(A) study on the effect of stringent regulator RelA on the production of amino acids in Escherichia coli = Stringent regulator RelA의 대장균에서의 아미노산 생산에 대한 효과에 관한 연구

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Recent genetics and biochemical studies have identified a novel control mechanism of gene expression during amino acid starvation, the so-called ‘stringent response’. The stringent response is triggered by the elevated concentrations of guanosine tetraphosphate (ppGpp), which is synthesized by the RelA ppGpp synthetase in $\it{Escherichia coli}$. Here, we have applied directed evolution techniques to create novel mutant $\it{relA}$ genes with increased ppGpp synthetase activities. Using the constructed mutants, the intracellular concentration of ppGpp in $\it{E. coli}$ cells were increased without amino acid starvation condition. In addition, dramatic changes in gene expression profiles were observed. One hand, the expression of genes involved in translation system was inhibited; on the other, enzymes required for amino acid biosynthesis and transport were activated. And the introduction of the mutant $\it{relA}$ gene into the amino acid producing strains was resulted in increased production of amino acids such as L-threonine, L-lysine, L-phenylalanine, and L-tyrosine. Our results clearly demonstrate that the artificial increase of the intracellular ppGpp by the mutant $\it{relA}$ gene reprogrammed the genome-wide gene expression for the production of amino acids without stringent conditions, and these mutant $\it{relA}$ genes could be applied for the production of amino acids in other bacterial strains. Furthermore, we showed that the metabolic engineering of the minimized-genome E. coli strain constructed by precise elimination of unnecessary genes or genomic fragment could increase the production of amino acids such as L-threonine, L-lysine, L-phenylalanine, and L-tyrosine. In addition, genome-wide transcriptional profile analysis revealed that the elimination of unnecessary genes have resulted in nutrient and energy saving and an improved substrate yield coefficient, causing more efficient cellular metabolism without physiological compromise. And...
Kim, Sun-Changresearcher김선창researcher
한국과학기술원 : 생명과학과,
Issue Date
455351/325007  / 020045214

학위논문(박사) - 한국과학기술원 : 생명과학과, 2010.08, [ vii, 129 p. ]


Eschierichia coli; amino acids; RelA; Metabolic engineering; directed evolution; 단백질 구조; 대장균; RelA 유전자; 아미노산 생산; 대사공항

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