John E; Morett, Enrique

    2012-08-01

    Gene duplication and divergence are essential processes for the evolution of new activities. Divergence may be gradual, involving simple amino acid residue substitutions, or drastic, such that larger structural elements are inserted, deleted or rearranged. Vast protein sequence comparisons, supported by some experimental evidence, argue that large structural modifications have been necessary for certain catalytic activities to evolve. However, it is not clear whether these activities could not have been attained by gradual changes. Interestingly, catalytic promiscuity could play a fundamental evolutionary role: a preexistent secondary activity could be increased by simple amino acid residue substitutions that do not affect the enzyme's primary activity. The promiscuous profile of the enzyme may be modified gradually by genetic drift, making a pool of potentially useful activities that can be selected before duplication. In this work, we used random mutagenesis and in vivo selection to evolve the Pseudomonas aeruginosa PAO1 carboxylesterase PA3859, a small protein, to attain the function of BioH, a much larger paralog involved in biotin biosynthesis. BioH was chosen as a target activity because it provides a highly sensitive selection for evolved enzymatic activities by auxotrophy complementation. After only two cycles of directed evolution, mutants with the ability to efficiently complement biotin auxotrophy were selected. The in vivo and in vitro characterization showed that the activity of one of our mutant proteins was similar to that of the wild-type BioH enzyme. Our results demonstrate that it is possible to evolve enzymatic activities present in larger proteins by discrete amino acid substitutions.

  6. Unique amino acid substitutions in the capsid proteins of foot-and-mouth disease virus from a persistent infection in cell culture.

    PubMed Central

    Díez, J; Dávila, M; Escarmís, C; Mateu, M G; Dominguez, J; Pérez, J J; Giralt, E; Melero, J A; Domingo, E

    1990-01-01

    Maintenance of a persistent foot-and-mouth disease virus (FMDV) infection in BHK-21 cells involves a coevolution of cells and virus (J. C. de la Torre, E. Martínez-Salas, J. Díez, A. Villaverde, F. Gebauer, E. Rocha, M. Dávila, and E. Domingo, J. Virol. 62:2050-2058, 1988). The resident FMDV undergoes a number of phenotypic changes, including a gradual decrease in virion stability. Here we report the nucleotide sequence of the P1 genomic segment of the virus rescued after 100 passages of the carrier cells (R100). Only 5 of 15 mutations in P1 of R100 were silent. Nine amino acid substitutions were fixed on the viral capsid during persistence, and three of the variant amino acids are not represented in the corresponding position of any picornavirus sequenced to date. Cysteine at position 7 of VP3, that provides disulfide bridges at the FMDV fivefold axis, was substituted by valine, as determined by RNA, cDNA, and protein sequencing. The modified virus shows high buoyant density in cesium chloride and depicts the same sensitivity to photoinactivation by intercalating dyes as the parental FMDV C-S8c1. Amino acid substitutions fixed in VP1 resulted in altered antigenicity, as revealed by reactivity with monoclonal antibodies. In addition to defining at the molecular level the alterations the FMDV capsid underwent during persistence, the results show that positions which are highly invariant in an RNA genome may change when viral replication occurs in a modified environment. Images PMID:2170684

  7. ENTPRISE: An Algorithm for Predicting Human Disease-Associated Amino Acid Substitutions from Sequence Entropy and Predicted Protein Structures

    PubMed Central

    Zhou, Hongyi; Gao, Mu; Skolnick, Jeffrey

    2016-01-01

    The advance of next-generation sequencing technologies has made exome sequencing rapid and relatively inexpensive. A major application of exome sequencing is the identification of genetic variations likely to cause Mendelian diseases. This requires processing large amounts of sequence information and therefore computational approaches that can accurately and efficiently identify the subset of disease-associated variations are needed. The accuracy and high false positive rates of existing computational tools leave much room for improvement. Here, we develop a boosted tree regression machine-learning approach to predict human disease-associated amino acid variations by utilizing a comprehensive combination of protein sequence and structure features. On comparing our method, ENTPRISE, to the state-of-the-art methods SIFT, PolyPhen-2, MUTATIONASSESSOR, MUTATIONTASTER, FATHMM, ENTPRISE exhibits significant improvement. In particular, on a testing dataset consisting of only proteins with balanced disease-associated and neutral variations defined as having the ratio of neutral/disease-associated variations between 0.3 and 3, the Mathews Correlation Coefficient by ENTPRISE is 0.493 as compared to 0.432 by PPH2-HumVar, 0.406 by SIFT, 0.403 by MUTATIONASSESSOR, 0.402 by PPH2-HumDiv, 0.305 by MUTATIONTASTER, and 0.181 by FATHMM. ENTPRISE is then applied to nucleic acid binding proteins in the human proteome. Disease-associated predictions are shown to be highly correlated with the number of protein-protein interactions. Both these predictions and the ENTPRISE server are freely available for academic users as a web service at http://cssb.biology.gatech.edu/entprise/. PMID:26982818

  8. ENTPRISE: An Algorithm for Predicting Human Disease-Associated Amino Acid Substitutions from Sequence Entropy and Predicted Protein Structures.

    PubMed

    Zhou, Hongyi; Gao, Mu; Skolnick, Jeffrey

    2016-01-01

    The advance of next-generation sequencing technologies has made exome sequencing rapid and relatively inexpensive. A major application of exome sequencing is the identification of genetic variations likely to cause Mendelian diseases. This requires processing large amounts of sequence information and therefore computational approaches that can accurately and efficiently identify the subset of disease-associated variations are needed. The accuracy and high false positive rates of existing computational tools leave much room for improvement. Here, we develop a boosted tree regression machine-learning approach to predict human disease-associated amino acid variations by utilizing a comprehensive combination of protein sequence and structure features. On comparing our method, ENTPRISE, to the state-of-the-art methods SIFT, PolyPhen-2, MUTATIONASSESSOR, MUTATIONTASTER, FATHMM, ENTPRISE exhibits significant improvement. In particular, on a testing dataset consisting of only proteins with balanced disease-associated and neutral variations defined as having the ratio of neutral/disease-associated variations between 0.3 and 3, the Mathews Correlation Coefficient by ENTPRISE is 0.493 as compared to 0.432 by PPH2-HumVar, 0.406 by SIFT, 0.403 by MUTATIONASSESSOR, 0.402 by PPH2-HumDiv, 0.305 by MUTATIONTASTER, and 0.181 by FATHMM. ENTPRISE is then applied to nucleic acid binding proteins in the human proteome. Disease-associated predictions are shown to be highly correlated with the number of protein-protein interactions. Both these predictions and the ENTPRISE server are freely available for academic users as a web service at http://cssb.biology.gatech.edu/entprise/.

  9. Design, synthesis and biological evaluation of di-substituted cinnamic hydroxamic acids bearing urea/thiourea unit as potent histone deacetylase inhibitors.

    PubMed

    Ning, Chengqing; Bi, Yanjing; He, Yujun; Huang, WenYuan; Liu, Lifei; Li, Yi; Zhang, Sihan; Liu, Xiaoyu; Yu, Niefang

    2013-12-01

    A novel class of di-substituted cinnamic hydroxamic acid derivatives containing urea or thiourea unit was designed, synthesized and evaluated as HDAC inhibitors. All tested compounds demonstrated significant HDAC inhibitory activities and anti-proliferative effects against diverse human tumor cell lines. Among them, 7l exhibited most potent pan-HDAC inhibitory activity, with an IC50 value of 130 nM. It also showed strong cellular inhibition against diverse cell lines including HCT-116, MCF-7, MDB-MB-435 and NCI-460, with GI50 values of 0.35, 0.22, 0.51 and 0.48 μM, respectively.

  10. Structural and Functional Characteristics of Oxysterol 7α-Hydroxylase with Amino-Acid Substitution R486C and Their Relation to the Appearance of Neurodegenerative Diseases

    NASA Astrophysics Data System (ADS)

    Dichenko, Ya. V.; Yantsevich, A. V.; Usanov, S. A.

    2015-03-01

    The influence of the amino-acid substitution Arg486Cys on the conformational stability of recombinant cytochrome P450 7B1 (CYP7B1, oxysterol 7α-hydroxylase) was studied. The single base change was shown to decrease the free energy of the transition of the heme-protein from its native state to a denatured one, which pointed to a lower thermodynamic stability for the mutant form of the enzyme. This could be the cause of the metabolic disruption of neurosteroids and, as a consequence, the appearance of neurodegenerative diseases.

  11. Amino acid rating method for evaluating protein adequacy of infant formulas.

    PubMed

    Sarwar, G; Botting, H G; Peace, R W

    1989-01-01

    Amino acid profiles and/or protein digestibility (by the rat balance method) were determined for various forms (powder, ready-to-use, liquid concentrate, etc.) of cow's milk- and soy-based infant formulas obtained from 4 manufacturers. The essential amino acid data of the formulas were compared with that of human milk for the calculation of amino acid scores (based on the single most limiting amino acid). The product of amino acid score and total protein (g/100 kcal) was then termed "amino acid rating." Amino acid scores for the milk- and soy-based formulas ranged from 59 to 90 and from 59 to 81%, respectively, due to deficiencies in sulfur amino acids and/or tryptophan. Because of significantly higher total protein contents (g/100 kcal) of soy- (2.65-3.68) and milk-based (2.20-2.95) infant formulas compared to human milk (1.5), the relative amino acid ratings (human milk = 100) for all infant formulas except 2 liquid concentrates (having values of 87%) were above 100%. Values for true digestibility of protein in milk- and soy-based formulas ranged from 87 to 97 and from 92 to 95%, respectively. When corrected for protein digestibility, the relative amino acid ratings for all the milk-based liquid concentrates were below 100% (77-98%).

  12. Reliability of Urinary Excretion Rate Adjustment in Measurements of Hippuric Acid in Urine

    PubMed Central

    Nicolli, Annamaria; Chiara, Federica; Gambalunga, Alberto; Carrieri, Mariella; Bartolucci, Giovanni Battista; Trevisan, Andrea

    2014-01-01

    The urinary excretion rate is calculated based on short-term, defined time sample collections with a known sample mass, and this measurement can be used to remove the variability in urine concentrations due to urine dilution. Adjustment to the urinary excretion rate of hippuric acid was evaluated in 31 healthy volunteers (14 males and 17 females). Urine was collected as short-term or spot samples and tested for specific gravity, creatinine and hippuric acid. Hippuric acid values were unadjusted or adjusted to measurements of specific gravity, creatinine or urinary excretion rate. Hippuric acid levels were partially independent of urinary volume and urinary flow rate, in contrast to specific gravity and creatinine, which were both highly dependent on the hippuric acid level. Accordingly, hippuric acid was independent on urinary specific gravity and creatinine excretion. Unadjusted and adjusted values for specific gravity or creatinine were generally closely correlated, especially in spot samples. Values adjusted to the urinary excretion rate appeared well correlated to those unadjusted and adjusted to specific gravity or creatinine values. Thus, adjustment of crude hippuric acid values to the urinary excretion rate is a valid procedure but is difficult to apply in the field of occupational medicine and does not improve the information derived from values determined in spot urine samples, either unadjusted or adjusted to specific gravity and creatinine. PMID:25019265

  13. The Prevalence of Hepatitis C Virus Core Amino Acid 70 Substitution and Genotypes of Polymorphisms Near the IFNL3 Gene in Iranian Patients With Chronic Hepatitis C

    PubMed Central

    Kadjbaf, Danesh; Keshvari, Maryam; Alavian, Seyed Moayed; Pouryasin, Ali; Behnava, Bita; Salimi, Shima; Mehrnoush, Leila; Karimi Elizee, Pegah; Sharafi, Heidar

    2016-01-01

    Background Molecular studies have demonstrated that the hepatitis C virus (HCV) genotype and host genetics play predictive roles in the management of patients infected with HCV. Objectives This study aimed to investigate the HCV genotype, core amino acid (aa) 70 substitution, and polymorphisms near the IFNL3 gene (including rs12979860 and rs8099917) among Iranian patients with chronic hepatitis C (CHC). Patients and Methods In this cross-sectional study, the molecular profiles of the HCV genotype, core aa 70 substitution, and rs12979860 and rs8099917 polymorphisms and plasma HCV RNA levels were determined in 429 CHC patients including 141 hemophilic, 84 thalassemic, and 204 non-hemophilic, non-thalassemic patients. Results The hepatitis C virus subtype 1a was the most common subtype in the study population. Core aa substitution Arg70Gln was strongly associated with cirrhosis (OR = 2.49; 95% CI = 1.13 - 5.50; P = 0.020). Core aa 70 substitutions were more frequently observed in patients with the HCV subtype 1b than in patients with any other HCV subtypes (P < 0.001). Core aa 70 substitutions were also more common in patients with the rs12979860 TT genotype than in patients with non-TT genotypes (17.3% vs. 8.5%, P = 0.022) and also in rs8099917 non-TT genotypes than in the TT genotype (14.0% vs. 7.0%, P = 0.026). The HCV genotypes and rs8099917 polymorphisms were correlated in which HCV subtype 1b was in favor of rs8099917 GG and HCV subtype 3a favored rs8099917 TT (P = 0.021). Furthermore, the rs12979860 TT and rs8099917 GG genotypes showed significantly lower HCV RNA levels than the other genotypes (P < 0.001). Conclusions There is an as yet unexplained association between HCV and host parameters with unknown mechanisms in patients with chronic HCV infection. The assessments of core aa 70 substitution and polymorphisms near the IFNL3 gene could offer promising steps to improve the management of patients with HCV.

  14. The Prevalence of Hepatitis C Virus Core Amino Acid 70 Substitution and Genotypes of Polymorphisms Near the IFNL3 Gene in Iranian Patients With Chronic Hepatitis C

    PubMed Central

    Kadjbaf, Danesh; Keshvari, Maryam; Alavian, Seyed Moayed; Pouryasin, Ali; Behnava, Bita; Salimi, Shima; Mehrnoush, Leila; Karimi Elizee, Pegah; Sharafi, Heidar

    2016-01-01

    Background Molecular studies have demonstrated that the hepatitis C virus (HCV) genotype and host genetics play predictive roles in the management of patients infected with HCV. Objectives This study aimed to investigate the HCV genotype, core amino acid (aa) 70 substitution, and polymorphisms near the IFNL3 gene (including rs12979860 and rs8099917) among Iranian patients with chronic hepatitis C (CHC). Patients and Methods In this cross-sectional study, the molecular profiles of the HCV genotype, core aa 70 substitution, and rs12979860 and rs8099917 polymorphisms and plasma HCV RNA levels were determined in 429 CHC patients including 141 hemophilic, 84 thalassemic, and 204 non-hemophilic, non-thalassemic patients. Results The hepatitis C virus subtype 1a was the most common subtype in the study population. Core aa substitution Arg70Gln was strongly associated with cirrhosis (OR = 2.49; 95% CI = 1.13 - 5.50; P = 0.020). Core aa 70 substitutions were more frequently observed in patients with the HCV subtype 1b than in patients with any other HCV subtypes (P < 0.001). Core aa 70 substitutions were also more common in patients with the rs12979860 TT genotype than in patients with non-TT genotypes (17.3% vs. 8.5%, P = 0.022) and also in rs8099917 non-TT genotypes than in the TT genotype (14.0% vs. 7.0%, P = 0.026). The HCV genotypes and rs8099917 polymorphisms were correlated in which HCV subtype 1b was in favor of rs8099917 GG and HCV subtype 3a favored rs8099917 TT (P = 0.021). Furthermore, the rs12979860 TT and rs8099917 GG genotypes showed significantly lower HCV RNA levels than the other genotypes (P < 0.001). Conclusions There is an as yet unexplained association between HCV and host parameters with unknown mechanisms in patients with chronic HCV infection. The assessments of core aa 70 substitution and polymorphisms near the IFNL3 gene could offer promising steps to improve the management of patients with HCV. PMID:27630727

  15. 40 CFR 721.10202 - Benzoic acid, 4-chloro-2- [(substituted)azo]-, strontium salt (1:1) (generic).

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false Benzoic acid, 4-chloro-2- -, strontium... New Uses for Specific Chemical Substances § 721.10202 Benzoic acid, 4-chloro-2- -, strontium salt (1:1... identified generically as benzoic acid, 4-chloro-2- -, strontium salt (1:1) (PMN P-08-701) is subject...

  16. Calibration of a molecular clock in tits (Paridae)--do nucleotide substitution rates of mitochondrial genes deviate from the 2% rule?

    PubMed

    Päckert, Martin; Martens, Jochen; Tietze, Dieter Thomas; Dietzen, Christian; Wink, Michael; Kvist, Laura

    2007-07-01

    The ongoing debate on the reliability of avian molecular clocks is actually based on only a small number of calibrations carried out under different assumptions with respect to the choice and constraints of calibration points or to the use of substitution models. In this study, we provide substitution rate estimates for two mitochondrial genes, cytochrome b and the control region, and age estimates for lineage splits within four subgenera of tits (Paridae: Parus, Cyanistes, Poecile and Periparus). Overall sequence divergence between cytochrome b lineages covers a range of 0.4-1.8% per million years and is thus consistent with the frequently adopted approximation for a sequence divergence between avian lineages of 1.6-2% per my. Overall rate variation is high and encompasses the 2% value in a 95% CI for model corrected data. Mean rate estimates for cytochrome b range between 1.9 and 8.9 x 10(-3) substitutions per site per lineage. Local rates differ significantly between taxonomic levels with lowest estimates for haplotype lineages. At the population/subspecies level mean sequence divergence between lineages matches the 2% rule best for most cytochrome b datasets (1.5-1.9% per my) with maximum estimates for small isolated populations like those of the Canarian P. teneriffae complex (up to 3.9% per my). Overall rate estimates for the control region range at similar values like those for cytochrome b (2.7-8.8 x 10(-3), 0.5-1.8% per my), however, within some subgenera mean rates are higher than those for cytochrome b for uncorrected sequence data. The lowest rates for both genes were calculated for coal tits of subgenus Periparus (0.04-0.6% per my). Model-corrected sequence data tend to result in higher rate estimates than uncorrected data. Increase of the gamma shape parameter goes along with a significant decrease of rate and partly age estimates, too. Divergence times for earliest deep splits within tit subgenera Periparus and Parus were dated to the mid Miocene at

  17. Isotope-specific and amino acid-specific heavy atom substitutions alter barrier crossing in human purine nucleoside phosphorylase.

    PubMed

    Suarez, Javier; Schramm, Vern L

    2015-09-01

    Computational chemistry predicts that atomic motions on the femtosecond timescale are coupled to transition-state formation (barrier-crossing) in human purine nucleoside phosphorylase (PNP). The prediction is experimentally supported by slowed catalytic site chemistry in isotopically labeled PNP (13C, 15N, and 2H). However, other explanations are possible, including altered volume or bond polarization from carbon-deuterium bonds or propagation of the femtosecond bond motions into slower (nanoseconds to milliseconds) motions of the larger protein architecture to alter catalytic site chemistry. We address these possibilities by analysis of chemistry rates in isotope-specific labeled PNPs. Catalytic site chemistry was slowed for both [2H]PNP and [13C, 15N]PNP in proportion to their altered protein masses. Secondary effects emanating from carbon-deuterium bond properties can therefore be eliminated. Heavy-enzyme mass effects were probed for local or global contributions to catalytic site chemistry by generating [15N, 2H]His8-PNP. Of the eight His per subunit, three participate in contacts to the bound reactants and five are remote from the catalytic sites. [15N, 2H]His8-PNP had reduced catalytic site chemistry larger than proportional to the enzymatic mass difference. Altered barrier crossing when only His are heavy supports local catalytic site femtosecond perturbations coupled to transition-state formation. Isotope-specific and amino acid specific labels extend the use of heavy enzyme methods to distinguish global from local isotope effects.

  18. Isotope-specific and amino acid-specific heavy atom substitutions alter barrier crossing in human purine nucleoside phosphorylase.

