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Biochemical characterization of branched chain fatty acid biosynthesis
Get PDFConsistent with the fact that Escherichia coli synthesizes straight chain fatty acids, extracts from this organism can utilize acetyl-CoA, but not isovaleryl-CoA, to prime a single cycle of fatty acid biosynthesis. In contrast, extract of Bacillus subtilis, an organism that synthesizes methyl-branched fatty acids, can utilize both acetyl-CoA and isovaleryl-CoA as a primer for a single cycle of fatty acid biosynthesis. These observations indicate that B. subtilis contains a novel 13 ketoacyl-acyl carrier protein synthase that can initiate branched chain fatty biosynthesis using isovalerylCoA as a substrate. This enzyme, which we call 13-ketoacyl-acyl carrier protein synthase IIIB was purified to homogeneity by a three step procedure. The molecular weight of the protein is 44,000 as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified enzyme is highly specific for isovaleryl CoA; Kₘ for isovaleryl-CoA was 35.8 µM. Acetyl CoA could not substitute for the branched chain substrates. This specificity of KAS IIIB suggests it has a role in catalyzing the initial reaction of branched chain fatty acid biosynthesis.
The biosynthesis of fatty acid requires the action of three condensing enzymes, β-ketoacyl-ACP synthase I, II and Ill. β-ketoacyl-ACP synthase Ill catalyzes the first condensation reaction of this process in Escherichia coli and this is encoded by the fabH gene. Using a recombination-based procedure we have disrupted the fabH gene of E.coli. The resulting strain can grow, although at a reduced rate. The exogenous supply of butyrate but not valerate or isovalerate alleviates this reduced growth rate. These results indicate that the fabH gene is not essential and that the deficiency in β-ketoacyl-ACP synthase Ill can be partially overcome by the action of β-ketoacyl-ACP synthase I
Molecular biology of the 3 methylcrotonyl CoA carboxylase subunits
Get PDFThe cDNA and gene coding for a 64 kDa polypeptide have been cloned from Arabidopsis thaliana. The polypeptide gene product has been immunologically identified as the non-biotinylated subunit of 3-methylcrotonyl-CoA carboxylase (MCCase). This is the first time this subunit has been cloned from any organism. The Arabidopsis non-biotinylated subunit of MCCase is encoded by a single gene containing nine introns. The N-terminus of the deduced amino acid sequence contains several transit peptide characteristics, including a predicted amphiphilic alpha-helix. The primary sequence of the non-biotinylated subunit of MCCase shows highest homology to the carboxyltransferase domains of biotin-containing enzymes which have methyl-branched thiol ester substrates or products: methylmalonyl-CoA decarboxylase, propionyl-CoA carboxylase, and transcarboxylase (34.75%, 32.36%, and 33.20% amino acid identity, respectively). In addition, the non-biotinylated subunit of MCCase shows 56.28% amino acid identity to an unidentified Caenorhabditis elegans protein; this C. elegans homolog is likely a MCCase non-biotinylated subunit from the animal kingdom
Going Beyond Counting First Authors in Author Co-citation Analysis
Get PDFThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Profiling of metabolites and volatile flavour compounds from Solanum species using gas chromatograph-mass spectrometry
No full textImproving thermostability of Aspergillus awamori glucoamylase by directed evolution
Get PDFTo improve the thermostability of glucoamylase (GA), random mutagenesis followed by random recombination (directional evolution) were performed on a cloned Aspergillus awamori GA gene expressed in Saccharomyces cerevisiae. We have successfully developed a starch plate assay method to screen thermostable GAs. The results of the plate assay and kinetic analyses of irreversible thermoinactivation were consistent. Several multiple mutants, which produced the most thermostable GAs according to the plate assay, were isolated and characterized by DNA sequencing. Their enhanced thermostability has been confirmed by kinetic analysis of irreversible thermoinactivation. Four new mutations, T62A, S119P, T290A and H391Y, have been identified that are more thermostable than wild type GA and their improvements in thermostability are cumulative. Combinations of these individual mutations and the thermostable mutations constructed previously in the lab by site-directed mutagenesis have been made to create a multiple mutant THS8 (D20C/A27C/S30P/T62A/S119P/T290A/H391Y) with substantially increased thermostability, as revealed by an increase of 50C in melting temperature, and 5.0 kJ/mol in thermoinactivation free energy at 800C, compared to wild-type GA. All single or multiple mutant GAs have similar activity and catalytic efficiency (k[Subscript cat]/K[Subscript m]) as wild-type GA
Variations on the Author
Get PDF“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Get PDFWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
The Effects of Carbon Source and Growth Temperature on the Fatty Acid Profiles of Thermobifida fusca
Get PDFThe aerobic, thermophilic Actinobacterium, Thermobifida fusca has been proposed as an organism to be used for the efficient conversion of plant biomass to fatty acid-derived precursors of biofuels or biorenewable chemicals. Despite the potential of T. fusca to catabolize plant biomass, there is remarkably little data available concerning the natural ability of this organism to produce fatty acids. Therefore, we determined the fatty acids that T. fusca produces when it is grown on different carbon sources (i.e., glucose, cellobiose, cellulose and avicel) and at two different growth temperatures, namely at the optimal growth temperature of 50°C and at a suboptimal temperature of 37°C. These analyses establish that T. fusca produces a combination of linear and branched chain fatty acids (BCFAs), including iso-, anteiso-, and 10-methyl BCFAs that range between 14- and 18-carbons in length. Although different carbon sources and growth temperatures both quantitatively and qualitatively affect the fatty acid profiles produced by T. fusca, growth temperature is the greater modifier of these traits. Additionally, genome scanning enabled the identification of many of the fatty acid biosynthetic genes encoded by T. fusca.This article is published as Winkelman DC and Nikolau BJ (2022) The Effects of Carbon Source and Growth Temperature on the Fatty Acid Profiles of Thermobifida fusca. Front. Mol. Biosci. 9:896226. doi: 10.3389/fmolb.2022.896226.© 2022 Winkelman and Nikolau. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms
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