Abstract
There are three kinds of aspartate kinase in the metabolic pathway of Escherichia coli,including LysC,MetL and ThrA. Lysine,methionine and threonine are synthesized after aspartic acid phosphorylation pathway caused by aspartate kinase so that aspartic acid is not able to be accumulated to a high concentration in E. coli. Aspartic acid phosphorylation could partly inhibit by gene knock-out. Single gene mutants which lack LysC,ThrA and MetL respectively,are constructed from W3110. After chloroamphenicol resistance removed by pCP20,double gene mutants which lack LysC-ThrA and LysC-MetL respectively are constructed. All mutants are finally confirmed by check primer. These constructions are based on Red recombination system. Concentration of aspartic acid is determined by high performance liquid chromatography. The results show that all mutants except MetL single gene mutants are able to accumulate more L-aspartic acid than wild type. This will lay a foundation for strain improvement by metabolic engineering and contribute to production of aspartic acid by fermentation.
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