    PubMed

    Suarez, Javier; Schramm, Vern L

    2015-09-01

    Computational chemistry predicts that atomic motions on the femtosecond timescale are coupled to transition-state formation (barrier-crossing) in human purine nucleoside phosphorylase (PNP). The prediction is experimentally supported by slowed catalytic site chemistry in isotopically labeled PNP (13C, 15N, and 2H). However, other explanations are possible, including altered volume or bond polarization from carbon-deuterium bonds or propagation of the femtosecond bond motions into slower (nanoseconds to milliseconds) motions of the larger protein architecture to alter catalytic site chemistry. We address these possibilities by analysis of chemistry rates in isotope-specific labeled PNPs. Catalytic site chemistry was slowed for both [2H]PNP and [13C, 15N]PNP in proportion to their altered protein masses. Secondary effects emanating from carbon-deuterium bond properties can therefore be eliminated. Heavy-enzyme mass effects were probed for local or global contributions to catalytic site chemistry by generating [15N, 2H]His8-PNP. Of the eight His per subunit, three participate in contacts to the bound reactants and five are remote from the catalytic sites. [15N, 2H]His8-PNP had reduced catalytic site chemistry larger than proportional to the enzymatic mass difference. Altered barrier crossing when only His are heavy supports local catalytic site femtosecond perturbations coupled to transition-state formation. Isotope-specific and amino acid specific labels extend the use of heavy enzyme methods to distinguish global from local isotope effects. PMID:26305965

  19. Isotope-specific and amino acid-specific heavy atom substitutions alter barrier crossing in human purine nucleoside phosphorylase

    PubMed Central

    Suarez, Javier; Schramm, Vern L.

    2015-01-01

    Computational chemistry predicts that atomic motions on the femtosecond timescale are coupled to transition-state formation (barrier-crossing) in human purine nucleoside phosphorylase (PNP). The prediction is experimentally supported by slowed catalytic site chemistry in isotopically labeled PNP (13C, 15N, and 2H). However, other explanations are possible, including altered volume or bond polarization from carbon-deuterium bonds or propagation of the femtosecond bond motions into slower (nanoseconds to milliseconds) motions of the larger protein architecture to alter catalytic site chemistry. We address these possibilities by analysis of chemistry rates in isotope-specific labeled PNPs. Catalytic site chemistry was slowed for both [2H]PNP and [13C, 15N]PNP in proportion to their altered protein masses. Secondary effects emanating from carbon–deuterium bond properties can therefore be eliminated. Heavy-enzyme mass effects were probed for local or global contributions to catalytic site chemistry by generating [15N, 2H]His8-PNP. Of the eight His per subunit, three participate in contacts to the bound reactants and five are remote from the catalytic sites. [15N, 2H]His8-PNP had reduced catalytic site chemistry larger than proportional to the enzymatic mass difference. Altered barrier crossing when only His are heavy supports local catalytic site femtosecond perturbations coupled to transition-state formation. Isotope-specific and amino acid specific labels extend the use of heavy enzyme methods to distinguish global from local isotope effects. PMID:26305965

  20. Liquid-Phase Heat-Release Rates of the Systems Hydrazine-Nitric Acid and Unsymmetrical Dimethylhydrazine-Nitric Acid

    NASA Technical Reports Server (NTRS)

    Somogyi, Dezso; Feiler, Charles E.

    1960-01-01

    The initial rates of heat release produced by the reactions of hydrazine and unsymmetrical dimethylhydrazine with nitric acid were determined in a bomb calorimeter under conditions of forced mixing. Fuel-oxidant weight ratio and injection velocity were varied. The rate of heat release apparently depended on the interfacial area between the propellants. Above a narrow range of injection velocities representing a critical amount of interfacial area, the rates reached a maximum and were almost constant with injection velocity. The maximum rate for hydrazine was about 70 percent greater than that for unsymmetrical dimethylhydrazine. The total heat released did not vary with mixture ratio over the range studied.

  1. Protodeboronation of Heteroaromatic, Vinyl, and Cyclopropyl Boronic Acids: pH-Rate Profiles, Autocatalysis, and Disproportionation.

    PubMed

    Cox, Paul A; Leach, Andrew G; Campbell, Andrew D; Lloyd-Jones, Guy C

    2016-07-27

    pH-rate profiles for aqueous-organic protodeboronation of 18 boronic acids, many widely viewed as unstable, have been studied by NMR and DFT. Rates were pH-dependent, and varied substantially between the boronic acids, with rate maxima that varied over 6 orders of magnitude. A mechanistic model containing five general pathways (k1-k5) has been developed, and together with input of [B]tot, KW, Ka, and KaH, the protodeboronation kinetics can be correlated as a function of pH (1-13) for all 18 species. Cyclopropyl and vinyl boronic acids undergo very slow protodeboronation, as do 3- and 4-pyridyl boronic acids (t0.5 > 1 week, pH 12, 70 °C). In contrast, 2-pyridyl and 5-thiazolyl boronic acids undergo rapid protodeboronation (t0.5 ≈ 25-50 s, pH 7, 70 °C), via fragmentation of zwitterionic intermediates. Lewis acid additives (e.g., Cu, Zn salts) can attenuate (2-pyridyl) or accelerate (5-thiazolyl and 5-pyrazolyl) fragmentation. Two additional processes compete when the boronic acid and the boronate are present in sufficient proportions (pH = pKa ± 1.6): (i) self-/autocatalysis and (ii) sequential disproportionations of boronic acid to borinic acid and borane. PMID:27355973

  2. Measurement of Fatty Acid Oxidation Rates in Animal Tissues and Cell Lines

    PubMed Central

    Huynh, Frank K.; Green, Michelle F.; Koves, Timothy R.; Hirschey, Matthew D.

    2014-01-01

    While much oncological research has focused on metabolic shifts in glucose and amino acid oxidation, recent evidence suggests that fatty acid oxidation (FAO) may also play an important role in the metabolic reprogramming of cancer cells. Here, we present a simple method for measuring FAO rates using radiolabeled palmitate, common laboratory reagents, and standard supplies. This protocol is broadly applicable for measuring FAO rates in cultured cancer cells as well as in both malignant and nontransformed animal tissues. PMID:24862277

  3. An Amino Acid Substitution Inhibits Specialist Herbivore Production of an Antagonist Effector and Recovers Insect-Induced Plant Defenses1[W][OA

    PubMed Central

    Schmelz, Eric A.; Huffaker, Alisa; Carroll, Mark J.; Alborn, Hans T.; Ali, Jared G.; Teal, Peter E.A.

    2012-01-01

    Plants respond to insect herbivory through the production of biochemicals that function as either direct defenses or indirect defenses via the attraction of natural enemies. While attack by closely related insect pests can result in distinctive levels of induced plant defenses, precise biochemical mechanisms responsible for differing responses remain largely unknown. Cowpea (Vigna unguiculata) responds to Fall armyworm (Spodoptera frugiperda) herbivory through the detection of fragments of chloroplastic ATP synthase γ-subunit proteins, termed inceptin-related peptides, present in larval oral secretions (OS). In contrast to generalists like Fall armyworm, OS of the legume-specializing velvetbean caterpillar (VBC; Anticarsia gemmatalis) do not elicit ethylene production and demonstrate significantly lower induced volatile emission in direct herbivory comparisons. Unlike all other Lepidoptera OS examined, which preferentially contain inceptin (Vu-In; +ICDINGVCVDA−), VBC OS contain predominantly a C-terminal truncated peptide, Vu-In−A (+ICDINGVCVD−). Vu-In−A is both inactive and functions as a potent naturally occurring antagonist of Vu-In-induced responses. To block antagonist production, amino acid substitutions at the C terminus were screened for differences in VBC gut proteolysis. A valine-substituted peptide (Vu-InΔV; +ICDINGVCVDV−) retaining full elicitor activity was found to accumulate in VBC OS. Compared with the native polypeptide, VBC that previously ingested 500 pmol of the valine-modified chloroplastic ATP synthase γ-subunit precursor elicited significantly stronger plant responses in herbivory assays. We demonstrate that a specialist herbivore minimizes the activation of defenses by converting an elicitor into an antagonist effector and identify an amino acid substitution that recovers these induced plant defenses to a level observed with generalist herbivores. PMID:23008466

  4. Identification and localization of amino acid substitutions between two phenobarbital-inducible rat hepatic microsomal cytochromes P-450 by micro sequence analyses.

    PubMed Central

    Yuan, P M; Ryan, D E; Levin, W; Shively, J E

    1983-01-01

    Two isozymes of rat liver microsomal cytochrome P-450--P-450b and P-450e--were compared by micro sequence analyses of their NH2 termini and tryptic fragments. These two phenobarbital-inducible hemoproteins, which are immunochemically indistinguishable with antibody against cytochrome P-450b, have extensive sequence homology. Automated Edman degradation of the native proteins revealed identical amino acids for the first 35 residues. Sequence determinations of the tryptic peptides, which constitute approximately 75% of each protein molecule, have thus far shown 10 amino acid differences between the two isozymes. Results of our amino acid sequence analyses established that two of the cDNAs, pcP-450pb1 and pcP-450pb4, reported by Fujii-Kuriyama et al. [Fujii-Kuriyama, Y., Mizukami, Y., Kamajiri, K., Sogawa, K. & Muramatsu, M. (1982) Proc. Natl. Acad. Sci. USA 79, 2793-2797] encode cytochrome P-450b whereas pcP-450pb2, a third cDNA whose nucleotide sequence differed slightly from that of the other two (six amino acid substitutions), encodes cytochrome P-450e. In addition to establishing the identity of these cloned cDNAs we provide direct evidence for seven additional amino acid differences between cytochromes P-450b and P-450e that occur beyond the region (Arg358) encoded by the cloned cDNA for cytochrome P-450e. Together, the amino acid sequences determined by micro sequence analysis and recombinant DNA techniques reveal 13 amino acid differences between these two isozymes. This report highlights the complementary nature of two different molecular approaches to elucidation of the amino acid sequences of isozymes with extensive structural homology. PMID:6572377

  5. Functional dissection of naturally occurring amino acid substitutions in eIF4E that confers recessive potyvirus resistance in plants.

    PubMed

    Yeam, Inhwa; Cavatorta, Jason R; Ripoll, Daniel R; Kang, Byoung-Cheorl; Jahn, Molly M

    2007-09-01

    Naturally existing variation in the eukaryotic translation initiation factor 4E (eIF4E) homolog encoded at the pvr1 locus in Capsicum results in recessively inherited resistance against several potyviruses. Previously reported data indicate that the physical interaction between Capsicum-eIF4E and the viral genome-linked protein (VPg) is required for the viral infection in the Capsicum-Tobacco etch virus (TEV) pathosystem. In this study, the potential structural role(s) of natural variation in the eIF4E protein encoded by recessive resistance alleles and their biological consequences have been assessed. Using high-resolution three-dimensional structural models based on the available crystallographic structures of eIF4E, we show that the amino acid substitution G107R, found in many recessive plant virus resistance genes encoding eIF4E, is predicted to result in a substantial modification in the protein binding pocket. The G107R change was shown to not only be responsible for the interruption of VPg binding in planta but also for the loss of cap binding ability in vitro, the principal function of eIF4E in the host. Overexpression of the Capsicum-eIF4E protein containing the G107R amino acid substitution in Solanum lycopersicum indicated that this polymorphism alone is sufficient for the acquisition of resistance against several TEV strains.

  6. Amino acid substitutions in mosaic penicillin-binding protein 2 associated with reduced susceptibility to cefixime in clinical isolates of Neisseria gonorrhoeae.

    PubMed

    Takahata, Sho; Senju, Nami; Osaki, Yumi; Yoshida, Takuji; Ida, Takashi

    2006-11-01

    The molecular mechanisms of reduced susceptibility to cefixime in clinical isolates of Neisseria gonorrhoeae, particularly amino acid substitutions in mosaic penicillin-binding protein 2 (PBP2), were examined. The complete sequence of ponA, penA, and por genes, encoding, respectively, PBP1, PBP2, and porin, were determined for 58 strains isolated in 2002 from Japan. Replacement of leucine 421 by proline in PBP1 and the mosaic-like structure of PBP2 were detected in 48 strains (82.8%) and 28 strains (48.3%), respectively. The presence of mosaic PBP2 was the main cause of the elevated cefixime MIC (4- to 64-fold). In order to identify the mutations responsible for the reduced susceptibility to cefixime in isolates with mosaic PBP2, penA genes with various mutations were transferred to a susceptible strain by genetic transformation. The susceptibility of partial recombinants and site-directed mutants revealed that the replacement of glycine 545 by serine (G545S) was the primary mutation, which led to a two- to fourfold increase in resistance to cephems. Replacement of isoleucine 312 by methionine (I312M) and valine 316 by threonine (V316T), in the presence of the G545S mutation, reduced susceptibility to cefixime, ceftibuten, and cefpodoxime by an additional fourfold. Therefore, three mutations (G545S, I312M, and V316T) in mosaic PBP2 were identified as the amino acid substitutions responsible for reduced susceptibility to cefixime in N. gonorrhoeae.

  7. Amino acid substitutions in GyrA and ParC are associated with fluoroquinolone resistance in Mycoplasma bovis isolates from Japanese dairy calves.

    PubMed

    Sato, Toyotaka; Okubo, Torahiko; Usui, Masaru; Higuchi, Hidetoshi; Tamura, Yutaka

    2013-01-01

    We investigated the contribution of quinolone resistance-determining region (QRDR) mutations to fluoroquinolone (enrofloxacin, orbifloxacin and danofloxacin) susceptibility in 58 Mycoplasma bovis isolates from dairy cattle in Japan. Fluoroquinolone non-resistant isolates (fluoroquinolone-MICs≤2 μg/ml) possessed no QRDR mutations (19 isolates) or Ser83Leu in GyrA (32 isolates). Fluoroquinolone-resistant isolates (fluoroquinolone-MICs≥4 μg/ml) possessed Ser83Leu in GyrA and Ser81Pro in ParC (3 isolates) or Ser83Phe in GyrA and Ser80Ile in ParC (4 isolates). Laboratory-derived fluoroquinolone-resistant mutants selected from 2 isolates (possessing Ser83Leu in GyrA) had an amino acid substitution in ParC at the same position (Ser80Ile or Ser81Tyr) as fluoroquinolone-resistant isolates, suggesting a concurrent amino acid substitution in ParC (Ser80 or Ser81) is important in fluoroquinolone resistance in M. bovis isolates.

  8. Amino acid ratings of different forms of infant formulas based on varying degrees of processing.

    PubMed

    Sarwar, G

    1991-01-01

    Amino acid profiles, protein digestibility and/or amino acid bioavailability for the various forms (powder, liquid concentrate, ready-to-use, etc.) of infant formulas (involving varying degrees of heat processing during preparation) have been determined. Amino acid scores (based on the single most limiting amino acid) were calculated by comparing the essential amino acid data with that of human milk. Amino acid scores were multiplied by total protein (g/100 kcal) to obtain amino acid ratings, which take into account both quality and quantity of protein. Amino acid scores for milk- and soy-based formulas ranged from 49 to 90 and 59 to 81%, respectively, due to deficiencies in methionine plus cystine and/or tryptophan. The deficiency in the limiting amino acids was more marked in liquid concentrate than powder prepared by the same manufacturer. Because of significantly higher total protein contents (g/100 kcal) of soy- (2.65-3.68) and milk-based (2.20-2.95) formulas compared to human milk (1.5), the relative amino acid ratings (human milk = 100) of all formulas except two milk-based liquid concentrates and one ready-to-feed (with values of 77-87%) were greater than 100%. When corrected for protein digestibility, the relative amino acid ratings for all four liquid concentrates were less than 100%. Lower levels of digestible protein and bioavailable amino acids in liquid concentrate compared with powder (prepared by the same manufacturer) would suggest that inferior protein quality of liquid concentrates may be due to more severe heat treatment involved in their preparation.

  9. Support Effects on Bronsted acid site densities and alcohol dehydration turnover rates on tungsten oxide domains

    SciTech Connect

    Macht, Josef; Baertsch, Chelsey D.; May-Lozano, Marcos; Soled, Stuart L.; Wang, Yong; Iglesia, Enrique

    2005-03-01

    Initial activity and acid site density of several WAl, WSi (MCM41) and one WSn sample were determined. Trans/cis 2-butene selectivity is dependent on the support. Presumably, these differences are due to subtle differences in base strengths. 2-Butanol dehydration rates (per W-atom) reached maximum values at intermediate WOx surface densities on WAl, as reported for 2-butanol dehydration reactions on WZr. Titration results indicate that Bronsted acid sites are required for 2-butanol dehydration on WAl, WSi and WSn. UV-visible studies suggest that WAl is much more difficult to reduce than WZr. The detection of reduced centers on WAl, the number of which correlates to Bronsted acid site density and catalyst activity, as well as the temperature dependence of Bronsted acid site density indicate the in-situ formation of these active sites. We infer that this mechanism is common among all supported WOx samples described in this study. Turnover rates are a function of Bronsted acid site density only. High acid site densities lead to high turnover rates. Higher active site densities may cause stronger conjugate bases, as a higher electron density has to be stabilized, and thus weaker acidity, enabling a faster rate of product desorption. The maximum achievable active site density is dependent on the support. WZr reaches a higher active site density than WAl.

  10. Formation rates, stability and reactivity of sulfuric acid - amine clusters predicted by computational chemistry

    NASA Astrophysics Data System (ADS)

    Kurtén, Theo; Ortega, Ismael; Kupiainen, Oona; Olenius, Tinja; Loukonen, Ville; Reiman, Heidi; McGrath, Matthew; Vehkamäki, Hanna

    2013-04-01

    Despite the importance of atmospheric particle formation for both climate and air quality, both experiments and non-empirical models using e.g. sulfuric acid, ammonia and water as condensing vapors have so far been unable to reproduce atmospheric observations using realistic trace gas concentrations. Recent experimental and theoretical evidence has shown that this mystery is likely resolved by amines. Combining first-principles evaporation rates for sulfuric acid - dimethylamine clusters with cluster kinetic modeling, we show that even sub-ppt concentrations of amines, together with atmospherically realistic concentrations of sulfuric acid, result in formation rates close to those observed in the atmosphere. Our simulated cluster formation rates are also close to, though somewhat larger than, those measured at the CLOUD experiment in CERN for both sulfuric acid - ammonia and sulfuric acid - dimethylamine systems. A sensitivity analysis indicates that the remaining discrepancy for the sulfuric acid - amine particle formation rates is likely caused by steric hindrances to cluster formation (due to alkyl groups of the amine molecules) rather than by significant errors in the evaporation rates. First-principles molecular dynamic and reaction kinetic modeling shed further light on the microscopic physics and chemistry of sulfuric acid - amine clusters. For example, while the number and type of hydrogen bonds in the clusters typically reach their equilibrium values on a picosecond timescale, and the overall bonding patterns predicted by traditional "static" quantum chemical calculations seem to be stable, the individual atoms participating in the hydrogen bonds continuously change at atmospherically realistic temperatures. From a chemical reactivity perspective, we have also discovered a surprising phenomenon: clustering with sulfuric acid molecules slightly increases the activation energy required for the abstraction of alkyl hydrogens from amine molecules. This implies

  11. ESTIMATION OF PHOSPHATE ESTER HYDROLYSIS RATE CONSTANTS. II. ACID AND GENERAL BASE CATALYZED HYDROLYSIS

    EPA Science Inventory

    SPARC (SPARC Performs Automated Reasoning in Chemistry) chemical reactivity models were extended to calculate acid and neutral hydrolysis rate constants of phosphate esters in water. The rate is calculated from the energy difference between the initial and transition states of a ...

  12. Substituted benzeneseleninic acids as bidentate ligands. Synthesis and spectroscopic studies of manganese(II) and iron(II) complexes

    NASA Astrophysics Data System (ADS)

    Candrini, Giovanni; Malavasi, Wanda; Preti, Carlo; Tosi, Giuseppe; Zannini, Paolo

    The para- and meta-substituted seleninato anion, XC 6H 4SeO -2, forms complexes with manganese(II) and iron(II) of the type [M(XC 6H 4SeO 2) 2(H 2O) 2], which have been shown to contain the bidentate ligand in seleninato- O, O' derivatives, the water molecules being coordinated to the metals. From the electronic absorption spectra and from the magnetic susceptibility data we have proposed for all the complexes a distorted octahedral D4 h symmetry. The structure of the anhydrous para- and meta-substituted benzeneseleninato complexes of manganese(II) and iron(II) have been investigated by means of electrical conductance measurements, spectral (electronic and i.r.) studies and magnetic susceptibility measurements. The anhydrous complexes are always of the seleninato- O, O' type with the ligands tetrahedrally coordinated to the central atom. The wavelengths of the principal absorption peaks have been accounted for quantitatively in terms of the crystal field theory for manganese(II) derivatives. The nephelauxetic parameters are all indicative of an appreciable metal-ligand covalency.

