title: Development of L-tryptophan production strains by defined genetic modification in Escherichia coli.
abstract: Construction and improvement of industrial strains play a central role in the commercial development of microbial fermentation processes. L-tryptophan producers have usually been developed by classical random mutagenesis due to its complicated metabolic network and regulatory mechanism. However, in the present study, an L-tryptophan overproducing Escherichia coli strain was developed by defined genetic modification methodology. Feedback inhibitions of 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase (AroF) and anthranilate synthase (TrpED) were eliminated by site-directed mutagenesis. Expression of deregulated AroF and TrpED was achieved by using a temperature-inducible expression plasmid pSV. Transcriptional regulation of trp repressor was removed by deleting trpR. Pathway for L-Trp degradation was removed by deleting tnaA. L-phenylalanine and L-tyrosine biosynthesis pathways that compete with L-tryptophan biosynthesis were blocked by deleting their critical genes (pheA and tyrA). The final engineered E. coli can produce 13.3 g/l of L-tryptophan. Fermentation characteristics of the engineered strains were also analyzed.
authors.affiliation: State Key Laboratory of Food Science and Technology, Jiangnan University, Wuxi 214122, China.
authors.forename: Zhi-Jun
authors.initials: ZJ
authors.lastname: Zhao
authors.forename: Chun
authors.initials: C
authors.lastname: Zou
authors.forename: Yi-Xing
authors.initials: YX
authors.lastname: Zhu
authors.forename: Jun
authors.initials: J
authors.lastname: Dai
authors.forename: Sheng
authors.initials: S
authors.lastname: Chen
authors.forename: Dan
authors.initials: D
authors.lastname: Wu
authors.forename: Jing
authors.initials: J
authors.lastname: Wu
authors.forename: Jian
authors.initials: J
authors.lastname: Chen
mesh_terms: D000878:Anthranilate Synthase / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics; D025461:Feedback, Physiological; D055786:Gene Knockout Techniques; D009097:Multienzyme Complexes / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D010649:Phenylalanine / Q000096:biosynthesis; D010957:Plasmids; D011302:Prephenate Dehydratase / Q000235:genetics; D012097:Repressor Proteins / Q000378:metabolism; D014364:Tryptophan / Q000096:biosynthesis*; D014368:Tryptophanase / Q000235:genetics; D014443:Tyrosine / Q000096:biosynthesis
chemical_list: D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D009097:Multienzyme Complexes; C071216:P-protein, E coli; D012097:Repressor Proteins; C027190:TRPR protein, E coli; D014443:Tyrosine; D010649:Phenylalanine; D014364:Tryptophan; D000878:Anthranilate Synthase; D014368:Tryptophanase; D011302:Prephenate Dehydratase