Metabolic Engineering Of Corynebacterium Glutamicum ATCC13032to Produce L-methionine

Methionine is one of the essential amino acids for human, and plays an important role infood, medical and feed additive industries. Methionine is commercially produced mainly bychemical synthesis as a racemic mixture, which costs a lot and will lead to environmentalpollution. As a result, L-methionine production by bacterial fermentation is becoming moreattractive. In this study, an L-methionine producing strain was constructed from the wild typeCorynebacterium glutamicum strain ATCC13032, using metabolic engineering strategies.The main results are listed below:(1) Strain WTQ101was constructed from ATCC13032via deleting gene thrB encodinghomoserine kinase. The strain has an L-threonine auxotroph, and produces a large scale ofL-aspartate and L-homoserine along with the unexpected L-lysine.(2) Strain WTQ101/pJYW-4-homm-lysCmwas constructed from strain WTQ101by theoverexpression of gene lysCmencoding feedback resistant aspartate kinase and gene hommencoding feedback resistant homoserine dehydrogenase. The expression of these two genesleads more L-aspartate to L-homoserine, and further increases the precursor of L-methionine.Strain WTQ102was constructed from WTQ101by the deletion of gene mcbR encoding theregulator McbR to release the transcriptional repression to various genes in the L-methioninebiosynthetic pathway. The strain gains less L-lysine and produces more L-methionine. Tofurther increase the L-methionine production, gene lysCmand hommwere introduced intostrain WTQ102, and obtained strain WTQ102/pJYW-4-homm-lysCm.(3) Gene cluster brnF and brnE encoding the export protein complex BrnFE wasoverexpressed to increase the extracellular L-methionine concentration.(4) The fermentation condition was optimized via the addition of L-cysteine andL-isoleucine, and the control of the pH value. The final condition was selected as2mML-isoleucine addition and pH6.5. Strain WTQ102/pJYW-4-homm-lysCm-brnFE was furtherfermented in a3-L fermentor using the optimized condition, and42mM L-methionine wasproduced after72h fermentation.