  13. A single amino acid substitution makes ERK2 susceptible to pyridinyl imidazole inhibitors of p38 MAP kinase.

    PubMed Central

    Fox, T.; Coll, J. T.; Xie, X.; Ford, P. J.; Germann, U. A.; Porter, M. D.; Pazhanisamy, S.; Fleming, M. A.; Galullo, V.; Su, M. S.; Wilson, K. P.

    1998-01-01

    Mitogen-activated protein (MAP) kinases are serine/threonine kinases that mediate intracellular signal transduction pathways. Pyridinyl imidazole compounds block pro-inflammatory cytokine production and are specific p38 kinase inhibitors. ERK2 is related to p38 in sequence and structure, but is not inhibited by pyridinyl imidazole inhibitors. Crystal structures of two pyridinyl imidazoles complexed with p38 revealed these compounds bind in the ATP site. Mutagenesis data suggested a single residue difference at threonine 106 between p38 and other MAP kinases is sufficient to confer selectivity of pyridinyl imidazoles. We have changed the equivalent residue in human ERK2, Q105, into threonine and alanine, and substituted four additional ATP binding site residues. The single residue change Q105A in ERK2 enhances the binding of SB202190 at least 25,000-fold compared to wild-type ERK2. We report enzymatic analyses of wild-type ERK2 and the mutant proteins, and the crystal structure of a pyridinyl imidazole, SB203580, bound to an ERK2 pentamutant, I103L, Q105T, D106H, E109G. T110A. These ATP binding site substitutions induce low nanomolar sensitivity to pyridinyl imidazoles. Furthermore, we identified 5-iodotubercidin as a potent ERK2 inhibitor, which may help reveal the role of ERK2 in cell proliferation. PMID:9827991

  14. Effect of partial substitution of NaCl with KCl on Halloumi cheese during storage: chemical composition, lactic bacterial count, and organic acids production.

    PubMed

    Ayyash, Mutamed M; Shah, Nagendra P

    2010-08-01

    The effect of partial substitution of NaCl with KCl on chemical composition, lactic bacterial count, and organic acids profile of Halloumi cheese was investigated. Halloumi cheeses were made and kept in 4 different brine solutions at 18% including NaCl only (HA), 3NaCl : 1KCl (HB), 1NaCl : 1KCl (HC), and 1NaCl : 3KCl (HD) and then stored at 4 degrees C for 56 d. No significant effect was observed between control and experimental cheeses in terms of moisture, fat, protein, lactic bacterial count, and pH values at the same storage period. There was a significant difference in ash, sodium, and potassium contents among experimental cheeses at the same storage period. Ash, sodium, and potassium contents increased significantly during storage at same salt treatment. There was no significant difference in lactic and citric acid contents among experimental cheeses and that of the control. In contrary, there was a significant difference in acetic acid among experimental cheeses. A strong positive correlation was observed between ash, Na, and K contents. An inverse correlation between organic acids and both Na and K contents was also observed.

  15. The effect of limestone treatments on the rate of acid generation from pyritic mine gangue.

    PubMed

    Burt, R A; Caruccio, F T

    1986-09-01

    Surface water enters the Haile Gold Mine, Lancaster County, South Carolina by means of a small stream and is ponded behind a dam and in an abandoned pit. This water is affected by acidic drainage. In spite of the large exposures of potentially acid producing pyritic rock, the flux of acid to the water is relatively low. Nevertheless, the resulting pH values of the mine water are low (around 3.5) due to negligible buffering capacity. In view of the observed low release of acidity, the potential for acid drainage abatement by limestone ameliorants appears feasible.This study investigated the effects of limestone treatment on acid generation rates of the Haile mine pyritic rocks through a series of leaching experiments. Below a critical alkalinity threshold value, solutions of dissolved limestone were found consistently to accelerate the rate of pyrite oxidation by varying degrees. The oxidation rates were further accelerated by admixing solid limestone with the pyritic rock. However, after a period of about a month, the pyrite oxidation rate of the admixed samples declined to a level lower than that of untreated pyrite. Leachates produced by the pyrite and limestone mixtures contained little if any iron. Further, in the mixtures, an alteration of the pyrite surface was apparent.The observed behaviour of the treated pyrite appears to be related to the immersion of the pyrite grains within a high alkalinity/high pH environment. The high pH increases the rate of oxidation of ferrous iron which results in a higher concentration of ferric iron at the pyrite surface. This, in turn, increases the rate of pyrite oxidation. Above a threshold alkalinity value, the precipitation of hydrous iron oxides at the pyrite surface eventually outpaces acid generation and coats the pyrite surface, retarding the rate of pyrite oxidation. PMID:24214013

  16. Fast evolving 18S rRNA sequences from Solenogastres (Mollusca) resist standard PCR amplification and give new insights into mollusk substitution rate heterogeneity

    PubMed Central

    2010-01-01

    Background The 18S rRNA gene is one of the most important molecular markers, used in diverse applications such as molecular phylogenetic analyses and biodiversity screening. The Mollusca is the second largest phylum within the animal kingdom and mollusks show an outstanding high diversity in body plans and ecological adaptations. Although an enormous amount of 18S data is available for higher mollusks, data on some early branching lineages are still limited. Despite of some partial success in obtaining these data from Solenogastres, by some regarded to be the most "basal" mollusks, this taxon still remained problematic due to contamination with food organisms and general amplification difficulties. Results We report here the first authentic 18S genes of three Solenogastres species (Mollusca), each possessing a unique sequence composition with regions conspicuously rich in guanine and cytosine. For these GC-rich regions we calculated strong secondary structures. The observed high intra-molecular forces hamper standard amplification and appear to increase formation of chimerical sequences caused by contaminating foreign DNAs from potential prey organisms. In our analyses, contamination was avoided by using RNA as a template. Indication for contamination of previously published Solenogastres sequences is presented. Detailed phylogenetic analyses were conducted using RNA specific models that account for compensatory substitutions in stem regions. Conclusions The extreme morphological diversity of mollusks is mirrored in the molecular 18S data and shows elevated substitution rates mainly in three higher taxa: true limpets (Patellogastropoda), Cephalopoda and Solenogastres. Our phylogenetic tree based on 123 species, including representatives of all mollusk classes, shows limited resolution at the class level but illustrates the pitfalls of artificial groupings formed due to shared biased sequence composition. PMID:20214780

  17. [Bicycle test: measure of anaerobic power, heart rate and blood lactic acid].

    PubMed

    Faye, J; Fall, A; Seck, D; Badji, L; Faye, E M; Cisse, F

    2002-01-01

    Seven sportsmen, 100 meters and 400 meters runners are submitted to an effort test of 30 seconds. The subjects are on average 23.7 +/-2 years old. The purpose of our work is to study on the one hand the evlution of the anaerobic power. the heart rate and the lactic acid in blood during and after a bicycle test. and their relation, and on the other hand. to know the suitable pratical importance of the heart rate and the lactic acid in blood in connection with the intermittent efforts recovery aiming the anaerobic power developpement. These physiological parameters have been measured by a Monark bicycle 864, a sport-tester PE 3000 and a spectrophotometer JASCO 7800 UV/VIS. The power and the heart rate increase quickly in the 5 first seconds. Our subjects reach their average maximal anaerobic power at the 10th second, and then this power decreases progressively, while the heart rate continues to increase, without being maximal at the end of te test. Five minutes later it decreases in a half, while the lactic acid level calculated at the 30th second is continuing significantly. We have not found a significant relation between the measured parameters at the test stopping and during the recovery period (except between the lactic acid and the recovery index of the heart rate at the 25th minute). The lactic acid in blood would inform better about a good recovery during an interval training effort.

  18. [Studies of gene regulation of de novo biosynthetic pathway of purine in Salmonella typhimurium. X. Isolation of purR(am) mutants and preliminary studies of amino acid substitution].

    PubMed

    Zhang, H S; Wang, A Q

    2000-01-01

    Starting from a super-repressed mutant of purR, 3-18, 439 independent candidates of purR- mutants were isolated by using NCE selecting plate with lactose as sole carbon source. Among these mutants. 11 amber mutants were detected by introducing a tRNA suppressor gene. Cotransduction analysis proved that the amber mutation sites of 11 amber mutants all located on purR. Amino acid substitution experiments were performed with three tRNA suppressors, supD, supE and supF, for each purR(am). The results showed that the same amino acid substitution occurred in different site of PurR protein could result in varied effects on purR function; different amino acid substitution occurred at the same position of PurR protein also could produced varied effects on purR function.

  19. Site-directed substitution of Ser1406 of hamster CAD with glutamic acid alters allosteric regulation of carbamyl phosphate synthetase II.

    PubMed

    Banerjei, L C; Davidson, J N

    1997-01-01

    Ser1406 of the allosteric region of the hamster CAD enzyme, carbamyl phosphate synthetase II (CPSase), is known to be phosphorylated in vitro by cAMP-dependent protein kinase (PKA). Metabolic labeling experiments described here demonstrate that CAD is phosphorylated in somatic cells in culture. Phosphorylation is stimulated by treating cells with 8-bromo-cAMP, a PKA activator. The stimulation is essentially prevented by pretreatment with H-89, a PKA specific inhibitor. Substitution of Ser1406 with alanine results in an enzyme with kinetics and allosteric regulation indistinguishable from unsubstituted CAD. However, substitution to glutamic acid increases CPSase activity by reducing the apparent Km (ATP). The UTP concentration required to give 50% inhibition is increased rendering this altered enzyme significantly less sensitive to feedback inhibition, but allosteric activation by PRPP is unaffected. While these data do not prove that Ser1406 is phosphorylated in vivo, they do indicate that a specific alteration at this residue can affect allosteric regulation. PMID:9218000

  20. Quantitative Structure-Property Relationship (QSPR) Models for a Local Quantum Descriptor: Investigation of the 4- and 3-Substituted-Cinnamic Acid Esterification.

    PubMed

    Rodrigues-Santos, Cláudio E; Echevarria, Aurea; Sant'Anna, Carlos M R; Bitencourt, Thiago B; Nascimento, Maria G; Bauerfeldt, Glauco F

    2015-09-22

    In this work, the theoretical description of the 4- and 3-substituted-cinnamic acid esterification with different electron donating and electron withdrawing groups was performed at the B3LYP and M06-2X levels, as a two-step process: the O-protonation and the nucleophile attack by ethanol. In parallel, an experimental work devoted to the synthesis and characterization of the substituted-cinnamate esters has also been performed. In order to quantify the substituents effects, quantitative structure-property relationship (QSPR) models based on the atomic charges, Fukui functions and the Frontier Effective-for-Reaction Molecular Orbitals (FERMO) energies were investigated. In fact, the Fukui functions, ƒ⁺C and ƒ(-)O, indicated poor correlations for each individual step, and in contrast with the general literature, the O-protonation step is affected both by the FERMO energies and the O-charges of the carbonyl group. Since the process was shown to not be totally described by either charge- or frontier-orbitals, it is proposed to be frontier-charge-miscere controlled. Moreover, the observed trend for the experimental reaction yields suggests that the electron withdrawing groups favor the reaction and the same was observed for Step 2, which can thus be pointed out as the determining step.

  1. Amino acid substitution at peptide-binding pockets of HLA class I molecules increases risk of severe acute GVHD and mortality

    PubMed Central

    Wang, Tao; Haagenson, Michael; Spellman, Stephen R.; Askar, Medhat; Battiwalla, Minoo; Baxter-Lowe, Lee Ann; Bitan, Menachem; Fernandez-Viña, Marcelo; Gandhi, Manish; Jakubowski, Ann A.; Maiers, Martin; Marino, Susana R.; Marsh, Steven G. E.; Oudshoorn, Machteld; Palmer, Jeanne; Prasad, Vinod K.; Reddy, Vijay; Ringden, Olle; Saber, Wael; Santarone, Stella; Schultz, Kirk R.; Setterholm, Michelle; Trachtenberg, Elizabeth; Turner, E. Victoria; Woolfrey, Ann E.; Lee, Stephanie J.; Anasetti, Claudio

    2013-01-01

    HLA disparity has a negative impact on the outcomes of hematopoietic cell transplantation (HCT). We studied the independent impact of amino acid substitution (AAS) at peptide-binding positions 9, 99, 116, and 156, and killer immunoglobulin-like receptor binding position 77 of HLA-A, B, or C, on the risks for grade 3-4 acute graft-versus-host disease (GVHD), chronic GVHD, treatment-related mortality (TRM), relapse, and overall survival. In multivariate analysis, a mismatch at HLA-C position 116 was associated with increased risk for severe acute GVHD (hazard ratio [HR] = 1.45, 95% confidence interval [CI] = 1.15-1.82, P = .0016). Mismatch at HLA-C position 99 was associated with increased transplant-related mortality (HR = 1.37, 95% CI = 1.1-1.69, P = .0038). Mismatch at HLA-B position 9 was associated with increased chronic GVHD (HR = 2.28, 95% CI = 1.36-3.82, P = .0018). No AAS were significantly associated with outcome at HLA-A. Specific AAS pair combinations with a frequency >30 were tested for association with HCT outcomes. Cysteine to tyrosine substitution at position 99 of HLA-C was associated with increased TRM (HR = 1.78, 95% = CI 1.27-2.51, P = .0009). These results demonstrate that donor-recipient mismatch for certain peptide-binding residues of the HLA class I molecule is associated with increased risk for acute and chronic GVHD and death. PMID:23982174

  2. A single amino acid substitution modulates low-pH-triggered membrane fusion of GP64 protein in Autographa californica and Bombyx mori nucleopolyhedroviruses

    SciTech Connect

    Katou, Yasuhiro; Yamada, Hayato; Ikeda, Motoko; Kobayashi, Michihiro

    2010-09-01

    We have previously shown that budded viruses of Bombyx mori nucleopolyhedrovirus (BmNPV) enter the cell cytoplasm but do not migrate into the nuclei of non-permissive Sf9 cells that support a high titer of Autographa californica multicapsid nucleopolyhedrovirus (AcMNPV) multiplication. Here we show, using the syncytium formation assay, that low-pH-triggered membrane fusion of BmNPV GP64 protein (Bm-GP64) is significantly lower than that of AcMNPV GP64 protein (Ac-GP64). Mutational analyses of GP64 proteins revealed that a single amino acid substitution between Ac-GP64 H155 and Bm-GP64 Y153 can have significant positive or negative effects on membrane fusion activity. Studies using bacmid-based GP64 recombinant AcMNPV harboring point-mutated ac-gp64 and bm-gp64 genes showed that Ac-GP64 H155Y and Bm-GP64 Y153H substitutions decreased and increased, respectively, the multiplication and cell-to-cell spread of progeny viruses. These results indicate that Ac-GP64 H155 facilitates the low-pH-triggered membrane fusion reaction between virus envelopes and endosomal membranes.

  3. Effects of two novel amino acid substitutions on the penicillin binding properties of the PBP5 C‑terminal from Enterococcus faecium.

    PubMed

    Zhou, Chengjiang; Niu, Haiying; Yu, Hui; Zhou, Lishe; Wang, Zhanli

    2015-10-01

    The low‑affinity penicillin‑binding protein (PBP)5 is responsible for resistance to β‑lactam antibiotics in Enterococcus faecium. (E. faecium). In order to evaluate more fully the potential of this species for the development of resistance to β-lactam antibiotics, the present study aimed to examine the extent of penicillin-binding protein (PBP) variations in a collection of clinical E. faecium isolates. In the present study, the C‑terminal domain of PBP5 (PBP5‑CD) of 13 penicillin‑resistant clinical isolates of E. faecium were sequenced and the correlation between penicillin resistance and particular amino acid changes were analyzed. The present study identified for the first time, to the best of our knowledge, two novel substitutions (Tyr460Phe and Ala462Thr or Val462Thr) of E. faecium PBP5‑CD. The covalent interaction between penicillin and PBP5‑CD was also investigated using homology modeling and molecular docking methods. The theoretical calculation revealed that Phe460 and Thr462 were involved in penicillin binding, suggesting that substitutions at these positions exert effects on the affinity for penicillin, and this increased affinity translates into lower resistance in vitro.

  4. Ultra-superovulation for the CRISPR-Cas9-mediated production of gene-knockout, single-amino-acid-substituted, and floxed mice.

    PubMed

    Nakagawa, Yoshiko; Sakuma, Tetsushi; Nishimichi, Norihisa; Yokosaki, Yasuyuki; Yanaka, Noriyuki; Takeo, Toru; Nakagata, Naomi; Yamamoto, Takashi

    2016-01-01

    Current advances in producing genetically modified mice using genome-editing technologies have indicated the need for improvement of limiting factors including zygote collection for microinjection and their cryopreservation. Recently, we developed a novel superovulation technique using inhibin antiserum and equine chorionic gonadotropin to promote follicle growth. This method enabled the increased production of fertilized oocytes via in vitro fertilization compared with the conventional superovulation method. Here, we verify that the ultra-superovulation technique can be used for the efficient generation of clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-mediated knockout mice by microinjection of plasmid vector or ribonucleoprotein into zygotes. We also investigated whether single-amino-acid-substituted mice and conditional knockout mice could be generated. Founder mice bearing base substitutions were generated more efficiently by co-microinjection of Cas9 protein, a guide RNA and single-stranded oligodeoxynucleotide (ssODN) than by plasmid microinjection with ssODN. The conditional allele was successfully introduced by the one-step insertion of an ssODN designed to carry an exon flanked by two loxP sequences and homology arms using a double-cut CRISPR-Cas9 strategy. Our study presents a useful method for the CRISPR-Cas9-based generation of genetically modified mice from the viewpoints of animal welfare and work efficiency. PMID:27387532

  5. Ultra-superovulation for the CRISPR-Cas9-mediated production of gene-knockout, single-amino-acid-substituted, and floxed mice.

    PubMed

    Nakagawa, Yoshiko; Sakuma, Tetsushi; Nishimichi, Norihisa; Yokosaki, Yasuyuki; Yanaka, Noriyuki; Takeo, Toru; Nakagata, Naomi; Yamamoto, Takashi

    2016-08-15

    Current advances in producing genetically modified mice using genome-editing technologies have indicated the need for improvement of limiting factors including zygote collection for microinjection and their cryopreservation. Recently, we developed a novel superovulation technique using inhibin antiserum and equine chorionic gonadotropin to promote follicle growth. This method enabled the increased production of fertilized oocytes via in vitro fertilization compared with the conventional superovulation method. Here, we verify that the ultra-superovulation technique can be used for the efficient generation of clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-mediated knockout mice by microinjection of plasmid vector or ribonucleoprotein into zygotes. We also investigated whether single-amino-acid-substituted mice and conditional knockout mice could be generated. Founder mice bearing base substitutions were generated more efficiently by co-microinjection of Cas9 protein, a guide RNA and single-stranded oligodeoxynucleotide (ssODN) than by plasmid microinjection with ssODN. The conditional allele was successfully introduced by the one-step insertion of an ssODN designed to carry an exon flanked by two loxP sequences and homology arms using a double-cut CRISPR-Cas9 strategy. Our study presents a useful method for the CRISPR-Cas9-based generation of genetically modified mice from the viewpoints of animal welfare and work efficiency.

  6. Ultra-superovulation for the CRISPR-Cas9-mediated production of gene-knockout, single-amino-acid-substituted, and floxed mice

    PubMed Central

    Nakagawa, Yoshiko; Nishimichi, Norihisa; Yokosaki, Yasuyuki; Yanaka, Noriyuki; Takeo, Toru; Nakagata, Naomi; Yamamoto, Takashi

    2016-01-01

    ABSTRACT Current advances in producing genetically modified mice using genome-editing technologies have indicated the need for improvement of limiting factors including zygote collection for microinjection and their cryopreservation. Recently, we developed a novel superovulation technique using inhibin antiserum and equine chorionic gonadotropin to promote follicle growth. This method enabled the increased production of fertilized oocytes via in vitro fertilization compared with the conventional superovulation method. Here, we verify that the ultra-superovulation technique can be used for the efficient generation of clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR-associated protein 9 (Cas9)-mediated knockout mice by microinjection of plasmid vector or ribonucleoprotein into zygotes. We also investigated whether single-amino-acid-substituted mice and conditional knockout mice could be generated. Founder mice bearing base substitutions were generated more efficiently by co-microinjection of Cas9 protein, a guide RNA and single-stranded oligodeoxynucleotide (ssODN) than by plasmid microinjection with ssODN. The conditional allele was successfully introduced by the one-step insertion of an ssODN designed to carry an exon flanked by two loxP sequences and homology arms using a double-cut CRISPR-Cas9 strategy. Our study presents a useful method for the CRISPR-Cas9-based generation of genetically modified mice from the viewpoints of animal welfare and work efficiency. PMID:27387532

  7. Effects of salt concentration on the reaction rate of Glc with amino acids, peptides, and proteins.

    PubMed

    Yamaguchi, Keiko; Noumi, Yuri; Nakajima, Katsumi; Nagatsuka, Chiharu; Aizawa, Haruko; Nakawaki, Rie; Mizude, Eri; Otsuka, Yuzuru; Homma, Takeshi; Chuyen, Nguyen Van

    2009-11-01

    The reaction between the amino group and the carbonyl group is important in food quality control. Furthermore, advanced glycation end products from foods are considered to relate to aging and diabetes. Thus, it is important to control this reaction. In this study, we investigated the effects of salt concentration on the rates of browning reaction of amino acid, peptides, and proteins. A high concentration of sodium chloride retarded the reaction rate of Glc with amino acids as measured with the absorbance at 470 nm, but did not change the browning rate of Glc with peptides. On the other hand, sodium chloride retarded the browning reaction rate of proteins as measured with polymerization degree or by the loss of Lys. It is hoped that the results of this study will be applied in the control of amino-carbonyl reaction rates in the food industry. PMID:19897911

  8. Density functional theory study of hydrogen atom abstraction from a series of para-substituted phenols: why is the Hammett σ(p)+ constant able to represent radical reaction rates?

