Analysing overexpression of l-valine biosynthesis genes in pyruvate-dehydrogenase-deficient Corynebacterium glutamicum

Abstract

L-valine biosynthesis was analysed by comparing different plasmids in pyruvate-dehydrogenase-deficient Corynebacterium glutamicum strains in order to achieve an optimal production strain. The plasmids contained different combinations of the genes ilvBNCDE encoding for the L-valine forming pathway. It was shown that overexpression of the ilvBN genes encoding acetolactate synthase is obligatory for efficient pyruvate conversion and to prevent L-alanine as a by-product. In contrast to earlier studies, overexpression of ilvE encoding transaminase B is favourable in pyruvate-dehydrogenase-negative strains. Its amplification enhanced L-valine formation and avoided extra- and intracellular accumulation of ketoisovalerate.

Metabolic Engineering of Corynebacterium glutamicum for 2-Ketoisovalerate Production

“…50%) and relatively low final titers of 2-ketoisovalerate (55 mM) and L-valine (22 mM). The same strain overexpressing ilvE (instead of ilvD) formed only 10 mM 2-ketoisovalerate (Y P/S of 0.052 mol per mol of glucose) and 104 mM L-valine (3). In combination, these results indicate the importance of the transaminase B for L-valine formation and point to the contribution of the ilvE deletion to the production of 2-ketoisovalerate.…”

“…In fact, overexpression of ilvBNCD and, in particular, the deletion of pqo shifted the carbon flux significantly in the direction of 2-ketoisovalerate, leading to higher final titers of this product and only low concentrations of the by-products pyruvate, Lalanine, and L-valine. The overexpression of ilvBNC or ilvBNCD in C. glutamicum ⌬aceE has previously been shown to be necessary for a more efficient carbon flux from pyruvate toward L-valine (3,6,40). However, even the pJC4-based overexpression of ilvBNCD might not necessarily completely overcome a limitation in this pathway since expression of ilvBNC is partially controlled by an attenuation mechanism involving antitermination (34).…”

Improvement of l-valine production at high temperature in Brevibacterium flavum by overexpressing ilvEBNrC genes

“…Bartek et al [1] and Blombach et al [3] demonstrated that overexpression of the ilvBNC operon with the ilvE gene afforded higher L-valine production than overexpression of the ilvBNC operon with the ilvD gene. To study the separate contributions of ilvB, ilvN r , ilvC, and livE genes to the production of L-valine, the stains of B. flavum ATCC14067/ pDXW-8-ilvBN r , B. flavum ATCC14067/pDXW-8-ilvBN r C, B. flavum ATCC14067/pDXW-8-ilvEBNC, B. flavum ATCC 14067/pDXW-8-ilvEBN r C, and B. flavum ATCC14067/ pDXW-8-ilvE were constructed, fermentations with the resulting strains were carried out at 31, 34, 37, and 40°C with the fermentation with B. flavum ATCC14067 as control experiment.…”

“…TA catalyzes the conversion of 2-oxo-isovalerate into L-valine with L-glutamate as the amino group donor [15]. Overexpression of the ilvBNC operon with the ilvE gene afforded higher L-valine production than overexpression of the ilvBNC operon the with ilvD gene [1,3]. AHAS is feedback-inhibited by three branched-chain amino acids; the 50% inhibitory concentrations for AHAS with valine, isoleucine, and leucine have been determined (0.9, 3.1, and 6 mM, respectively) [14].…”

“…The ilvB gene encoding a catalytic subunit and the ilvN gene encoding a regulatory subunit form an operon with the ilvC gene [11], and the ilvBNC operon is a target for overexpression for L-valine accumulation [1,2,4,21]. TA catalyzes the conversion of 2-oxo-isovalerate into L-valine with L-glutamate as the amino group donor [15].…”