title: Engineering of Corynebacterium glutamicum for high-yield L-valine production under oxygen deprivation conditions.
abstract: We previously demonstrated efficient L-valine production by metabolically engineered Corynebacterium glutamicum under oxygen deprivation. To achieve the high productivity, a NADH/NADPH cofactor imbalance during the synthesis of l-valine was overcome by engineering NAD-preferring mutant acetohydroxy acid isomeroreductase (AHAIR) and using NAD-specific leucine dehydrogenase from Lysinibacillus sphaericus. Lactate as a by-product was largely eliminated by disrupting the lactate dehydrogenase gene ldhA. Nonetheless, a few other by-products, particularly succinate, were still produced and acted to suppress the L-valine yield. Eliminating these by-products therefore was deemed key to improving theL-valine yield. By additionally disrupting the phosphoenolpyruvate carboxylase gene ppc, succinate production was effectively suppressed, but both glucose consumption and L-valine production dropped considerably due to the severely elevated intracellular NADH/NAD(+) ratio. In contrast, this perturbed intracellular redox state was more than compensated for by deletion of three genes associated with NADH-producing acetate synthesis and overexpression of five glycolytic genes, including gapA, encoding NADH-inhibited glyceraldehyde-3-phosphate dehydrogenase. Inserting feedback-resistant mutant acetohydroxy acid synthase and NAD-preferring mutant AHAIR in the chromosome resulted in higher L-valine yield and productivity. Deleting the alanine transaminase gene avtA suppressed alanine production. The resultant strain produced 1,280 mM L-valine at a yield of 88% mol mol of glucose(-1) after 24 h under oxygen deprivation, a vastly improved yield over our previous best.
authors.affiliation: Research Institute of Innovative Technology for the Earth, Kizugawadai, Kizugawa, Kyoto, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Hasegawa
authors.forename: Masako
authors.initials: M
authors.lastname: Suda
authors.forename: Kimio
authors.initials: K
authors.lastname: Uematsu
authors.forename: Yumi
authors.initials: Y
authors.lastname: Natsuma
authors.forename: Kazumi
authors.initials: K
authors.lastname: Hiraga
authors.forename: Toru
authors.initials: T
authors.lastname: Jojima
authors.forename: Masayuki
authors.initials: M
authors.lastname: Inui
authors.forename: Hideaki
authors.initials: H
authors.lastname: Yukawa
mesh_terms: D000693:Anaerobiosis; D053898:Biosynthetic Pathways / Q000235:genetics*; D048230:Corynebacterium glutamicum / Q000235:genetics* / Q000378:metabolism*; D017353:Gene Deletion; D015870:Gene Expression; D019344:Lactic Acid / Q000378:metabolism; D060847:Metabolic Engineering; D009243:NAD / Q000378:metabolism; D010100:Oxygen / Q000378:metabolism; D011995:Recombination, Genetic; D019802:Succinic Acid / Q000378:metabolism; D014633:Valine / Q000096:biosynthesis*
chemical_list: D009243:NAD; D019344:Lactic Acid; D019802:Succinic Acid; D014633:Valine; D010100:Oxygen