    PubMed

    Yoshida, Tatsusada; Hirozumi, Koji; Harada, Masataka; Hitaoka, Seiji; Chuman, Hiroshi

    2011-06-01

    The rate of hydrogen atom abstraction from phenolic compounds by a radical is known to be often linear with the Hammett substitution constant σ(+), defined using the S(N)1 solvolysis rates of substituted cumyl chlorides. Nevertheless, a physicochemical reason for the above "empirical fact" has not been fully revealed. The transition states of complexes between the 2,2-diphenyl-1-picrylhydrazyl radical (dpph·) and a series of para-substituted phenols were determined by DFT (Density Functional Theory) calculations, and then the activation energy as well as the homolytic bond dissociation energy of the O-H bond and charge distribution in the transition state were calculated. The heterolytic bond dissociation energy of the C-Cl bond and charge distribution in the corresponding para-substituted cumyl chlorides were calculated in parallel. Excellent correlations among σ(+), charge distribution, and activation and bond dissociation energies revealed quantitatively that there is a strong similarity between the two reactions, showing that the electron-deficiency of the π-electron system conjugated with a substituent plays a crucial role in determining rates of the two reactions. The results provide a new insight into and physicochemical understanding of σ(+) in the hydrogen abstraction from substituted phenols by a radical.

  9. Probing the Active Center of Benzaldehyde Lyase with Substitutions and the Pseudosubstrate Analogue Benzoylphosphonic Acid Methyl Ester

    SciTech Connect

    Brandt, Gabriel S.; Nemeria, Natalia; Chakraborty, Sumit; McLeish, Michael J.; Yep, Alejandra; Kenyon, George L.; Petsko, Gregory A.; Jordan, Frank; Ringe, Dagmar

    2008-07-28

    Benzaldehyde lyase (BAL) catalyzes the reversible cleavage of (R)-benzoin to benzaldehyde utilizing thiamin diphosphate and Mg{sup 2+} as cofactors. The enzyme is important for the chemoenzymatic synthesis of a wide range of compounds via its carboligation reaction mechanism. In addition to its principal functions, BAL can slowly decarboxylate aromatic amino acids such as benzoylformic acid. It is also intriguing mechanistically due to the paucity of acid-base residues at the active center that can participate in proton transfer steps thought to be necessary for these types of reactions. Here methyl benzoylphosphonate, an excellent electrostatic analogue of benzoylformic acid, is used to probe the mechanism of benzaldehyde lyase. The structure of benzaldehyde lyase in its covalent complex with methyl benzoylphosphonate was determined to 2.49 {angstrom} (Protein Data Bank entry 3D7K) and represents the first structure of this enzyme with a compound bound in the active site. No large structural reorganization was detected compared to the complex of the enzyme with thiamin diphosphate. The configuration of the predecarboxylation thiamin-bound intermediate was clarified by the structure. Both spectroscopic and X-ray structural studies are consistent with inhibition resulting from the binding of MBP to the thiamin diphosphate in the active centers. We also delineated the role of His29 (the sole potential acid-base catalyst in the active site other than the highly conserved Glu50) and Trp163 in cofactor activation and catalysis by benzaldehyde lyase.

  10. Probing the active center of benzaldehyde lyase with substitutions and the pseudosubstrate analogue benzoylphosphonic acid methyl ester.

    PubMed

    Brandt, Gabriel S; Nemeria, Natalia; Chakraborty, Sumit; McLeish, Michael J; Yep, Alejandra; Kenyon, George L; Petsko, Gregory A; Jordan, Frank; Ringe, Dagmar

    2008-07-22

    Benzaldehyde lyase (BAL) catalyzes the reversible cleavage of ( R)-benzoin to benzaldehyde utilizing thiamin diphosphate and Mg (2+) as cofactors. The enzyme is important for the chemoenzymatic synthesis of a wide range of compounds via its carboligation reaction mechanism. In addition to its principal functions, BAL can slowly decarboxylate aromatic amino acids such as benzoylformic acid. It is also intriguing mechanistically due to the paucity of acid-base residues at the active center that can participate in proton transfer steps thought to be necessary for these types of reactions. Here methyl benzoylphosphonate, an excellent electrostatic analogue of benzoylformic acid, is used to probe the mechanism of benzaldehyde lyase. The structure of benzaldehyde lyase in its covalent complex with methyl benzoylphosphonate was determined to 2.49 A (Protein Data Bank entry 3D7K ) and represents the first structure of this enzyme with a compound bound in the active site. No large structural reorganization was detected compared to the complex of the enzyme with thiamin diphosphate. The configuration of the predecarboxylation thiamin-bound intermediate was clarified by the structure. Both spectroscopic and X-ray structural studies are consistent with inhibition resulting from the binding of MBP to the thiamin diphosphate in the active centers. We also delineated the role of His29 (the sole potential acid-base catalyst in the active site other than the highly conserved Glu50) and Trp163 in cofactor activation and catalysis by benzaldehyde lyase.

  11. Rational Design, Synthesis, and Preliminary Structure-Activity Relationships of α-Substituted-2-Phenylcyclopropane Carboxylic Acids as Inhibitors of Salmonella typhimurium O-Acetylserine Sulfhydrylase.

    PubMed

    Pieroni, Marco; Annunziato, Giannamaria; Beato, Claudia; Wouters, Randy; Benoni, Roberto; Campanini, Barbara; Pertinhez, Thelma A; Bettati, Stefano; Mozzarelli, Andrea; Costantino, Gabriele

    2016-03-24

    Cysteine is a building block for several biomolecules that are crucial for living organisms. The last step of cysteine biosynthesis is catalyzed by O-acetylserine sulfydrylase (OASS), a highly conserved pyridoxal 5'-phosphate (PLP)-dependent enzyme, present in different isoforms in bacteria, plants, and nematodes, but absent in mammals. Beside the biosynthesis of cysteine, OASS exerts a series of "moonlighting" activities in bacteria, such as transcriptional regulation, contact-dependent growth inhibition, swarming motility, and induction of antibiotic resistance. Therefore, the discovery of molecules capable of inhibiting OASS would be a valuable tool to unravel how this protein affects the physiology of unicellular organisms. As a continuation of our efforts toward the synthesis of OASS inhibitors, in this work we have used a combination of computational and spectroscopic approaches to rationally design, synthesize, and test a series of substituted 2-phenylcyclopropane carboxylic acids that bind to the two S. typhymurium OASS isoforms at nanomolar concentrations.

  12. Long-chain ethers as solvents can amplify the enantioselectivity of the Carica papaya lipase-catalyzed transesterification of 2-(substituted phenoxy)propanoic acid esters.

    PubMed

    Miyazawa, Toshifumi; Iguchi, Wakana

    2013-10-01

    The enantioselectivity of the transesterification of the 2,2,2-trifluoroethyl esters of 2-(substituted phenoxy)propanoic acids, as catalyzed by the lipase from Carica papaya, was greatly improved by using long-chain ethers, such as di-n-hexyl ether, as solvents instead of the conventional diisopropyl ether. Thus, for example, the E value was enhanced from 21 [in diisopropyl ether (0.8 ml)] to 57 [in di-n-hexyl ether (0.8 ml)] in the reaction of 2,2,2-trifluoroethyl(RS)-2-phenoxypropanoate (0.1 mmol) with methanol (0.4 mmol) in the presence of the plant lipase preparation (10 mg); it was also improved from 13 (in diisopropyl ether) to 44 (in di-n-hexyl ether) in the reaction of 2,2,2-trifluoroethyl(RS)-2-(2-chlorophenoxy)propanoate with methanol under the same reaction conditions.

  13. Rational Design, Synthesis, and Preliminary Structure-Activity Relationships of α-Substituted-2-Phenylcyclopropane Carboxylic Acids as Inhibitors of Salmonella typhimurium O-Acetylserine Sulfhydrylase.

    PubMed

    Pieroni, Marco; Annunziato, Giannamaria; Beato, Claudia; Wouters, Randy; Benoni, Roberto; Campanini, Barbara; Pertinhez, Thelma A; Bettati, Stefano; Mozzarelli, Andrea; Costantino, Gabriele

    2016-03-24

    Cysteine is a building block for several biomolecules that are crucial for living organisms. The last step of cysteine biosynthesis is catalyzed by O-acetylserine sulfydrylase (OASS), a highly conserved pyridoxal 5'-phosphate (PLP)-dependent enzyme, present in different isoforms in bacteria, plants, and nematodes, but absent in mammals. Beside the biosynthesis of cysteine, OASS exerts a series of "moonlighting" activities in bacteria, such as transcriptional regulation, contact-dependent growth inhibition, swarming motility, and induction of antibiotic resistance. Therefore, the discovery of molecules capable of inhibiting OASS would be a valuable tool to unravel how this protein affects the physiology of unicellular organisms. As a continuation of our efforts toward the synthesis of OASS inhibitors, in this work we have used a combination of computational and spectroscopic approaches to rationally design, synthesize, and test a series of substituted 2-phenylcyclopropane carboxylic acids that bind to the two S. typhymurium OASS isoforms at nanomolar concentrations. PMID:26894308

  14. Synthesis and biological evaluation of novel N-substituted 1H-dibenzo[a,c]carbazole derivatives of dehydroabietic acid as potential antimicrobial agents.

    PubMed

    Gu, Wen; Qiao, Chao; Wang, Shi-Fa; Hao, Yun; Miao, Ting-Ting

    2014-01-01

    A series of new N-substituted 1H-dibenzo[a,c]carbazole derivatives were synthesized from dehydroabietic acid, and their structures were characterized by IR, (1)H NMR and HRMS spectral data. All compounds were evaluated for their antibacterial and antifungal activities against four bacteria (Bacillus subtilis, Staphylococcus aureus, Escherichia coli and Pseudomonas fluorescens) and three fungi (Candida albicans, Candida tropicalis and Aspergillus niger) by serial dilution technique. Some of the synthesized compounds displayed pronounced antimicrobial activity against tested strains with low MIC values ranging from 0.9 to 15.6μg/ml. Among them, compounds 6j and 6r exhibited potent inhibitory activity comparable to reference drugs amikacin and ketoconazole. PMID:24300736

  15. Silica sulfuric acid: a reusable solid catalyst for one pot synthesis of densely substituted pyrrole-fused isocoumarins under solvent-free conditions.

    PubMed

    Pathak, Sudipta; Debnath, Kamalesh; Pramanik, Animesh

    2013-01-01

    A convenient and efficient methodology for the synthesis of densely substituted pyrrole-fused isocoumarins, which employs solid-supported silica sulfuric acid (SSA) as catalyst, has been developed. When the mixture of ninhydrin adducts of acetylacetone/ethyl acetoacetate and primary amines was heated on the solid surface of SSA under solvent-free conditions, the pyrrole-fused isocoumarins were formed in good yields. This synthetic method has several advantages such as the employment of solvent-free reaction conditions without the use of any toxic reagents and metal catalysts, the ease of product isolation, the use of a recyclable catalyst, the low cost, the easy availability of the starting materials, and the excellent yields of products. PMID:24367398

  16. Effect of amino acid substitutions in a potential metal-binding site of AnfA on expression from the anfH promoter in Azotobacter vinelandii.

    PubMed

    Premakumar, R; Loveless, T M; Bishop, P E

    1994-10-01

    AnfA, an activator required for transcription of the structural genes encoding nitrogenase 3 (anfHDGK) in Azotobacter vinelandii, has a potential metal-binding site [(S19)H(C21)FTGE(C26)R] in its N terminus. Growth studies and expression of an anfH-lacZ fusion in mutants containing amino acid substitutions in this site indicate that Ser-19 is not required for AnfA activity whereas Cys-21 and Cys-26 are required. Residual expression of the anfH-lacZ fusion in AnfA- mutants was found to be due to activation by VnfA, the activator required for expression of genes encoding nitrogenase 2.

  17. Technical note: Synergistic effect of iodide ions on inhibitive performance of substituted dithiobiurets during corrosion of mild steel in hot hydrochloric acid

    SciTech Connect

    Quraishi, M.A.; Rawat, J.; Ajmal, M.

    1999-10-01

    Four substituted dithiobiurets (i.e., 1,5-diphenyl-2,4-dithiobiuret [DPDTB]; 1-anisidyl-5-phenyl 2,4-dithiodiuret [APDTB]; 1-tolyl-5-phenyl 2,4-dithiobiuret [TPDTB]; and 1-chlorophenyl-5-phenyl 2,4-dithiobiuret [CPDTB]) were synthesized to study their inhibiting effect on mild steel (MS) corrosion in 5 N hot hydrochloric acid (HCl). The synergistic effect of these compounds with potassium iodide (KI) was studied at different concentrations, temperatures, and immersion periods by weight loss and potentiodynamic polarization methods. All compounds showed good inhibition efficiency (IE) at all temperatures and showed the enhancement in IE with the addition of small amounts of KI. Potentiodynamic polarization studies showed that APDTB and DPDTB are predominantly cathodic inhibitors, whereas TPDTB and CPDTB are mixed inhibitors. The adsorption of all these compounds followed Temkin's adsorption isotherm.

  18. The Amino Acid Substitution Q65H in the 2C Protein of Swine Vesicular Disease Virus Confers Resistance to Golgi Disrupting Drugs.

    PubMed

    Vázquez-Calvo, Ángela; Caridi, Flavia; González-Magaldi, Mónica; Saiz, Juan-Carlos; Sobrino, Francisco; Martín-Acebes, Miguel A

    2016-01-01

    Swine vesicular disease virus (SVDV) is a porcine pathogen and a member of the species Enterovirus B within the Picornaviridae family. Brefeldin A (BFA) is an inhibitor of guanine nucleotide exchange factors of Arf proteins that induces Golgi complex disassembly and alters the cellular secretory pathway. Since BFA has been shown to inhibit the RNA replication of different enteroviruses, including SVDV, we have analyzed the effect of BFA and of golgicide A (GCA), another Golgi disrupting drug, on SVDV multiplication. BFA and GCA similarly inhibited SVDV production. To investigate the molecular basis of the antiviral effect of BFA, SVDV mutants with increased resistance to BFA were isolated. A single amino acid substitution, Q65H, in the non-structural protein 2C was found to be responsible for increased resistance to BFA. These results provide new insight into the relationship of enteroviruses with the components of the secretory pathway and on the role of SVDV 2C protein in this process.

  19. Single-channel studies on linear gramicidins with altered amino acid sequences. A comparison of phenylalanine, tryptophane, and tyrosine substitutions at positions 1 and 11.

    PubMed Central

    Mazet, J L; Andersen, O S; Koeppe, R E

    1984-01-01

    The relation between chemical structure and permeability characteristics of transmembrane channels has been investigated with the linear gramicidins (A, B, and C), where the amino acid at position 1 was chemically replaced by phenylalanine, tryptophane or tyrosine. The purity of most of the compounds was estimated to be greater than 99.99%. The modifications resulted in a wide range of conductance changes in NaCl solutions: sixfold from tryptophane gramicidin A to tyrosine gramicidin B. The conductance changes induced by a given amino acid substitution at position 1 are not the same as at position 11. The only important change in the Na+ affinity was observed when the first amino acid was tyrosine. No major conformational changes of the polypeptide backbone structure could be detected on the basis of experiments with mixtures of different analogues and valine gramicidin A (except possibly with tyrosine at position 1), as all the compounds investigated could form hybrid channels with valine gramicidin A. The side chains are not in direct contact with the permeating ions. The results were therefore interpreted in terms of modifications of the energy profile for ion movement through the channel, possibly due to an electrostatic interaction between the dipoles of the side chains and ions in the channel. Images FIGURE 1 FIGURE 2 FIGURE 3 PMID:6201199

  20. Effects of simulated acid rain on growth rate in a spruce-living spider.

    PubMed

    Gunnarsson, B; Johnsson, J

    1989-01-01

    Growing juveniles of the spruce-living spider Pityohyphantes phrygianus were sprayed with water of different acidity--pH approximately 7 (control), 4.0 (acid rain) and 2.2-in a 2.5-month laboratory experiment. The growth rates did not differ between control and pH 4.0, while there was a significant growth reduction at pH 2.2. A low mortality occurred only in the pH 2.2 group. P. phrygianus seems to be resistant against acid rain although negative long-term effects cannot be ruled out.

  1. LASER BIOLOGY AND MEDICINE: Application of laser fluorimetry for determining the influence of a single amino-acid substitution on the individual photophysical parameters of a fluorescent form of a fluorescent protein mRFP1

    NASA Astrophysics Data System (ADS)

    Banishev, A. A.; Vrzheshch, E. P.; Shirshin, E. A.

    2009-03-01

    Individual photophysical parameters of the chromophore of a fluorescent protein mRFP1 and its two mutants (amino-acid substitution at position 66 - mRFP1/ Q66C and mRFP1/Q66S proteins) are determined. For this purpose, apart from conventional methods of fluorimetry and spectrophotometry, nonlinear laser fluorimetry is used. It is shown that the individual extinction coefficients of the chromophore of proteins correlate (correlation coefficient above 0.9) with the volume of the substituted amino-acid residue at position 66 (similar to the positions of the absorption, fluorescence excitation and emission maxima).

  2. Application of laser fluorimetry for determining the influence of a single amino-acid substitution on the individual photophysical parameters of a fluorescent form of a fluorescent protein mRFP1

    SciTech Connect

    Banishev, A A; Vrzheshch, E P; Shirshin, E A

    2009-03-31

    Individual photophysical parameters of the chromophore of a fluorescent protein mRFP1 and its two mutants (amino-acid substitution at position 66 - mRFP1/ Q66C and mRFP1/Q66S proteins) are determined. For this purpose, apart from conventional methods of fluorimetry and spectrophotometry, nonlinear laser fluorimetry is used. It is shown that the individual extinction coefficients of the chromophore of proteins correlate (correlation coefficient above 0.9) with the volume of the substituted amino-acid residue at position 66 (similar to the positions of the absorption, fluorescence excitation and emission maxima). (laser biology and medicine)

  3. A single amino acid substitution in a chitinase of the legume Medicago truncatula is sufficient to gain Nod-factor hydrolase activity.

