Browsing by Subject "Lysine"
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Item Characterization of the diverse substrate specificities and biological roles of polyamine biosynthetic(2008-09-18) Lee, Jeongmi; Phillips, Margaret A.The beta /alpha -barrel fold-type basic amino acid decarboxylases include eukaryotic ornithine decarboxylases (ODC), and bacterial and plant enzymes with activity on L-arginine and meso-diaminopimelate. These enzymes catalyze essential steps in polyamine and lysine biosynthesis. Phylogenetic analysis suggests that diverse bacterial species also contain ODC-like enzymes from this fold-type. However, in comparison to the eukaryotic ODCs, amino acid differences were identified in the sequence of the 310-helix that forms a key specificity element in the active site, suggesting they might function on novel substrates. Putative decarboxylases from a phylogenetically diverse range of bacteria were characterized to determine their substrate preference. Enzymes from species within Methanosarcina, Pseudomonas, Bartonella, Nitrosomonas, Thermotoga and Aquifex showed a strong preference for L-ornithine, while the enzyme from Vibrio vulnificus (VvL/ODC) had dual specificity functioning well on both L-ornithine and L-lysine. The X-ray structure of VvL/ODC was solved in the presence of the reaction products putrescine and cadaverine to 1.7 and 2.15 Å, respectively. The overall structure is similar to eukaryotic ODC, however reorientation of the 310-helix enlarges the substrate binding-pocket, thereby allowing L-lysine to be accommodated. The structure of the putrescine-bound enzyme suggests that a bridging water molecule between the shorter L-ornithine and key active site residues provides the structural basis for VvL/ODC to also function on this substrate. Our data demonstrate that there is greater structural and functional diversity in bacterial polyamine biosynthetic decarboxylases than previously suspected. The beta /alpha -barrel fold decarboxylases also include a group of bacterial putative carboxynorspermidine decarboxylases (CANSDCs), which were hypothesized to be involved in norspermidine biosynthesis. Putative norspermidine biosynthetic enzymes including V.vulnificus CANSDC were characterized to determine their substrate specificities. The polyamine biosynthetic pathway in V.cholerae was confirmed by in vivo reconstitution of norspermidine biosynthetic genes in E.coli. Knockout of polyamine biosynthetic genes in V.cholerae revealed that normal levels of norspermidine are not essential for cell viability and that norspermidine is important for biofilm formation as a signaling molecule in more than one regulatory pathways for this process. Our preliminary data from real time RT-PCR suggest that norspermidine may also be related to quorum sensing and virulence gene expressions in V.cholerae .Item Evolution, metabolism, and virulence of the oral microbiome(2013-12) Jorth, Peter Allan; Whiteley, MarvinThe human microbiome has important roles in maintaining health, but dysbiosis of the microbiota can lead to disease. Polymicrobial interactions can result in synergy, producing disease that is worse than the sum of the respective single species infections. Despite this significant impact, synergy is understudied due to the complexity of polymicrobial interactions. Periodontitis is a microbiome-associated disease, and is one of the most common infectious diseases worldwide. Therefore, we have used periodontal disease as a model to study polymicrobial synergy. I have used two complementary approaches to study polymicrobial infections. The opportunistic periodontal pathogen Aggregatibacter actinomycetemcomitans exhibits synergy with streptococci in model murine infections. Because polymicrobial interactions are dependent on organisms’ abilities to sense their environments, I have examined the genetic regulatory mechanisms used by A. actinomycetemcomitans to interact with its environment. Through Northern blot analyses and biochemical approaches, I show that A. actinomycetemcomitans uses non-coding RNAs to regulate amino acid transport. Taking a comparative genomics approach, I demonstrate that A. actinomycetemcomitans DNA uptake systems are evolutionarily linked to genome defense. To describe host-influenced changes in gene expression, I develop a new technique to transcriptionally profile A. actinomycetemcomitans in a murine abscess infection, thereby revealing the importance of specific fermentative and anaerobic respiratory genes for in vivo survival. The long-term goal is to use these studies as a basis to characterize genetic regulatory mechanisms mediating synergy in polymicrobial A. actinomycetemcomitans infections with streptococci and other oral microbes. As a second approach to study polymicrobial infections, I analyze gene expression of healthy and diseased human plaque communities from aggressive periodontitis patients. Profiling ribosome content of healthy and diseased communities, I show that disease communities adopt similar less diverse population structures distinct from healthy populations. In addition to changes in population composition, using community transcriptional profiling I show that a keystone species within diseased communities up-regulates expression of genes involved in making the oral inflammatory molecule butyrate. These studies demonstrate for the first time that microbiome based diseases are marked by gene expression changes in addition to compositional changes.Item Genetic Combining Analysis of Food-Grade Maize: Colored and Quality Protein(2012-10-19) Mahan, Adam LyleMaize genetic diversity includes an array of kernel colors (red, blue, purple) with blue concentrated in the aleurone and red primarily in the pericarp. Quality protein maize (QPM) is improved over normal maize in regards to grain concentration of the essential amino acids lysine and tryptophan but has not been widely adapted in part due to lower than conventional yields. These are minimally-utilized specialty corns when compared to the yellows and whites commonly grown. Red, blue, and purple pigments are antioxidant phytochemicals produced by the plant as secondary metabolites. Antioxidants have been linked to anti-cancer and other anti-inflammatory health benefits. QPM hybrids are desirable in developing countries where subsistent agriculture is commonly practiced and quality protein cereals are non-existent. These two diverse maize categories have been the subject of little breeding research compared to normal maize and the potential for high phenolic content as well as the characterization of these QPM hybrids has not been previously investigated. We evaluated 153 maize hybrids (84 colored, 69 QPM) across three locations. High heritability estimates were found for phenolic content (0.80), tryptophan (0.46), and endosperm opacity (0.82). It was encouraging that all three traits observed little genotype by environment (GxE) interaction across diverse environments. This proved the trait analysis procedure to be robust in detecting and separating genotypes for both total phenolic content in colored maize, and amino acids in QPM. Top combiners for phenolics were the purple maize "maize morado" and red lines, with blue, yellow and white maize performing in descending order. Within the tested hybrids, high per kernel antioxidants (measured by total phenolics) may be the answer for producing the most total phenolics, with the top hybrid yielding greater than twice the total phenolics as the top yielding yellow hybrid. The top QPM hybrid out yielded the top normal hybrid by 35 and 30% for lysine and tryptophan. Additionally, QPM endosperm opacity primarily followed an additive, mid-parent trend, with some hybrids (20%) from diverse germplasm backgrounds deviating from that trend displaying the complexity and recessive nature of multiple modifier loci. Additional agronomic and composition traits were minimally correlated with phenolics.Item Lysine supplementation of Iranian bread(Texas Tech University, 1973-08) Dadbakhsh, PariNot availableItem Studies on R. rubrum hemoprotein and interaction of cytochromes with R. rubrum reaction centers(Texas Tech University, 1987-05) Ayres, Mary LynnNot available