초록
<P><B>ABSTRACT</B><P> 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 <I>ldhA</I> . 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 the l -valine yield. By additionally disrupting the phosphoenolpyruvate carboxylase gene <I>ppc</I> , succinate production was effectively suppressed, but both glucose consumption and l -valine production dropped considerably due to the severely elevated intracellular NADH/NAD <SUP>+</SUP> 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 <I>gapA</I> , 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 <I>avtA</I> suppressed alanine production. The resultant strain produced 1,280 mM l -valine at a yield of 88% mol mol of glucose <SUP>−1</SUP> after 24 h under oxygen deprivation, a vastly improved yield over our previous best. </P></P>