    PubMed

    Zhang, Lan-Yue; Cai, Jie; Li, Ru-Jie; Liu, Wei; Wagner, Christian; Wong, Kam-Bo; Xie, Zhi-Ping; Staehelin, Christian

    2016-07-01

    The symbiotic interaction between nitrogen-fixing rhizobia and legumes depends on lipo-chitooligosaccharidic Nod-factors (NFs). The NF hydrolase MtNFH1 of Medicago truncatula is a symbiotic enzyme that hydrolytically inactivates NFs with a C16 : 2 acyl chain produced by the microsymbiont Sinorhizobium meliloti 1021. MtNFH1 is related to class V chitinases (glycoside hydrolase family 18) but lacks chitinase activity. Here, we investigated the substrate specificity of MtNFH1-related proteins. MtCHIT5a and MtCHIT5b of M. truncatula as well as LjCHIT5 of Lotus japonicus showed chitinase activity, suggesting a role in plant defence. The enzymes failed to hydrolyse NFs from S. meliloti. NFs from Rhizobium leguminosarum with a C18 : 4 acyl moiety were neither hydrolysed by these chitinases nor by MtNFH1. Construction of chimeric proteins and further amino acid replacements in MtCHIT5b were performed to identify chitinase variants that gained the ability to hydrolyse NFs. A single serine-to-proline substitution was sufficient to convert MtCHIT5b into an NF-cleaving enzyme. MtNFH1 with the corresponding proline-to-serine substitution failed to hydrolyse NFs. These results are in agreement with a substrate-enzyme model that predicts NF cleavage when the C16 : 2 moiety is placed into a distinct fatty acid-binding cleft. Our findings support the view that MtNFH1 evolved from the ancestral MtCHIT5b by gene duplication and subsequent symbiosis-related neofunctionalization. PMID:27383628

  4. A single amino acid substitution in a chitinase of the legume Medicago truncatula is sufficient to gain Nod-factor hydrolase activity

    PubMed Central

    Zhang, Lan-Yue; Cai, Jie; Li, Ru-Jie; Liu, Wei; Wagner, Christian; Wong, Kam-Bo; Xie, Zhi-Ping; Staehelin, Christian

    2016-01-01

    The symbiotic interaction between nitrogen-fixing rhizobia and legumes depends on lipo-chitooligosaccharidic Nod-factors (NFs). The NF hydrolase MtNFH1 of Medicago truncatula is a symbiotic enzyme that hydrolytically inactivates NFs with a C16 : 2 acyl chain produced by the microsymbiont Sinorhizobium meliloti 1021. MtNFH1 is related to class V chitinases (glycoside hydrolase family 18) but lacks chitinase activity. Here, we investigated the substrate specificity of MtNFH1-related proteins. MtCHIT5a and MtCHIT5b of M. truncatula as well as LjCHIT5 of Lotus japonicus showed chitinase activity, suggesting a role in plant defence. The enzymes failed to hydrolyse NFs from S. meliloti. NFs from Rhizobium leguminosarum with a C18 : 4 acyl moiety were neither hydrolysed by these chitinases nor by MtNFH1. Construction of chimeric proteins and further amino acid replacements in MtCHIT5b were performed to identify chitinase variants that gained the ability to hydrolyse NFs. A single serine-to-proline substitution was sufficient to convert MtCHIT5b into an NF-cleaving enzyme. MtNFH1 with the corresponding proline-to-serine substitution failed to hydrolyse NFs. These results are in agreement with a substrate-enzyme model that predicts NF cleavage when the C16 : 2 moiety is placed into a distinct fatty acid-binding cleft. Our findings support the view that MtNFH1 evolved from the ancestral MtCHIT5b by gene duplication and subsequent symbiosis-related neofunctionalization. PMID:27383628

  5. A single amino acid substitution (R441A) in the receptor-binding domain of SARS coronavirus spike protein disrupts the antigenic structure and binding activity

    SciTech Connect

    He Yuxian . E-mail: yhe@nybloodcenter.org; Li Jingjing; Jiang Shibo

    2006-05-26

    The spike (S) protein of severe acute respiratory syndrome coronavirus (SARS-CoV) has two major functions: interacting with the receptor to mediate virus entry and inducing protective immunity. Coincidently, the receptor-binding domain (RBD, residues 318-510) of SAR-CoV S protein is a major antigenic site to induce neutralizing antibodies. Here, we used RBD-Fc, a fusion protein containing the RBD and human IgG1 Fc, as a model in the studies and found that a single amino acid substitution in the RBD (R441A) could abolish the immunogenicity of RBD to induce neutralizing antibodies in immunized mice and rabbits. With a panel of anti-RBD mAbs as probes, we observed that R441A substitution was able to disrupt the majority of neutralizing epitopes in the RBD, suggesting that this residue is critical for the antigenic structure responsible for inducing protective immune responses. We also demonstrated that the RBD-Fc bearing R441A mutation could not bind to soluble and cell-associated angiotensin-converting enzyme 2 (ACE2), the functional receptor for SARS-CoV and failed to block S protein-mediated pseudovirus entry, indicating that this point mutation also disrupted the receptor-binding motif (RBM) in the RBD. Taken together, these data provide direct evidence to show that a single amino acid residue at key position in the RBD can determine the major function of SARS-CoV S protein and imply for designing SARS vaccines and therapeutics.

  6. 40 CFR 721.10202 - Benzoic acid, 4-chloro-2- [(substituted)azo]-, strontium salt (1:1) (generic).

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... salt (1:1) (generic). 721.10202 Section 721.10202 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) TOXIC SUBSTANCES CONTROL ACT SIGNIFICANT NEW USES OF CHEMICAL SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.10202 Benzoic acid, 4-chloro-2- -, strontium salt...

  7. 40 CFR 721.10202 - Benzoic acid, 4-chloro-2- [(substituted)azo]-, strontium salt (1:1) (generic).

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... salt (1:1) (generic). 721.10202 Section 721.10202 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) TOXIC SUBSTANCES CONTROL ACT SIGNIFICANT NEW USES OF CHEMICAL SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.10202 Benzoic acid, 4-chloro-2- -, strontium salt...

  8. 40 CFR 721.10202 - Benzoic acid, 4-chloro-2- [(substituted)azo]-, strontium salt (1:1) (generic).

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... salt (1:1) (generic). 721.10202 Section 721.10202 Protection of Environment ENVIRONMENTAL PROTECTION AGENCY (CONTINUED) TOXIC SUBSTANCES CONTROL ACT SIGNIFICANT NEW USES OF CHEMICAL SUBSTANCES Significant New Uses for Specific Chemical Substances § 721.10202 Benzoic acid, 4-chloro-2- -, strontium salt...

  9. Occurrence and metabolism of 4-substituted glutamic acids in the seedlings of various species of legumes. [Sophora japonica

    SciTech Connect

    Winter, H.C.; Dekker, E.E.

    1987-04-01

    The authors measured the levels of 4-methyleneglutamic acid (Meglu), 4-methyleneglutamine (Megln), erythro-4-methylglutamic acid (e-Mglu), and threo-4-methylglutamic acid (t-Mglu) in seedlings of various species of legumes by HPLC and ion exchange chromatography. High levels of e-Mglu and Megln but no t-Mglu or Meglu are present in Sophora japonica. Peanut seedling contain both e-Mglu and t-Mglu at 20-50% and 5%, resp., of the level of Meglu whereas only traces of Meglu and Mglu occur in soybean seedlings. Excised peanut embryos germinated on Linsmaier and Skoog medium + (U-/sup 14/C)-leucine incorporated isotope into e-Mglu, Meglu, and Megln; (U-/sup 14/C)-proline or glycine was not so incorporated. Soybean embryos rapidly converted added (2-/sup 14/C)-Meglu to a variety of non-amino acid products; peanut embryos, in contrast, retain 25% of added Meglu unchanged and 50% as Megln. These results suggest that in a variety of legumes leucine may serve as a precursor of Mglu and Meglu during germination; also, whereas Meglu remains as such or as Megln in some species, it is rapidly metabolized in others.

  10. Identification and Structural Analysis of Amino Acid Substitutions that Increase the Stability and Activity of Aspergillus niger Glucose Oxidase

    PubMed Central

    Marín-Navarro, Julia; Roupain, Nicole; Talens-Perales, David; Polaina, Julio

    2015-01-01

    Glucose oxidase is one of the most conspicuous commercial enzymes due to its many different applications in diverse industries such as food, chemical, energy and textile. Among these applications, the most remarkable is the manufacture of glucose biosensors and in particular sensor strips used to measure glucose levels in serum. The generation of ameliorated versions of glucose oxidase is therefore a significant biotechnological objective. We have used a strategy that combined random and rational approaches to isolate uncharacterized mutations of Aspergillus niger glucose oxidase with improved properties. As a result, we have identified two changes that increase significantly the enzyme's thermal stability. One (T554M) generates a sulfur-pi interaction and the other (Q90R/Y509E) introduces a new salt bridge near the interphase of the dimeric protein structure. An additional double substitution (Q124R/L569E) has no significant effect on stability but causes a twofold increase of the enzyme's specific activity. Our results disclose structural motifs of the protein which are critical for its stability. The combination of mutations in the Q90R/Y509E/T554M triple mutant yielded a version of A. niger glucose oxidase with higher stability than those previously described. PMID:26642312

  11. Does L to D-amino acid substitution trigger helix→sheet conformations in collagen like peptides adsorbed to surfaces?

    PubMed

    Velmurugan, Punitha; Jonnalagadda, Raghava Rao; Sankaranarayanan, Kamatchi; Dhathathreyan, Aruna

    2015-12-01

    The present work reports on the structural order, self assembling behaviour and the role in adsorption to hydrophilic or hydrophobic solid surfaces of modified sequence from the triple helical peptide model of the collagenase cleavage site in type I collagen (Uniprot accession number P02452 residues from 935 to 970) using (D)Ala and (D)Ile substitutions as given in the models below: Model-1: GSOGADGPAGAOGTOGPQGIAGQRGVV GLOGQRGER. Model-2: GSOGADGP(D)AGAOGTOGPQGIAGQRGVVGLOGQRGER. Model-3: GSOGADGPAGAOGTOGPQG(D)IAGQRGVVGLOGQRGER. Collagenase is an important enzyme that plays an important role in degrading collagen in wound healing, cancer metastasis and even in embryonic development. However, the mechanism by which this degradation occurs is not completely understood. Our results show that adsorption of the peptides to the solid surfaces, specifically hydrophobic triggers a helix to beta transition with order increasing in peptide models 2 and 3. This restricts the collagenolytic behaviour of collagenase and may find application in design of peptides and peptidomimetics for enzyme-substrate interaction, specifically with reference to collagen and other extra cellular matrix proteins.

  12. Effect of the amino acid substitution in the DNA-binding domain of the Fur regulator on production of pyoverdine.

    PubMed

    Valešová, Renáta; Palyzová, Andrea; Marešová, Helena; Stěpánek, Václav; Babiak, Peter; Kyslík, Pavel

    2013-07-01

    The ferric uptake regulator gene (fur), its promoter region and Fur box of pvdS gene involved in siderophore-mediated iron uptake system were sequenced in the parent strain Pseudomonas aeruginosa PAO1 and in the fur mutant FPA121 derived from the strain PAO1. We identified the gene fur 179 bearing a novel, single-point mutation that changed the amino acid residue Gln60Pro in the DNA-binding domain of the Fur protein. The synthesis of pyoverdine was studied in cultures of the strains PAO1 and FPA121 grown in iron-deplete and iron-replete (60 μmol/L FeIII) medium. The amino acid replacement in the regulatory Fur protein is responsible for the overproduction of pyoverdine in iron-deplete and iron-replete medium. No mutation was identified in the Fur box of the gene pvdS.

  13. Reduction of birth prevalence rates of neural tube defects after folic acid fortification in Chile.

    PubMed

    López-Camelo, Jorge S; Orioli, Iêda M; da Graça Dutra, Maria; Nazer-Herrera, Julio; Rivera, Nelson; Ojeda, María Elena; Canessa, Aurora; Wettig, Elisabeth; Fontannaz, Ana María; Mellado, Cecília; Castilla, Eduardo E

    2005-06-01

    To verify whether the decreasing neural tube defects birth prevalence rates in Chile are due to folic acid fortification or to pre-existing decreasing trends, we performed a population survey using a network of Estudio Colaborativo Latino Americano de Malformaciones Congenitas (ECLAMC, Latin American Collaborative Study of Congenital Malformations) maternity hospitals in Chile, between the years 1982 and 2002. Within each maternity hospital, birth prevalence rates of spina bifida and anencephaly were calculated from two pre-fortification periods (1982-1989 and 1990-2000), and from one fortified period (2001-2002). There was no historical trend for spina bifida birth prevalence rates before folic acid fortification, and there was a 51% (minimum 27%, maximum 66%) decrease in the birth prevalence rates of this anomaly in the fortified period. The relative risks of spina bifida were homogeneous among hospitals in the two period comparisons. There was no historical trend for the birth prevalence of anencephaly comparing the two pre-fortified periods, but the relative risks were heterogeneous among hospitals in this comparison. There was a 42% (minimum 10%, maximum 63%) decrease in the birth prevalence rate of anencephaly in the fortified period as compared with the immediately pre-fortified period, with homogeneous relative risks among hospitals. Within the methodological constraints of this study we conclude that the birth prevalence rates for both spina bifida and anencephaly decreased as a result of folic acid fortification, without interference of decreasing secular trends.

  14. Functional genomics reveals increases in cholesterol biosynthetic genes and highly unsaturated fatty acid biosynthesis after dietary substitution of fish oil with vegetable oils in Atlantic salmon (Salmo salar)

    PubMed Central

    Leaver, Michael J; Villeneuve, Laure AN; Obach, Alex; Jensen, Linda; Bron, James E; Tocher, Douglas R; Taggart, John B

    2008-01-01

    Background There is an increasing drive to replace fish oil (FO) in finfish aquaculture diets with vegetable oils (VO), driven by the short supply of FO derived from wild fish stocks. However, little is known of the consequences for fish health after such substitution. The effect of dietary VO on hepatic gene expression, lipid composition and growth was determined in Atlantic salmon (Salmo salar), using a combination of cDNA microarray, lipid, and biochemical analysis. FO was replaced with VO, added to diets as rapeseed (RO), soybean (SO) or linseed (LO) oils. Results Dietary VO had no major effect on growth of the fish, but increased the whole fish protein contents and tended to decrease whole fish lipid content, thus increasing the protein:lipid ratio. Expression levels of genes of the highly unsaturated fatty acid (HUFA) and cholesterol biosynthetic pathways were increased in all vegetable oil diets as was SREBP2, a master transcriptional regulator of these pathways. Other genes whose expression was increased by feeding VO included those of NADPH generation, lipid transport, peroxisomal fatty acid oxidation, a marker of intracellular lipid accumulation, and protein and RNA processing. Consistent with these results, HUFA biosynthesis, hepatic β-oxidation activity and enzymic NADPH production were changed by VO, and there was a trend for increased hepatic lipid in LO and SO diets. Tissue cholesterol levels in VO fed fish were the same as animals fed FO, whereas fatty acid composition of the tissues largely reflected those of the diets and was marked by enrichment of 18 carbon fatty acids and reductions in 20 and 22 carbon HUFA. Conclusion This combined gene expression, compositional and metabolic study demonstrates that major lipid metabolic effects occur after replacing FO with VO in salmon diets. These effects are most likely mediated by SREBP2, which responds to reductions in dietary cholesterol. These changes are sufficient to maintain whole body cholesterol

  15. Single substitutions to closely related amino acids contribute to the functional diversification of an insect-inducible, positively selected plant cystatin.

    PubMed

    Rasoolizadeh, Asieh; Goulet, Marie-Claire; Sainsbury, Frank; Cloutier, Conrad; Michaud, Dominique

    2016-04-01

    A causal link has been reported between positively selected amino acids in plant cystatins and the inhibitory range of these proteins against insect digestive cysteine (Cys) proteases. Here we assessed the impact of single substitutions to closely related amino acids on the contribution of positive selection to cystatin diversification. Cystatin sequence alignments, while confirming hypervariability, indicated a preference for related amino acids at positively selected sites. For example, the non-polar residues leucine (Leu), isoleucine (Ile) and valine (Val) were shown to predominate at positively selected site 2 in the N-terminal region, unlike selected sites 6 and 10, where polar residues are preferred. The model cystatin SlCYS8 and single variants with Leu, Ile or Val at position 2 were compared with regard to their ability to bind digestive proteases of the coleopteran pest Leptinotarsa decemlineata and to induce compensatory responses in this insect. A functional proteomics procedure to capture target Cys proteases in midgut extracts allowed confirmation of distinct binding profiles for the cystatin variants. A shotgun proteomics procedure to monitor whole Cys protease complements revealed protease family specific compensatory responses in the insect, dependent on the variant ingested. Our data confirm the contribution of closely related amino acids to the functional diversity of positively selected plant cystatins in a broader structure/function context imposing physicochemical constraints to primary structure alterations. They also underline the complexity of protease/inhibitor interactions in plant-insect systems, and the challenges still to be met in order to harness the full potential of ectopically expressed protease inhibitors in crop protection.

  16. Towards Sustainable Aquafeeds: Complete Substitution of Fish Oil with Marine Microalga Schizochytrium sp. Improves Growth and Fatty Acid Deposition in Juvenile Nile Tilapia (Oreochromis niloticus)

    PubMed Central

    Sarker, Pallab K.; Kapuscinski, Anne R.; Lanois, Alison J.; Livesey, Erin D.; Bernhard, Katie P.; Coley, Mariah L.

    2016-01-01

    We conducted a 84-day nutritional feeding experiment with dried whole cells of DHA-rich marine microalga Schizochytrium sp. (Sc) to determine the optimum level of fish-oil substitution (partial or complete) for maximum growth of Nile tilapia. When we fully replaced fish oil with Schizochytrium (Sc100 diet), we found significantly higher weight gain and protein efficiency ratio (PER), and lower (improved) feed conversion ratio (FCR) and feed intake compared to a control diet containing fish oil (Sc0); and no significant change in SGR and survival rate among all diets. The Sc100 diet had the highest contents of 22:6n3 DHA, led to the highest DHA content in fillets, and consequently led to the highest DHA:EPA ratios in tilapia fillets. Schizochytrium sp. is a high quality candidate for complete substitution of fish oil in juvenile Nile tilapia feeds, providing an innovative means to formulate and optimize the composition of tilapia juvenile feed while simultaneously raising feed efficiency of tilapia aquaculture and to further develop environmentally and socially sustainable aquafeeds. Results show that replacing fish oil with DHA-rich marine Sc improves the deposition of n3 LC PUFA levels in tilapia fillet. These results support further studies to lower Schizochytrium production costs and to combine different marine microalgae to replace fish oil and fishmeal into aquafeeds. PMID:27258552

  17. Towards Sustainable Aquafeeds: Complete Substitution of Fish Oil with Marine Microalga Schizochytrium sp. Improves Growth and Fatty Acid Deposition in Juvenile Nile Tilapia (Oreochromis niloticus).

    PubMed

    Sarker, Pallab K; Kapuscinski, Anne R; Lanois, Alison J; Livesey, Erin D; Bernhard, Katie P; Coley, Mariah L

    2016-01-01

    We conducted a 84-day nutritional feeding experiment with dried whole cells of DHA-rich marine microalga Schizochytrium sp. (Sc) to determine the optimum level of fish-oil substitution (partial or complete) for maximum growth of Nile tilapia. When we fully replaced fish oil with Schizochytrium (Sc100 diet), we found significantly higher weight gain and protein efficiency ratio (PER), and lower (improved) feed conversion ratio (FCR) and feed intake compared to a control diet containing fish oil (Sc0); and no significant change in SGR and survival rate among all diets. The Sc100 diet had the highest contents of 22:6n3 DHA, led to the highest DHA content in fillets, and consequently led to the highest DHA:EPA ratios in tilapia fillets. Schizochytrium sp. is a high quality candidate for complete substitution of fish oil in juvenile Nile tilapia feeds, providing an innovative means to formulate and optimize the composition of tilapia juvenile feed while simultaneously raising feed efficiency of tilapia aquaculture and to further develop environmentally and socially sustainable aquafeeds. Results show that replacing fish oil with DHA-rich marine Sc improves the deposition of n3 LC PUFA levels in tilapia fillet. These results support further studies to lower Schizochytrium production costs and to combine different marine microalgae to replace fish oil and fishmeal into aquafeeds.

  18. Towards Sustainable Aquafeeds: Complete Substitution of Fish Oil with Marine Microalga Schizochytrium sp. Improves Growth and Fatty Acid Deposition in Juvenile Nile Tilapia (Oreochromis niloticus).

    PubMed

    Sarker, Pallab K; Kapuscinski, Anne R; Lanois, Alison J; Livesey, Erin D; Bernhard, Katie P; Coley, Mariah L

    2016-01-01

    We conducted a 84-day nutritional feeding experiment with dried whole cells of DHA-rich marine microalga Schizochytrium sp. (Sc) to determine the optimum level of fish-oil substitution (partial or complete) for maximum growth of Nile tilapia. When we fully replaced fish oil with Schizochytrium (Sc100 diet), we found significantly higher weight gain and protein efficiency ratio (PER), and lower (improved) feed conversion ratio (FCR) and feed intake compared to a control diet containing fish oil (Sc0); and no significant change in SGR and survival rate among all diets. The Sc100 diet had the highest contents of 22:6n3 DHA, led to the highest DHA content in fillets, and consequently led to the highest DHA:EPA ratios in tilapia fillets. Schizochytrium sp. is a high quality candidate for complete substitution of fish oil in juvenile Nile tilapia feeds, providing an innovative means to formulate and optimize the composition of tilapia juvenile feed while simultaneously raising feed efficiency of tilapia aquaculture and to further develop environmentally and socially sustainable aquafeeds. Results show that replacing fish oil with DHA-rich marine Sc improves the deposition of n3 LC PUFA levels in tilapia fillet. These results support further studies to lower Schizochytrium production costs and to combine different marine microalgae to replace fish oil and fishmeal into aquafeeds. PMID:27258552

  19. Effect of diffusion layer pH and solubility on the dissolution rate of pharmaceutical acids and their sodium salts. II: Salicylic acid, theophylline, and benzoic acid.

