Corynebacterium glutamicum tailored for high-yield L-valine production

Abstract

We recently engineered the wild type of Corynebacterium glutamicum for the growth-decoupled production of L: -valine from glucose by inactivation of the pyruvate dehydrogenase complex and additional overexpression of the ilvBNCE genes, encoding the L-valine biosynthetic enzymes acetohydroxyacid synthase, isomeroreductase, and transaminase B. Based on the first generation of pyruvate-dehydrogenase-complex-deficient C. glutamicum strains, a second generation of high-yield L-valine producers was constructed by successive deletion of the genes encoding pyruvate:quinone oxidoreductase, phosphoglucose isomerase, and pyruvate carboxylase and overexpression of ilvBNCE. In fed-batch fermentations at high cell densities, the newly constructed strains produced up to 410 mM (48 g/l) L-valine, showed a maximum yield of 0.75 to 0.86 mol/mol (0.49 to 0.56 g/g) of glucose in the production phase and, in contrast to the first generation strains, excreted neither pyruvate nor any other by-product tested.

Platform Engineering of Corynebacterium glutamicum with Reduced Pyruvate Dehydrogenase Complex Activity for Improved Production of l -Lysine, l -Valine, and 2-Ketoisovalerate

“…Compared to the value for C. glutamicum ⌬aceE ⌬pqo (pJC4 ilvBNCE) (27), this titer is 3.4 times higher. Furthermore, C. glutamicum aceE A16 ⌬pqo ⌬ppc (pJC4 ilvBNCE) showed a 43% higher overall Y P/S and a 3-times-higher volumetric productivity (Q P ) ( Table 4).…”

“…Plasmid-bound overexpression of the L-valine biosynthesis genes ilvBNCE shifted the product spectrum toward L-valine (26), and inactivation of the pyruvate:quinone oxidoreductase (PQO) (pqo gene product; Fig. 1) and phosphoglucose isomerase (pgi gene product) in C. glutamicum ⌬aceE (pJC4 ilvBNCE) resulted in even more efficient L-valine production (up to 410 mM, with a maximum yield of 0.86 mol per mol of glucose in the production phase [27]). Based on these results, we engineered the wild type (WT) of C. glutamicum for the aerobic, growth-decoupled production of 2-ketoisovalerate (KIV) from glucose by deletion of the aceE, pqo, and ilvE genes and additional overexpression of the ilvBNCD genes (11).…”

Enhanced Valine Production inCorynebacterium glutamicumwith Defective H+-ATPase and C-Terminal Truncated Acetohydroxyacid Synthase

“…In addition, valine production using a pyruvate dehydrogenase knockout mutant of C. glutamicum was reported recently. 8,9) According to these reports, C. glutamicum with multiple gene-knockouts (pyruvate: quinone oxidoreductase, phosphoglucose isomerase, and pyruvate dehydrogenase gene deletion) and overexpressed ilvBNCE, produced up to 410 mM valine. A combination of the pyruvate dehydrogenase gene knockout and an ATPase-defective mutation might also be effective in improving valine production.…”

“…7) Overexpression of wild-type ilvBNCE has also been reported to be effective in the production of valine. 8,9) Protein engineering studies of AHAS from Escherichia coli has emphasized the application of C-terminal truncations of its regulatory subunit for reducing the feedback inhibition of this enzyme. [10][11][12] Based on these studies, we constructed a feedback-insensitive AHAS gene with C-terminal truncation of the regulatory subunit (ilvN) to investigate valine production in C. glutamicum.…”

Current knowledge on isobutanol production withEscherichia coli,Bacillus subtilisandCorynebacterium glutamicum

“…7 Additional inactivation of the PQO and overexpression of the ilvBNCE genes, encoding AHAS, AHAIR and transaminase B (TA) in C. glutamicum ΔaceE shifted the product spectrum toward L-valine and the resulting strain C. glutamicum ΔaceE Δpqo (pJC4ilvBNCE) produced about 225 mM L-valine with a Y P/S of 0.52 mol L-valine per mol glucose in fed-batch fermentations. 8 Based on these results, we subsequently engineered C. glutamicum for the aerobic production Figure 1. enzymes of the biosynthetic pathway of L-valine and the synthetic pathway from 2-ketoisovalerate to isobutanol.…”