Improvement of l-Leucine Production in Corynebacterium glutamicum by Altering the Redox Flux

The production of l-leucine was improved by the disruption of ltbR encoding transcriptional regulator and overexpression of the key genes (leuAilvBNCE) of the l-leucine biosynthesis pathway in Corynebacterium glutamicum XQ-9. In order to improve l-leucine production, we rationally engineered C. glutamicum to enhance l-leucine production, by improving the redox flux. On the basis of this, we manipulated the redox state of the cells by mutating the coenzyme-binding domains of acetohydroxyacid isomeroreductase encoded by ilvC, inserting NAD-specific leucine dehydrogenase, encoded by leuDH from Lysinibacillus sphaericus, and glutamate dehydrogenase encoded by rocG from Bacillus subtilis, instead of endogenous branched-chain amino acid transaminase and glutamate dehydrogenase, respectively. The yield of l-leucine reached 22.62 ± 0.17 g·L(−1) by strain ΔLtbR-acetohydroxyacid isomeroreductase (AHAIR)(M)/ABNC(M)E, and the concentrations of the by-products (l-valine and l-alanine) increased, compared to the strain ΔLtbR/ABNCE. Strain ΔLtbR-AHAIR(M)LeuDH/ABNC(M)LDH accumulated 22.87±0.31 g·L(−1) l-leucine, but showed a drastically low l-valine accumulation (from 8.06 ± 0.35 g·L(−1) to 2.72 ± 0.11 g·L(−1)), in comparison to strain ΔLtbR-AHAIR(M)/ABNC(M)E, which indicated that LeuDH has much specificity for l-leucine synthesis but not for l-valine synthesis. Subsequently, the resultant strain ΔLtbR-AHAIR(M)LeuDHRocG/ABNC(M)LDH accumulated 23.31 ± 0.24 g·L(−1) l-leucine with a glucose conversion efficiency of 0.191 g·g(−1).