    PubMed

    Serajuddin, A T; Jarowski, C I

    1985-02-01

    The pH-solubility profiles of salicylic acid and theophylline, as determined by the addition of HCl or NaOH to their aqueous suspensions, were identical with those of their sodium salts except during phase transitions from acid to salt or vice versa. Supersaturated solutions were formed during phase transitions. Unlike the solubility profiles, the pH-intrinsic dissolution rate profiles of an acid and its salt differed greatly. Good conformity with the Noyes-Whitney equation was demonstrated when the solubility values under pH conditions as the diffusion layer thickness, h, approaches zero (Cs,h = 0) were used rather than solubilities under pH conditions of the bulk media (Cs). The pH when h approaches zero (pHh = 0) was estimated by equilibration of a dissolution medium with an excess of material. Good correlation was shown between the pHh = 0 values of benzoic acid estimated according to this method and the pHh = 0 values reported in the literature. The intrinsic dissolution rate constant, the ratio of the diffusion coefficient to the diffusion layer thickness (D/h), may be assumed constant when comparing the dissolution rates of salicylic acid, theophylline and sodium theophylline. On the other hand, D/h decreased significantly during dissolution of sodium salicylate due to a large increase in Cs,h = 0 and the consequent increase in viscosity in the diffusion layer. A simple method of predicting the dissolution rate of an acid or a salt at different pH values has been developed.

  20. Effect of α-Methyl versus α-Hydrogen Substitution on Brain Availability and Tumor Imaging Properties of Heptanoic [F-18]Fluoroalkyl Amino Acids for Positron Emission Tomography (PET).

    PubMed

    Bouhlel, Ahlem; Alyami, Wadha; Li, Aixiao; Yuan, Liya; Rich, Keith; McConathy, Jonathan

    2016-04-14

    Two [(18)F]fluoroalkyl substituted amino acids differing only by the presence or absence of a methyl group on the α-carbon, (S)-2-amino-7-[(18)F]fluoro-2-methylheptanoic acid ((S)-[(18)F]FAMHep, (S)-[(18)F]14) and (S)-2-amino-7-[(18)F]fluoroheptanoic acid ((S)-[(18)F]FAHep, (S)-[(18)F]15), were developed for brain tumor imaging and compared to the well-established system L amino acid tracer, O-(2-[(18)F]fluoroethyl)-l-tyrosine ([(18)F]FET), in the delayed brain tumor (DBT) mouse model of high-grade glioma. Cell uptake, biodistribution, and PET/CT imaging studies showed differences in amino acid transport of these tracer by DBT cells. Recognition of (S)-[(18)F]15 but not (S)-[(18)F]14 by system L amino acid transporters led to approximately 8-10-fold higher uptake of the α-hydrogen substituted analogue (S)-[(18)F]15 in normal brain. (S)-[(18)F]15 had imaging properties similar to those of (S)-[(18)F]FET in the DBT tumor model while (S)-[(18)F]14 afforded higher tumor to brain ratios due to much lower uptake by normal brain. These results have important implications for the future development of α-alkyl and α,α-dialkyl substituted amino acids for brain tumor imaging.

  1. How the folding rates of two- and multistate proteins depend on the amino acid properties.

    PubMed

    Huang, Jitao T; Huang, Wei; Huang, Shanran R; Li, Xin

    2014-10-01

    Proteins fold by either two-state or multistate kinetic mechanism. We observe that amino acids play different roles in different mechanism. Many residues that are easy to form regular secondary structures (α helices, β sheets and turns) can promote the two-state folding reactions of small proteins. Most of hydrophilic residues can speed up the multistate folding reactions of large proteins. Folding rates of large proteins are equally responsive to the flexibility of partial amino acids. Other properties of amino acids (including volume, polarity, accessible surface, exposure degree, isoelectric point, and phase transfer energy) have contributed little to folding kinetics of the proteins. Cysteine is a special residue, it triggers two-state folding reaction and but inhibits multistate folding reaction. These findings not only provide a new insight into protein structure prediction, but also could be used to direct the point mutations that can change folding rate.

  2. Factors influencing the rate of non-enzymatic activation of carboxylic and amino acids by ATP

    NASA Technical Reports Server (NTRS)

    Mullins, D. W., Jr.; Lacey, J. C., Jr.

    1981-01-01

    The nonenzymatic formation of adenylate anhydrides of carboxylic and amino acids is discussed as a necessary step in the origin of the genetic code and protein biosynthesis. Results of studies are presented which have shown the rate of activation to depend on the pKa of the carboxyl group, the pH of the medium, temperature, the divalent metal ion catalyst, salt concentration, and the nature of the amino acid. In particular, it was found that of the various amino acids investigated, phenylalanine had the greatest affinity for the adenine derivatives adenosine and ATP. Results thus indicate that selective affinities between amino acids and nucleotides were important during prebiotic chemical evolution, and may have played a major role in the origin of protein synthesis and genetic coding.

  3. Non-peptide angiotensin II receptor antagonists. 2. Design, synthesis, and biological activity of N-substituted (phenylamino)phenylacetic acids and acyl sulfonamides.

    PubMed

    Dhanoa, D S; Bagley, S W; Chang, R S; Lotti, V J; Chen, T B; Kivlighn, S D; Zingaro, G J; Siegl, P K; Patchett, A A; Greenlee, W J

    1993-12-24

    The design, synthesis, and biological activity of a new class of highly potent non-peptide AII receptor antagonists derived from N-substituted (phenylamino)phenylacetic acids and acyl sulfonamides which exhibit a high selectivity for the AT1 receptor are described. A series of N-substituted (phenylamino)phenylacetic acids (9) and acyl sulfonamides (16) and a tetrazole derivative (19) were synthesized and evaluated in the in vitro AT1 (rabbit aorta) and AT2 (rat midbrain) binding assay. The (phenylamino)phenylacetic acids 9c (AT1 IC50 = 4 nM, AT2 IC50 = 0.74 microM), 9d (AT1 IC50 = 5.3 nM, AT2 IC50 = 0.49 microM), and 9e (AT1 IC50 = 5.3 nM, AT2 IC50 = 0.56 microM) were found to be the most potent AT1-selective AII antagonists in the acid series. Incorporation of various substituents in the central and bottom phenyl rings led to a decrease in the AT1 and AT2 binding affinity of the resulting compounds. Replacement of the carboxylic acid (CO2H) in 9c, 9d, and 9e with the bioisostere acyl sulfonamide (CONHSO2Ph) resulted in a (5-7)-fold increase in the AT1 potency of 16a (AT1 IC50 = 0.9 nM, AT2 IC50 = 0.2 microM), 16b (AT1 IC50 = 1 nM, AT2 IC50 = 2.9 microM), and 16c (AT1 IC50 = 0.8 nM, AT2 IC50 = 0.42 microM) and yielded acyl sulfonamides with subnanomolar AT1 activity. Incorporation of the acyl sulfonamide (CONHSO2Ph) for the CO2H of 9c not only enhanced the AT1 potency but also effected a marked increase in the AT2 potency of 16a (AT2 IC50 = 0.74 microM of 9c vs 0.2 microM of 16a) and made it the most potent AT2 antagonist in this study. Replacement of the CO2H of 9b with the bioisostere tetrazole resulted in 19 (AT1 IC50 = 15 nM) with a 2-fold loss in the AT1 and a complete loss in the AT2 binding affinity. (Phenylamino)phenylacetic acid 9c demonstrated good oral activity in AII-infused conscious normotensive rats at an oral dose of 1.0 mg/kg by inhibiting the pressor response for > 6 h. Acyl sulfonamides 16a-c displayed excellent in vivo activity by blocking the

  4. Fundamental Study on Temperature Dependence of Deposition Rate of Silicic Acid - 13270

    SciTech Connect

    Shinmura, Hayata; Niibori, Yuichi; Mimura, Hitoshi

    2013-07-01

    The dynamic behavior of the silicic acid is one of the key factors to estimate the condition of the repository system after the backfill. This study experimentally examined the temperature dependence of dynamic behavior of supersaturated silicic acid in the co-presence of solid phase, considering Na ions around the repository, and evaluated the deposition rate constant, k, of silicic acid by using the first-order reaction equation considering the specific surface area. The values of k were in the range of 1.0x10{sup -11} to 1.0x10{sup -9} m/s in the temperature range of 288 K to 323 K. The deposition rate became larger with increments of temperature under the Na ion free condition. Besides, in the case of Na ions 0.6 M, colloidal silicic acid decreased dramatically at a certain time. This means that the diameter of the colloidal silicic acid became larger than the pore size of filter (0.45 μm) due to bridging of colloidal silicic acid. Furthermore, this study estimated the range of altering area and the aperture of flow-path in various value of k corresponding to temperature by using advection-dispersion model. The concentration in the flow-path became lower with increments of temperature, and when the value of k is larger than 1.0x10{sup -11} m/s, the deposition range of supersaturated silicic acid was estimated to be less than 20 m around the repository. In addition, the deposition of supersaturated silicic acid led the decrement of flow-path aperture, which was remarkable under the condition of relatively high temperature. Such a clogging in flow paths is expected as a retardation effect of radionuclides. (authors)

  5. Novel high-affinity and selective biaromatic 4-substituted gamma-hydroxybutyric acid (GHB) analogues as GHB ligands: design, synthesis, and binding studies.

    PubMed

    Høg, Signe; Wellendorph, Petrine; Nielsen, Birgitte; Frydenvang, Karla; Dahl, Ivar F; Bräuner-Osborne, Hans; Brehm, Lotte; Frølund, Bente; Clausen, Rasmus P

    2008-12-25

    Gamma-hydroxybutyrate (GHB) is a metabolite of gamma-aminobutyric acid (GABA) and has been proposed to function as a neurotransmitter or neuromodulator. GHB is used in the treatment of narcolepsy and is a drug of abuse. GHB binds to both GABA(B) receptors and specific high-affinity GHB sites in brain, of which the latter have not been linked unequivocally to function, but are speculated to be GHB receptors. In this study, a series of biaromatic 4-substituted GHB analogues, including 4'-phenethylphenyl, 4'-styrylphenyl, and 4'-benzyloxyphenyl GHB analogues, were synthesized and characterized pharmacologically in a [3H](E,RS)-(6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene)acetic acid ([3H]NCS-382) binding assay and in GABA(A) and GABA(B) receptor binding assays. The compounds were selective for the high-affinity GHB binding sites and several displayed Ki values below 100 nM. The affinity of the 4-[4'-(2-iodobenzyloxy)phenyl] GHB analogue 17b was shown to reside predominantly with the R-enantiomer (Ki = 22 nM), which has higher affinity than previously reported GHB ligands.

  6. Adaptive amino acid substitutions enhance the virulence of an H7N7 avian influenza virus isolated from wild waterfowl in mice.

    PubMed

    Chen, Qiang; Yu, Zhijun; Sun, Weiyang; Li, Xue; Chai, Hongliang; Gao, Xiaolong; Guo, Jiao; Zhang, Kun; Feng, Na; Zheng, Xuexing; Wang, Hualei; Zhao, Yongkun; Qin, Chuan; Huang, Geng; Yang, Songtao; Qian, Jun; Gao, Yuwei; Xia, Xianzhu; Wang, Tiecheng; Hua, Yuping

    2015-05-15

    Although H7N7 AIVs primarily circulate in wild waterfowl, documented cases of human infection with H7N7 viruses suggest they may pose a pandemic threat. Here, we generated mouse-adapted variants of a wild waterfowl-origin H7N7 virus to identify adaptive changes that confer enhanced virulence in mammals. The mouse lethal doses (MLD50) of the adapted variants were reduced >5000-fold compared to the parental virus. Mouse-adapted variants viruses displayed enhanced replication in vitro and in vivo, and acquired the ability to replicate in extrapulmonary tissues. These observations suggest that enhanced growth characteristics and modified cell tropism may increase the virulence of H7N7 AIVs in mice. Genomic analysis of the adapted variant viruses revealed amino acid changes in the PB2 (E627K), PB1 (R118I), PA (L550M), HA (G214R), and NA (S372N) proteins. Our results suggest that these amino acid substitutions collaboratively enhance the ability of H7N7 virus to replicate and cause severe disease in mammals. PMID:25769645

  7. Straightforward and effective synthesis of γ-aminobutyric acid transporter subtype 2-selective acyl-substituted azaspiro[4.5]decanes.

    PubMed

    Ma, Xiaofeng; Lubin, Hodney; Ioja, Enikő; Kékesi, Orsolya; Simon, Ágnes; Apáti, Ágota; Orbán, Tamás I; Héja, László; Kardos, Julianna; Markó, István E

    2016-01-15

    Supply of major metabolites such as γ-aminobutyric acid (GABA), β-alanine and taurine is an essential instrument that shapes signalling, proper cell functioning and survival in the brain and peripheral organs. This background motivates the synthesis of novel classes of compounds regulating their selective transport through various fluid-organ barriers via the low-affinity γ-aminobutyric acid (GABA) transporter subtype 2 (GAT2). Natural and synthetic spirocyclic compounds or therapeutics with a range of structures and biological activity are increasingly recognised in this regard. Based on pre-validated GABA transport activity, straightforward and efficient synthesis method was developed to provide an azaspiro[4.5]decane scaffold, holding a variety of charge, substituent and 3D constrain of spirocyclic amine. Investigation of the azaspiro[4.5]decane scaffold in cell lines expressing the four GABA transporter subtypes led to the discovery of a subclass of a GAT2-selective compounds with acyl-substituted azaspiro[4.5]decane core.

  8. Enantioselective nitrile anion cyclization to substituted pyrrolidines. A highly efficient synthesis of (3S,4R)-N-tert-butyl-4-arylpyrrolidine-3-carboxylic acid.

    PubMed

    Chung, John Y L; Cvetovich, Raymond; Amato, Joseph; McWilliams, J Christopher; Reamer, Robert; DiMichele, Lisa

    2005-04-29

    [reaction: see text] A practical asymmetric synthesis of N-tert-butyl disubstituted pyrrolidines via a nitrile anion cyclization strategy is described. The five-step chromatography-free synthesis of (3S,4R)-1-tert-butyl-4-(2,4-difluorophenyl)pyrrolidine-3-carboxylic acid (2) from 2-chloro-1-(2,4-difluorophenyl)-ethanone achieved a 71% overall yield. The cyclization substrate was prepared via a catalytic CBS asymmetric reduction, t-butylamine displacement of the chlorohydrin, and a conjugate addition of the hindered secondary amine to acrylonitrile. The key nitrile anion 5-exo-tet cyclization concomitantly formed the pyrrolidine ring with clean inversion of the C-4 center to afford 1,3,4-trisubstituted chiral pyrrolidine in >95% yield and 94-99% ee. Diethyl chlorophosphate and lithium hexamethyldisilazide were shown to be the respective optimum activating group and base in this cyclization. The trans-cis mixture of the pyrrolidine nitrile undergoes a kinetically controlled epimerization/ saponification to afford the pure trans-pyrrolidine carboxylic acid target compound in >99.9% chemical and optical purity. This chemistry was also shown to be applicable to both electronically neutral and rich substituted phenyl substrates.

  9. Peruvian and globally reported amino acid substitutions on the Mycobacterium tuberculosis pyrazinamidase suggest a conserved pattern of mutations associated to pyrazinamide resistance

    PubMed Central

    Zimic, Mirko; Sheen, Patricia; Quiliano, Miguel; Gutierrez, Andrés; Gilman, Robert H.

    2010-01-01

    Resistance to pyrazinamide in Mycobacterium tuberculosis is usually associated with a reduction of pyrazinamidase activity caused by mutations in pncA, the pyrazinamidase coding gene. Pyrazinamidase is a hydrolase that converts pyrazinamide, the antituberculous drug against the latent stage, to the active compound, pyrazinoic acid. To better understand the relationship between pncA mutations and pyrazinamide-resistance, it is necessary to analyze the distribution of pncA mutations from pyrazinamide resistant strains. We determined the distribution of Peruvian and globally reported pncA missense mutations from M. tuberculosis clinical isolates resistant to pyrazinamide. The distributions of the single amino acid substitutions were compared at the secondary-structure-domains level. The distribution of the Peruvian mutations followed a similar pattern as the mutations reported globally. A consensus clustering of mutations was observed in hot-spot regions located in the metal coordination site and to a lesser extent in the active site of the enzyme. The data was not able to reject the null hypothesis that both distributions are similar, suggesting that pncA mutations associated to pyrazinamide resistance in M. tuberculosis, follow a conserved pattern responsible to impair the pyrazinamidase activity. PMID:19963078

  10. IMB-6G, a novel N-substituted sophoridinic acid derivative, induces endoplasmic reticulum stress-mediated apoptosis via activation of IRE1α and PERK signaling

    PubMed Central

    Zhang, Na; Bi, Chongwen; Liu, Lu; Dou, Yueying; Tang, Sheng; Pang, Weiqiang; Deng, Hongbin; Song, Danqing

    2016-01-01

    Sophoridinic acid derivatives have received considerable attentions for their potencies in cancer therapy. IMB-6G is a novel N-substituted sophoridinic acid derivative with potent cytotoxicity against tumor cells. In the present study, we explored the antitumor abilities of IMB-6G in human hepatocellular carcinoma (HCC) cells and investigated the underlying mechanisms. We found that IMB-6G inhibited cell growth and induced mitochondrial-dependent apoptosis in HepG2 and SMMC7721 cells. Analyses of the molecular mechanism of IMB-6G-induced apoptosis indicated IMB-6G induced endoplasmic reticulum (ER) stress activation. Incubation of HCC cells with IMB-6G induced increase in Bip and CHOP levels, which precede induction of apoptosis. Further study showed IMB-6G activated IRE1α and PERK pathways but did not stimulated ATF6 pathway in HCC cells. Moreover, silencing of IRE1α dramatically abrogated IMB-6G-induced pro-apoptotic ASK1-JNK signaling. Importantly, interruption of CHOP rendered HCC cells sensitive to IMB-6G-induced apoptosis via inactivation of Bim, PUMA and Bax. Thus, the IRE1α-ASK1 and PERK-CHOP pathways may be a novel molecular mechanism of IMB-6G-induced apoptosis. Collectively, our study demonstrates that IMB-6G induces ER stress-mediated apoptosis by activating IRE1α and PERK pathways. Our findings provide a rationale for the potential application of IMB-6G in HCC therapy. PMID:27009865

  11. Straightforward and effective synthesis of γ-aminobutyric acid transporter subtype 2-selective acyl-substituted azaspiro[4.5]decanes.

    PubMed

    Ma, Xiaofeng; Lubin, Hodney; Ioja, Enikő; Kékesi, Orsolya; Simon, Ágnes; Apáti, Ágota; Orbán, Tamás I; Héja, László; Kardos, Julianna; Markó, István E

    2016-01-15

    Supply of major metabolites such as γ-aminobutyric acid (GABA), β-alanine and taurine is an essential instrument that shapes signalling, proper cell functioning and survival in the brain and peripheral organs. This background motivates the synthesis of novel classes of compounds regulating their selective transport through various fluid-organ barriers via the low-affinity γ-aminobutyric acid (GABA) transporter subtype 2 (GAT2). Natural and synthetic spirocyclic compounds or therapeutics with a range of structures and biological activity are increasingly recognised in this regard. Based on pre-validated GABA transport activity, straightforward and efficient synthesis method was developed to provide an azaspiro[4.5]decane scaffold, holding a variety of charge, substituent and 3D constrain of spirocyclic amine. Investigation of the azaspiro[4.5]decane scaffold in cell lines expressing the four GABA transporter subtypes led to the discovery of a subclass of a GAT2-selective compounds with acyl-substituted azaspiro[4.5]decane core. PMID:26706177

  12. The Effect of Ring Substitution Position on the Structural Conformation of Mercaptobenzoic Acid Self-Assembled Monolayers on Au(111)

    SciTech Connect

    Lee, J; Willey, T; Nilsson, J; Terminello, L; De Yoreo, J; van Buuren, T

    2006-04-12

    Near edge X-ray absorption fine structure (NEX-AFS) spectroscopy, photoemission spectroscopy (PES) and contact angle measurements have been used to examine the structure and bonding of self-assembled monolayers (SAMs) prepared on Au(111) from the positional isomers of mercaptobenzoic acid (MBA). The isomer of MBA and solvent chosen in SAM preparation has considerable bearing upon film morphology. Carbon K-edge NEXAFS measurements indicate that the monomers of 2-, 3- and 4-MBA have well-defined orientations within their respective SAMs. Monomers of 3- and 4-MBA assume an upright orientation on the Au substrates in monolayers prepared using an acetic acid in ethanol solvent. The aryl ring and carboxyl group of these molecules are tilted from the surface normal by a colatitudal angle of {approx} 30{sup o}. Preparation of 4-MBA SAMs using pure ethanol solvent, a more traditional means of synthesis, had no appreciable effect upon the monomer orientation. Nonetheless, S(2p) PES measurements illustrate that it results in extensive bilayer formation via carboxyl group hydrogen-bonding between 4-MBA monomers. In 2-MBA monolayers prepared using acetic acid/ethanol solvent, the monomers adopt a more prostrate orientation on the Au substrates, in which the aryl ring and carboxyl group of the molecules are tilted {approx} 50{sup o} from the surface normal. This configuration is consistent with an interaction between both the mercaptan sulfur and carboxyl group of 2-MBA with the underlying substrate. S(2p) and C(1s) PES experiments provide supporting evidence for a bidentate interaction between 2-MBA and Au(111).

  13. Synthesis of 5-Substituted Derivatives of Isophthalic Acid as Non-Polymeric Amphiphilic Coating for Metal Oxide Nanoparticles

    PubMed Central

    Nilov, Denis; Kucheryavy, Pavel; Walker, Verina; Kidd, Clayton; Kolesnichenko, Vladimir L.; Goloverda, Galina Z.

    2014-01-01

    In the course of development of novel capping ligands with variable steric factor, which will be used as an organic coating for metal oxide nanoparticles, a base-catalyzed nucleophilic oxirane ring-opening addition reaction between dimethyl 5-hydroxyisophthalate and allyl glycidyl ether was studied. The allyl-terminated 1-1, 1-2 and 1-3 adducts and dihydroxylated derivative of the 1-1 adduct, 5-diglyceroxy isophthalic acid, were synthesized. The latter binds to the surface of 5 nm γ-Fe2O3 nanoparticles in reaction with their surfactant-free diethylene glycol colloids. PMID:25152545

  14. Highly efficient asymmetric hydrogenation of cyano-substituted acrylate esters for synthesis of chiral γ-lactams and amino acids.

    PubMed

    Kong, Duanyang; Li, Meina; Wang, Rui; Zi, Guofu; Hou, Guohua

    2016-01-28

    A highly efficient and enantioselective synthesis of γ-lactams and γ-amino acids by Rh-catalyzed asymmetric hydrogenation has been developed. Using the Rh-(S,S)-f-spiroPhos complex, under mild conditions a wide range of 3-cyano acrylate esters including both E and Z-isomers and β-cyano-α-aryl-α,β-unsaturated ketones were first hydrogenated with excellent enantioselectivities (up to 98% ee) and high turnover numbers (TON up to 10,000). PMID:26661067

  15. Solvent substitution

    SciTech Connect

    Not Available

    1990-01-01

    The DOE Environmental Restoration and Waste Management Office of Technology Development and the Air Force Engineering and Services Center convened the First Annual International Workshop on Solvent Substitution on December 4--7, 1990. The primary objectives of this joint effort were to share information and ideas among attendees in order to enhance the development and implementation of required new technologies for the elimination of pollutants associated with industrial use of hazardous and toxic solvents; and to aid in accelerating collaborative efforts and technology transfer between government and industry for solvent substitution. There were workshop sessions focusing on Alternative Technologies, Alternative Solvents, Recovery/Recycling, Low VOC Materials and Treatment for Environmentally Safe Disposal. The 35 invited papers presented covered a wide range of solvent substitution activities including: hardware and weapons production and maintenance, paint stripping, coating applications, printed circuit boards, metal cleaning, metal finishing, manufacturing, compliance monitoring and process control monitoring. This publication includes the majority of these presentations. In addition, in order to further facilitate information exchange and technology transfer, the US Air Force and DOE solicited additional papers under a general Call for Papers.'' These papers, which underwent review and final selection by a peer review committee, are also included in this combined Proceedings/Compendium. For those involved in handling, using or managing hazardous and toxic solvents, this document should prove to be a valuable resource, providing the most up-to-date information on current technologies and practices in solvent substitution. Individual papers are abstracted separated.

  16. The effect of random precipitation times on the scavenging rate for tropospheric nitric acid

    NASA Technical Reports Server (NTRS)

    Stewart, Richard W.

    1988-01-01

    A model for the effective scavenging rate of a soluble species has been developed. The model takes into account the possibility of positive as well as negative correlations between departures from the mean of the scavenging rate and species concentration. The model is demonstrated for the case of late afternoon rainout of nitric acid occurring just prior to the nighttime cessation of its chemical production. The calculations give effective scavenging rates which are about a factor of 2 to 3 greater than those calculated using the models of Rodhe and Grandell (1972) and Giorgi and Chameides (1985).

  17. Effect of defatting on acid hydrolysis rate of maize starch with different amylose contents.

    PubMed

    Wei, Benxi; Hu, Xiuting; Zhang, Bao; Li, Hongyan; Xu, Xueming; Jin, Zhengyu; Tian, Yaoqi

    2013-11-01

    The effect of defatting on the physiochemical properties and the acid hydrolysis rate of maize starch with different amylose contents was evaluated in this study. The increase in the number of pores and the stripping of starch surface layers were observed after defatting by scanning electron microscopy. X-ray diffraction spectrum showed that the peaks attributing to the amylose-lipid complex disappeared. The relative crystallinity increased by 19% for high-amylose maize starch (HMS) on defatting, while the other tested starches virtually unchanged. Differential scanning calorimetry study indicated an increase in the thermal stability for the defatted starches. Compared with native waxy maize starch, the acid hydrolysis rate of the defatted one increased by 6% after 10 days. For normal maize starch (NMS) and HMS, the higher rate of hydrolysis was observed during the first 5 days. Thereafter, the hydrolysis rate was lower than that of their native counterpart. The increase in susceptibility to acid hydrolysis (in the first 5 days) was mainly attributed to the defective and porous structures formed during defatting process, while the decrease of hydrolysis rate for NMS and HMS samples (after the first 5 days) probably resulted from the increase in the relative crystallinity.

  18. Comparison of parameterized nitric acid rainout rates using a coupled stochastic-photochemical tropospheric model

    NASA Technical Reports Server (NTRS)

    Stewart, Richard W.; Thompson, Anne M.; Owens, Melody A.; Herwehe, Jerold A.

    1989-01-01

    A major tropospheric loss of soluble species such as nitric acid results from scavenging by water droplets. Several theoretical formulations have been advanced which relate an effective time-independent loss rate for soluble species to statistical properties of precipitation such as the wet fraction and length of a precipitation cycle. In this paper, various 'effective' loss rates that have been proposed are compared with the results of detailed time-dependent model calculations carried out over a seasonal time scale. The model is a stochastic precipitation model coupled to a tropospheric photochemical model. The results of numerous time-dependent seasonal model runs are used to derive numerical values for the nitric acid residence time for several assumed sets of preciptation statistics. These values are then compared with the results obtained by utilizing theoretical 'effective' loss rates in time-independent models.

  19. Synthesis, antiproliferative and antifungal activities of 1,2,3-triazole-substituted carnosic Acid and carnosol derivatives.

    PubMed

    Pertino, Mariano Walter; Theoduloz, Cristina; Butassi, Estefania; Zacchino, Susana; Schmeda-Hirschmann, Guillermo

    2015-01-01

    Abietane diterpenes exhibit an array of interesting biological activities, which have generated significant interest among the pharmacological community. Starting from the abietane diterpenes carnosic acid and carnosol, twenty four new triazole derivatives were synthesized using click chemistry. The compounds differ in the length of the linker and the substituent on the triazole moiety. The compounds were assessed as antiproliferative and antifungal agents. The antiproliferative activity was determined on normal lung fibroblasts (MRC-5), gastric epithelial adenocarcinoma (AGS), lung cancer (SK-MES-1) and bladder carcinoma (J82) cells while the antifungal activity was assessed against Candida albicans ATCC 10231 and Cryptococcus neoformans ATCC 32264. The carnosic acid γ-lactone derivatives 1-3 were the most active antiproliferative compounds of the series, with IC50 values in the range of 43.4-46.9 μM and 39.2-48.9 μM for MRC-5 and AGS cells, respectively. Regarding antifungal activity, C. neoformans was the most sensitive fungus, with nine compounds inhibiting more than 50% of its fungal growth at concentrations ≤250 µg∙mL-1. Compound 22, possessing a p-Br-benzyl substituent on the triazole ring, showed the best activity (91% growth inhibition) at 250 µg∙mL-1 In turn, six compounds inhibited 50% C. albicans growth at concentrations lower than 250 µg∙mL-1. PMID:26007173

  20. Amino acid substitutions in the coat protein result in loss of insect transmissibility of a plant virus.

    PubMed Central

    Atreya, P L; Atreya, C D; Pirone, T P

    1991-01-01

    Amino acids near the N terminus of the coat protein of tobacco vein mottling virus were deleted or altered by site-directed mutagenesis to determine the effect on aphid transmissibility of the virus. Deletion of a three amino acid sequence Asp-Ala-Gly, which is conserved in aphid-transmissible potyvirus isolates, abolished transmission. The mutation Ala----Thr in this triplet drastically reduced transmission, whereas the mutation Asp----Asn had no effect, and the mutation Asp----Lys consistently reverted to the wild-type residue. The mutation Lys----Glu, in the residue adjacent to the glycine of the triplet, drastically reduced transmission, whereas the mutation Gln----Pro, seven residues downstream from the glycine had no effect. Comparison of the sequences of other potyviruses suggests that the presence of a glycine residue at the third position of the Asp-Ala-Gly triplet is critical for aphid transmissibility and that certain changes in the residues adjacent to this position abolish or greatly reduce aphid transmissibility. PMID:1881922

  1. Evidence That the Origin of Naked Kernels During Maize Domestication Was Caused by a Single Amino Acid Substitution in tga1.

    PubMed

    Wang, Huai; Studer, Anthony J; Zhao, Qiong; Meeley, Robert; Doebley, John F

    2015-07-01

    teosinte glume architecture1 (tga1), a member of the SBP-box gene family of transcriptional regulators, has been identified as the gene conferring naked kernels in maize vs. encased kernels in its wild progenitor, teosinte. However, the identity of the causative polymorphism within tga1 that produces these different phenotypes has remained unknown. Using nucleotide diversity data, we show that there is a single fixed nucleotide difference between maize and teosinte in tga1, and this difference confers a Lys (teosinte allele) to Asn (maize allele) substitution. This substitution transforms TGA1 into a transcriptional repressor. While both alleles of TGA1 can bind a GTAC motif, maize-TGA1 forms more stable dimers than teosinte-TGA1. Since it is the only fixed difference between maize and teosinte, this alteration in protein function likely underlies the differences in maize and teosinte glume architecture. We previously reported a difference in TGA1 protein abundance between maize and teosinte based on relative signal intensity of a Western blot. Here, we show that this signal difference is not due to tga1 but to a second gene, neighbor of tga1 (not1). Not1 encodes a protein that has 92% amino acid similarity to TGA1 and that is recognized by the TGA1 antibody. Genetic mapping and phenotypic data show that tga1, without a contribution from not1, controls the difference in covered vs. naked kernels. No trait differences could be associated with the maize vs. teosinte alleles of not1. Our results document how morphological evolution can be driven by a simple nucleotide change that alters protein function.

  2. Modulation of Enzymatic Activity and Biological Function of Listeria monocytogenes Broad-Range Phospholipase C by Amino Acid Substitutions and by Replacement with the Bacillus cereus Ortholog

    PubMed Central

    Zückert, Wolfram R.; Marquis, Hélène; Goldfine, Howard

    1998-01-01

    The secreted broad-range phosphatidylcholine (PC)-preferring phospholipase C (PC-PLC) of Listeria monocytogenes plays a role in the bacterium’s ability to escape from phagosomes and spread from cell to cell. Based on comparisons with two orthologs, Clostridium perfringens α-toxin and Bacillus cereus PLC (PLCBc), we generated PC-PLC mutants with altered enzymatic activities and substrate specificities and analyzed them for biological function in tissue culture and mouse models of infection. Two of the conserved active-site zinc-coordinating histidines were confirmed by single amino acid substitutions H69G and H118G, which resulted in proteins inactive in broth culture and unstable intracellularly. Substitutions D4E and H56Y remodeled the PC-PLC active site to more closely resemble the PLCBc active site, while a gene replacement resulted in L. monocytogenes secreting PLCBc. All of these mutants yielded similar amounts of active enzyme as wild-type PC-PLC both in broth culture and intracellularly. D4E increased activity on and specificity for PC, while H56Y and D4E H56Y showed higher activity on both PC and sphingomyelin, with reduced specificity for PC. As expected, PLCBc expressed by L. monocytogenes was highly specific for PC. During early intracellular growth in human epithelial cells, the D4E mutant and the PLCBc-expressing strain performed significantly better than the wild type, while the H56Y and D4E H56Y mutants showed a significant defect. In assays for cell-to-cell spread, the H56Y and D4E mutants had close to wild-type characteristics, while the spreading efficiency of PLCBc was significantly lower. These studies emphasize the species-specific features of PC-PLC important for growth in mammalian cells. PMID:9746585

  3. A sensitive real-time PCR based assay to estimate the impact of amino acid substitutions on the competitive replication fitness of human immunodeficiency virus type 1 in cell culture.

    PubMed

    Liu, Yi; Holte, Sarah; Rao, Ushnal; McClure, Jan; Konopa, Philip; Swain, J Victor; Lanxon-Cookson, Erinn; Kim, Moon; Chen, Lennie; Mullins, James I

    2013-04-01

    Fixation of mutations in human immunodeficiency virus type 1 (HIV-1), such as those conferring drug resistance and immune escape, can result in a change in replication fitness. To assess these changes, a real-time TaqMan PCR detection assay and statistical methods for data analysis were developed to estimate sensitively relative viral fitness in competitive viral replication experiments in cell culture. Chimeric viruses with the gene of interest in an HIV-1NL4-3 backbone were constructed in two forms, vifA (native vif gene in NL4-3) and vifB (vif gene with six synonymous nucleotide differences from vifA). Subsequently, mutations of interest were introduced into the chimeric viruses in NL4-3VifA backbones, and the mutants were competed against the chimera with the isogenic viral sequence in the NL4-3VifB backbone in cell culture. In order to assess subtle fitness differences, culture supernatants were sampled longitudinally, and the viruses differentially quantified using vifA- and vifB-specific primers in real-time PCR assays. Based on an exponential net growth model, the growth rate of each virus was determined and the fitness cost of the mutation(s) distinguishing the two viruses represented as the net growth rate difference between the mutant and the native variants. Using this assay, the fitness impact of eight amino acid substitutions was quantitated at highly conserved sites in HIV-1 Gag and Env. PMID:23201292

  4. Measurement and Analysis of Extracellular Acid Production to Determine Glycolytic Rate.

    PubMed

    Mookerjee, Shona A; Brand, Martin D

    2015-12-12

    Extracellular measurement of oxygen consumption and acid production is a simple and powerful way to monitor rates of respiration and glycolysis(1). Both mitochondrial (respiration) and non-mitochondrial (other redox) reactions consume oxygen, but these reactions can be easily distinguished by chemical inhibition of mitochondrial respiration. However, while mitochondrial oxygen consumption is an unambiguous and direct measurement of respiration rate(2), the same is not true for extracellular acid production and its relationship to glycolytic rate (3-6). Extracellular acid produced by cells is derived from both lactate, produced by anaerobic glycolysis, and CO2, produced in the citric acid cycle during respiration. For glycolysis, the conversion of glucose to lactate(-) + H(+) and the export of products into the assay medium is the source of glycolytic acidification. For respiration, the export of CO2, hydration to H2CO3 and dissociation to HCO3(-) + H(+) is the source of respiratory acidification. The proportions of glycolytic and respiratory acidification depend on the experimental conditions, including cell type and substrate(s) provided, and can range from nearly 100% glycolytic acidification to nearly 100% respiratory acidification (6). Here, we demonstrate the data collection and calculation methods needed to determine respiratory and glycolytic contributions to total extracellular acidification by whole cells in culture using C2C12 myoblast cells as a model.

  5. Dating the origin of hepatitis B virus reveals higher substitution rate and adaptation on the branch leading to F/H genotypes.

    PubMed

    Paraskevis, Dimitrios; Angelis, Konstantinos; Magiorkinis, Gkikas; Kostaki, Evangelia; Ho, Simon Y W; Hatzakis, Angelos

    2015-12-01

    The evolution of hepatitis B virus (HBV), particularly its origins and evolutionary timescale, has been the subject of debate. Three major scenarios have been proposed, variously placing the origin of HBV in humans and great apes from some million years to only a few thousand years ago (ka). To compare these scenarios, we analyzed 105 full-length HBV genome sequences from all major genotypes sampled globally. We found a high correlation between the demographic histories of HBV and humans, as well as coincidence in the times of origin of specific subgenotypes with human migrations giving rise to their host indigenous populations. Together with phylogenetic evidence, this suggests that HBV has co-expanded with modern humans. Based on the co-expansion, we conducted a Bayesian dating analysis to estimate a precise evolutionary timescale for HBV. Five calibrations were used at the origins of F/H genotypes, D4, C3 and B6 from respective indigenous populations in the Pacific and Arctic and A5 from Haiti. The estimated time for the origin of HBV was 34.1ka (95% highest posterior density interval 27.6-41.3ka), coinciding with the dispersal of modern non-African humans. Our study, the first to use full-length HBV sequences, places a precise timescale on the HBV epidemic and also shows that the "branching paradox" of the more divergent genotypes F/H from Amerindians is due to an accelerated substitution rate, probably driven by positive selection. This may explain previously observed differences in the natural history of HBV between genotypes F1 and A2, B1, and D.

  6. Genetic Basis of Differential Heat Resistance between Two Species of Congeneric Freshwater Snails: Insights from Quantitative Proteomics and Base Substitution Rate Analysis.

    PubMed

    Mu, Huawei; Sun, Jin; Fang, Ling; Luan, Tiangang; Williams, Gray A; Cheung, Siu Gin; Wong, Chris K C; Qiu, Jian-Wen

    2015-10-01

    We compared the heat tolerance, proteomic responses to heat stress, and adaptive sequence divergence in the invasive snail Pomacea canaliculata and its noninvasive congener Pomacea diffusa. The LT50 of P. canaliculata was significantly higher than that of P. diffusa. More than 3350 proteins were identified from the hepatopancreas of the snails exposed to acute and chronic thermal stress using iTRAQ-coupled mass spectrometry. Acute exposure (3 h exposure at 37 °C with 25 °C as control) resulted in similar numbers (27 in P. canaliculata and 23 in P. diffusa) of differentially expressed proteins in the two species. Chronic exposure (3 weeks of exposure at 35 °C with 25 °C as control) caused differential expression of more proteins (58 in P. canaliculata and 118 in P. diffusa), with many of them related to restoration of damaged molecules, ubiquitinating dysfunctional molecules, and utilization of energy reserves in both species; but only in P. diffusa was there a shift from carbohydrate to lipid catabolism. Analysis of orthologous genes encoding the differentially expressed proteins revealed two genes having clear evidence of positive selection (Ka/Ks > 1) and seven candidates for more detailed analysis of positive selection (Ka/Ks between 0.5 and 1). These nine genes are related to energy metabolism, cellular oxidative homeostasis, signaling, and binding processes. Overall, the proteomic and base substitution rate analyses indicate genetic basis of differential resistance to heat stress between the two species, and such differences could affect their further range expansion in a warming climate.

  7. Phylogeny of Arthropoda inferred from mitochondrial sequences: strategies for limiting the misleading effects of multiple changes in pattern and rates of substitution.

    PubMed

    Hassanin, Alexandre

    2006-01-01

    In this study, mitochondrial sequences were used to investigate the relationships among the major lineages of Arthropoda. The data matrix used for the analyses includes 84 taxa and 3918 nucleotides representing six mitochondrial protein-coding genes (atp6 and 8, cox1-3, and nad2). The analyses of nucleotide composition show that a reverse strand-bias, i.e., characterized by an excess of T relative to A nucleotides and of G relative to C nucleotides, was independently acquired in six different lineages of Arthropoda: (1) the honeybee mite (Varroa), (2) Opisthothelae spiders (Argiope, Habronattus, and Ornithoctonus), (3) scorpions (Euscorpius and Mesobuthus), (4) Hutchinsoniella (Cephalocarid), (5) Tigriopus (Copepod), and (6) whiteflies (Aleurodicus and Trialeurodes). Phylogenetic analyses confirm that these convergences in nucleotide composition can be particularly misleading for tree reconstruction, as unrelated taxa with reverse strand-bias tend to group together in MP, ML, and Bayesian analyses. However, the use of a specific model for minimizing effects of the bias, the "Neutral Transition Exclusion" (NTE) model, allows Bayesian analyses to rediscover most of the higher taxa of Arthropoda. Furthermore, the analyses of branch lengths suggest that three main factors explain accelerated rates of substitution: (1) genomic rearrangements, including duplication of the control region and gene translocation, (2) parasitic lifestyle, and (3) small body size. The comparisons of Bayesian Bootstrap percentages show that the support for many nodes increases when taxa with long branches are excluded from the analyses. It is therefore recommended to select taxa and genes of the mitochondrial genome for inferring phylogenetic relationships among arthropod lineages. The phylogenetic analyses support the existence of a major dichotomy within Arthropoda, separating Pancrustacea and Paradoxopoda. Basal relationships between Pancrustacean lineages are not robust, and the question

  8. Substitution of a single amino acid residue in the aromatic/arginine selectivity filter alters the transport profiles of tonoplast aquaporin homologs.

    PubMed

    Azad, Abul Kalam; Yoshikawa, Naoki; Ishikawa, Takahiro; Sawa, Yoshihiro; Shibata, Hitoshi

    2012-01-01

    Aquaporins are integral membrane proteins that facilitate the transport of water and some small solutes across cellular membranes. X-ray crystallography of aquaporins indicates that four amino acids constitute an aromatic/arginine (ar/R) pore constriction known as the selectivity filter. On the basis of these four amino acids, tonoplast aquaporins called tonoplast intrinsic proteins (TIPs) are divided into three groups in Arabidopsis. Herein, we describe the characterization of two group I TIP1s (TgTIP1;1 and TgTIP1;2) from tulip (Tulipa gesneriana). TgTIP1;1 and TgTIP1;2 have a novel isoleucine in loop E (LE2 position) of the ar/R filter; the residue at LE2 is a valine in all group I TIPs from model plants. The homologs showed mercury-sensitive water channel activity in a fast kinetics swelling assay upon heterologous expression in Pichia pastoris. Heterologous expression of both homologs promoted the growth of P. pastoris on ammonium or urea as sole sources of nitrogen and decreased growth and survival in the presence of H(2)O(2). TgTIP1;1- and TgTIP1;2-mediated H(2)O(2) conductance was demonstrated further by a fluorescence assay. Substitutions in the ar/R selectivity filter of TgTIP1;1 showed that mutants that mimicked the ar/R constriction of group I TIPs could conduct the same substrates that were transported by wild-type TgTIP1;1. In contrast, mutants that mimicked group II TIPs showed no evidence of urea or H(2)O(2) conductance. These results suggest that the amino acid residue at LE2 position is critical for the transport selectivity of the TIP homologs and group I TIPs might have a broader spectrum of substrate selectivity than group II TIPs.

  9. Fluorogenic Thorium Sensors Based on 2,6-Pyridinedicarboxylic Acid-Substituted Tetraphenylethenes with Aggregation-Induced Emission Characteristics.

    PubMed

    Wen, Jun; Dong, Liang; Hu, Sheng; Li, Weiyi; Li, Shuo; Wang, Xiaolin

    2016-01-01

    A novel fluorescent sensor based on tetraphenylethene (TPE) modified with 2,6-pyridinedicarboxylic acid (PDA) that shows aggregation-induced emission (AIE) characteristics for thorium recognition with remarkable fluoresence enhancement response has been synthesized. This sensor is capable of visually distinguishing Th(4+) among lanthanides, transition metals, and alkali metals under UV light. Th(4+) can be detected by the naked eye at ppb levels owing to the AIE phenomenon. The sensor showed high selectivity for Th(4+) compared to all other metals tested, and this recognition displayed good anti-interference qualities. This study represents the first application of a AIE fluorescence sensor in actinide metal recognition and it has potential applications in environmental systems for thorium ion detection. PMID:26419754

  10. ModelOMatic: fast and automated model selection between RY, nucleotide, amino acid, and codon substitution models.

    PubMed

    Whelan, Simon; Allen, James E; Blackburne, Benjamin P; Talavera, David

    2015-01-01

    Molecular phylogenetics is a powerful tool for inferring both the process and pattern of evolution from genomic sequence data. Statistical approaches, such as maximum likelihood and Bayesian inference, are now established as the preferred methods of inference. The choice of models that a researcher uses for inference is of critical importance, and there are established methods for model selection conditioned on a particular type of data, such as nucleotides, amino acids, or codons. A major limitation of existing model selection approaches is that they can only compare models acting upon a single type of data. Here, we extend model selection to allow comparisons between models describing different types of data by introducing the idea of adapter functions, which project aggregated models onto the originally observed sequence data. These projections are implemented in the program ModelOMatic and used to perform model selection on 3722 families from the PANDIT database, 68 genes from an arthropod phylogenomic data set, and 248 genes from a vertebrate phylogenomic data set. For the PANDIT and arthropod data, we find that amino acid models are selected for the overwhelming majority of alignments; with progressively smaller numbers of alignments selecting codon and nucleotide models, and no families selecting RY-based models. In contrast, nearly all alignments from the vertebrate data set select codon-based models. The sequence divergence, the number of sequences, and the degree of selection acting upon the protein sequences may contribute to explaining this variation in model selection. Our ModelOMatic program is fast, with most families from PANDIT taking fewer than 150 s to complete, and should therefore be easily incorporated into existing phylogenetic pipelines. ModelOMatic is available at https://code.google.com/p/modelomatic/.

  11. A single amino acid substitution confers high cinchonidine oxidation activity comparable with that of rabbit to monkey aldehyde oxidase 1.

    PubMed

    Fukiya, Kensuke; Itoh, Kunio; Yamaguchi, Satoshi; Kishiba, Akiko; Adachi, Mayuko; Watanabe, Nobuaki; Tanaka, Yorihisa

    2010-02-01

    Aldehyde oxidase 1 (AOX1) is a major member of the xanthine oxidase family belonging to the class of complex molybdo-flavoenzymes and plays an important role in the nucleophilic oxidation of N-heterocyclic aromatic compounds and various aldehydes. The enzyme has been well known to show remarkable species differences. Comparing the rabbit and monkey enzymes, the former showed extremely high activity toward cinchonidine and methotrexate, but the latter exhibited only marginal activities. In contrast, monkey had several times greater activity than did rabbit toward zonisamide and (+)-4-(4-cyanoanilino)-5,6-dihydro-7-hydroxy-7H-cyclopenta[d]-pyrimidine [(S)-RS-8359]. In this report, we tried to confer high cinchonidine oxidation activity comparable with that of rabbit AOX1 to monkey AOX1. The chimera proteins prepared by restriction enzyme digestion and recombination methods between monkey and rabbit AOX1s indicated that the sequences from Asn993 to Ala1088 of rabbit AOX1 are essential for the activity. The kinetic parameters were then measured using monkey AOX1 mutants prepared by site-directed mutagenesis. The monkey V1085A mutant acquired the high cinchonidine oxidation activity. Inversely, the reciprocal rabbit A1081V mutant lost the activity entirely: amino acid 1081 of rabbit AOX1 corresponding to amino acid 1085 of monkey AOX1. Thus, cinchonidine oxidation activity was drastically changed by mutation of a single residue in AOX1. However, this might be true for bulky substrates such as cinchonidine but not for small substrates. The mechanism of substrate-dependent species differences in AOX1 activity toward bulky substrates is discussed.

  12. Acid-base chemical reaction model for nucleation rates in the polluted atmospheric boundary layer

    NASA Astrophysics Data System (ADS)

    Chen, Modi; Titcombe, Mari; Jiang, Jingkun; Jen, Coty; Kuang, Chongai; Fischer, Marc L.; Eisele, Fred L.; Siepmann, J. Ilja; Hanson, David R.; Zhao, Jun; McMurry, Peter H.

    2013-05-01

    Measurements of aerosol number distributions down to one molecule have provided information that we've used to develop a new approach for modeling atmospheric nucleation rates. Measurements were carried out with the Cluster Chemical Ionization Mass Spectrometer (Cluster CIMS), the scanning mobility spectrometer using a diethylene glycol condensation particle counter as detector (DEG SMPS), and an ambient pressure proton transfer mass spectrometer for ammonia and amines (AmPMS). The model explains nucleation as a result of cluster evolution due to a sequence of acid-base reactions. We conclude that the smallest stable cluster contains four sulfuric acid molecules. The model leads to a simple analytic expression for nucleation rates that is reasonably consistent (i.e., ± 10x) with atmospheric observations. The model predicts that nucleation rates are equal to a prefactor, P<1, times the sulfuric acid vapor collision rate, (i.e., J=Pṡ0.5ṡk11 *[H2SO4]2).

  13. Diverse driving forces underlie the invariant occurrence of the T42A, E139D, I282V and T468M SHP2 amino acid substitutions causing Noonan and LEOPARD syndromes

    PubMed Central

    Martinelli, Simone; Torreri, Paola; Tinti, Michele; Stella, Lorenzo; Bocchinfuso, Gianfranco; Flex, Elisabetta; Grottesi, Alessandro; Ceccarini, Marina; Palleschi, Antonio; Cesareni, Gianni; Castagnoli, Luisa; Petrucci, Tamara C.; Gelb, Bruce D.; Tartaglia, Marco

    2008-01-01

    Missense PTPN11 mutations cause Noonan and LEOPARD syndromes (NS and LS), two developmental disorders with pleiomorphic phenotypes. PTPN11 encodes SHP2, an SH2 domain-containing protein tyrosine phosphatase functioning as a signal transducer. Generally, different substitutions of a particular amino acid residue are observed in these diseases, indicating that the crucial factor is the residue being replaced. For a few codons, only one substitution is observed, suggesting the possibility of specific roles for the residue introduced. We analyzed the biochemical behavior and ligand-binding properties of all possible substitutions arising from single-base changes affecting codons 42, 139, 279, 282 and 468 to investigate the mechanisms underlying the invariant occurrence of the T42A, E139D and I282V substitutions in NS and the Y279C and T468M changes in LS. Our data demonstrate that the isoleucine-to-valine change at codon 282 is the only substitution at that position perturbing the stability of SHP2's closed conformation without impairing catalysis, while the threonine-to-alanine change at codon 42, but not other substitutions of that residue, promotes increased phosphopeptide-binding affinity. The recognition specificity of the C-SH2 domain bearing the E139D substitution differed substantially from its wild-type counterpart acquiring binding properties similar to those observed for the N-SH2 domain, revealing a novel mechanism of SHP2's functional dysregulation. Finally, while functional selection does not seem to occur for the substitutions at codons 279 and 468, we point to deamination of the methylated cytosine at nucleotide 1403 as the driving factor leading to the high prevalence of the T468M change in LS. PMID:18372317

  14. Sensory Substitution

    NASA Astrophysics Data System (ADS)

    Verrillo, Ronald T.

    The idea that the cutaneous surface may be employed as a substitute for the eyes and ears is by no means a modern notion. Although the sense of touch has long been considered as a surrogate for both the visual and auditory modalities, the focus of this chapter will be on the efforts to develop a tactile substitute for hearing, especially that of human speech. The visual system is our primary means of processing information about environmental space such as orientation, distance, direction and size. It is much less effective in making temporal discriminations. The auditory system is unparalleled in processing information that involves rapid sequences of temporal events, such as speech and music. The tactile sense is capable of processing both spatial and temporal information although not as effective in either domain as the eye or the ear.

  15. Cytochrome c Trp65Ser substitution results in inhibition of acetic acid-induced programmed cell death in Saccharomyces cerevisiae.

    PubMed

    Guaragnella, Nicoletta; Passarella, Salvatore; Marra, Ersilia; Giannattasio, Sergio

    2011-11-01

    To gain further insight into the role of cytochrome c (cyt c) in yeast programmed cell death induced by acetic acid (AA-PCD), comparison was made between wild type and two mutant cells, one lacking cyt c and the other (W65Scyc1) expressing a mutant iso-1-cyt c in a form unable to reduce cyt c oxidase, with respect to occurrence of AA-PCD, cyt c release, ROS production and caspase-like activity. We show that in W65Scyc1 cells: i. no release of mutant cyt c occurs with inhibition of W65Scyc1 cell AA-PCD shown to be independent on impairment of electron flow, ii. there is a decrease in ROS production and an increase in caspase-like activity. We conclude that cyt c release does not depend on cyt c function as an electron carrier and that when still associated to the mitochondrial membrane, cyt c in its reduced form has a role in AA-PCD, by regulating ROS production and caspase-like activity. PMID:21907312

  16. Separation of 1,3-substituted imidazoles for quality control of a Lewis acidic ionic liquid for aluminum electroplating.

    PubMed

    Kosmus, Patrick; Steiner, Oliver; Goessler, Walter; Gollas, Bernhard

    2014-05-01

    Ionic liquids (ILs) are already used or have great potential in many industrial applications. Knowledge about their unique physicochemical characteristics makes ILs suitable for the electrodeposition of metals with very low negative potentials. Aluminum with its good corrosion protection behavior has great capability to be electroplated from IL electrolytes on steel substrates. The stability of the chosen electrolyte is very important to ensure industrial applicability. In this study, temperature and electrochemical long-term stability from electrolytes based on a Lewis acidic mixture of AlCl3 and 1-ethyl-3-methylimidazolium chloride are investigated. A published method was modified to identify possible degradation products using mass spectrometric detection. The optimized method used an Agilent Zorbax SB-Phenyl column (2.0 × 150 mm, 5 μm particles) with a 20 mmol TFA and 5% ACN mobile phase. This method allowed the quantification of several imidazoles from 0.1 to 100 mg/L. When analyzing the long-term stressed electrolytes, no significant changes in electrolyte composition could be observed.

  17. A single amino acid substitution in the ORF1 of cymbidium ringspot virus determines the accumulation of two satellite RNAs.

    PubMed

    Rubino, Luisa; Russo, Marcello

    2012-09-01

    Tombusviruses may support the replication of satellite (sat) RNAs. In particular, two satRNAs, sat L and Cymsat RNAs, are replicated by carnation Italian ringspot (CIRV) and tomato bushy stunt (TBSV) virus, but not by cymbidium ringspot virus (CymRSV) in vitro transcripts unless they contain a poly(A) tail at the 3' end. Conversely, the replication of both satRNAs was supported by virus particles or viral RNA of the original CymRSV inoculum even in the absence of the poly(A) tail. Sequence and mutational analyses revealed that the full-length infectious CymRSV clone contains one relevant sequence variation in the ORF 1-encoded protein (p33) compared with the original inoculum, i.e. a Ser₁₉ TCC codon instead of a Phe₁₉ TTC codon, which inhibited the replication of sat L and Cymsat RNAs. It is suggested that this amino acid is contained in a domain essential for the replication of some subviral RNAs. PMID:22709553

  18. Comparison and preparation of multilayered polylactic acid fabric strengthen calcium phosphate-based bone substitutes for orthopedic applications.

    PubMed

    Chen, Wen-Cheng; Ko, Chia-Ling; Yang, Jia-Kai; Wu, Hui-Yu; Lin, Jia-Horng

    2016-03-01

    An attempt to maintain the three-dimensional space into restorative sites through the conveniently pack porous fillers are general used strategy. Advancement in the manufacturing protective shells in the scaffolds, which would be filled with brittle ceramic grafts for the development of highly connective pores provides the approach to solve crack problem for generating the tissues. Therefore, multilayered braided and alkalized poly(lactic acid) (PLA) composites with calcium phosphate bone cement (CPC) were synthesized and compared. The PLA/CPC composites were divided into various groups according to a series of heat-treatment temperatures (100-190 °C) and periods (1-3 h) and then characterized. The effects of 24-h immersion on the strength decay resistance of the samples were compared. Results showed that the residual oil capped on the surfaces of alkalized PLA braid was removed, and the structure was unaltered. However, the reduced tensile stress of alkalized PLA braids was due to ester-group formation by hydrolysis. Mechanical test results of PLA/CPC composites showed that the strength significantly increased after heat treatment, except when the heating temperature was higher than the PLA melting point at approximately 160-170 °C. The degree of PLA after recrystallization became higher than that of unheated composites, thereby leading to reduced strength and toughness of the specimen. Braiding fibers of biodegradable PLA reinforced and toughened the structure particularly of the extra-brittle material of thin-sheet CPC after implantation.

  19. Hemoglobin substitutes.

    PubMed

    Anbari, Kevin K; Garino, Jonathan P; Mackenzie, Colin F

    2004-10-01

    Orthopaedic patients frequently require blood transfusions to treat peri-operative anemia. Research in the area of hemoglobin substitutes has been of great interest since it holds the promise of reducing the reliance on allogeneic blood transfusions. The three categories of hemoglobin substitutes are (1) cell-free, extracellular hemoglobin preparations made from human or bovine hemoglobin (hemoglobin-based oxygen carriers or HBOCs); (2) fluorine-substituted linear or cyclic carbon chains with a high oxygen-carrying capacity (perfluorocarbons); and (3) liposome-encapsulated hemoglobin. Of the three, HBOCs have been the most extensively studied and tested in preclinical and clinical trials that have shown success in diminishing the number of blood transfusions as well as an overall favorable side-effect profile. This has been demonstrated in vascular, cardiothoracic, and orthopaedic patients. HBOC-201, which is a preparation of cell-free bovine hemoglobin, has been approved for clinical use in South Africa. These products may well become an important tool for physicians treating peri-operative anemia in orthopaedic patients.

  20. A Single Amino-Acid Substitution in the Sodium Transporter HKT1 Associated with Plant Salt Tolerance.

    PubMed

    Ali, Akhtar; Raddatz, Natalia; Aman, Rashid; Kim, Songmi; Park, Hyeong Cheol; Jan, Masood; Baek, Dongwon; Khan, Irfan Ullah; Oh, Dong-Ha; Lee, Sang Yeol; Bressan, Ray A; Lee, Keun Woo; Maggio, Albino; Pardo, Jose M; Bohnert, Hans J; Yun, Dae-Jin

    2016-07-01

    A crucial prerequisite for plant growth and survival is the maintenance of potassium uptake, especially when high sodium surrounds the root zone. The Arabidopsis HIGH-AFFINITY K(+) TRANSPORTER1 (HKT1), and its homologs in other salt-sensitive dicots, contributes to salinity tolerance by removing Na(+) from the transpiration stream. However, TsHKT1;2, one of three HKT1 copies in Thellungiella salsuginea, a halophytic Arabidopsis relative, acts as a K(+) transporter in the presence of Na(+) in yeast (Saccharomyces cerevisiae). Amino-acid sequence comparisons indicated differences between TsHKT1;2 and most other published HKT1 sequences with respect to an Asp residue (D207) in the second pore-loop domain. Two additional T salsuginea and most other HKT1 sequences contain Asn (n) in this position. Wild-type TsHKT1;2 and altered AtHKT1 (AtHKT1(N-D)) complemented K(+)-uptake deficiency of yeast cells. Mutant hkt1-1 plants complemented with both AtHKT1(N) (-) (D) and TsHKT1;2 showed higher tolerance to salt stress than lines complemented by the wild-type AtHKT1 Electrophysiological analysis in Xenopus laevis oocytes confirmed the functional properties of these transporters and the differential selectivity for Na(+) and K(+) based on the n/d variance in the pore region. This change also dictated inward-rectification for Na(+) transport. Thus, the introduction of Asp, replacing Asn, in HKT1-type transporters established altered cation selectivity and uptake dynamics. We describe one way, based on a single change in a crucial protein that enabled some crucifer species to acquire improved salt tolerance, which over evolutionary time may have resulted in further changes that ultimately facilitated colonization of saline habitats.