l-Valine production with minimization of by-products’ synthesis in Corynebacterium glutamicum and Brevibacterium flavum

Hou, Xiaohu; Chen, Xinde; Zhang, Yue; Qian, He; Zhang, Weiguo

doi:10.1007/s00726-012-1308-9

Cited by 46 publications

(41 citation statements)

References 25 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The intracellular concentration of L-valine determined was as much as 10-fold higher than that of L-glutamate during L-valine production under oxygen deprivation in strain Val-9. Consequently, AvtA-catalyzed alanine synthesis by transamination from L-valine must be the predominant reaction under these conditions (34).…”

Section: Discussionmentioning

confidence: 99%

Engineering of Corynebacterium glutamicum for High-Yieldl-Valine Production under Oxygen Deprivation Conditions

Hasegawa

Suda

Uematsu

et al. 2013

Appl Environ Microbiol

108

View full text Add to dashboard Cite

bWe 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 the L-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.

show abstract

Section: Discussionmentioning

confidence: 99%

Engineering of Corynebacterium glutamicum for High-Yieldl-Valine Production under Oxygen Deprivation Conditions

Hasegawa

Suda

Uematsu

et al. 2013

Appl Environ Microbiol

108

View full text Add to dashboard Cite

show abstract

“…Luria-Bertani (LB) was used as the standard medium for Escherichia coli. LBG (LB supplemented with 5 g L −1 glucose) was used for C. glutamicum [2]. Epo medium was used for preparing electroporation-competent cells.…”

Section: Strains Growth Media and Culturing Conditionsmentioning

confidence: 99%

“…acids [1,2]. Metabolic engineering via genetic modification in C. glutamicum is based on intensifying or shifting carbon flux towards the biosynthetic pathway of the aimed product or on enhancing the degradation of some special substrates [3].…”

mentioning

confidence: 99%

A method for simultaneous gene overexpression and inactivation in the Corynebacterium glutamicum genome

Zhang²,

Han

et al. 2016

Journal of Industrial Microbiology and Biotechnology

Self Cite

View full text Add to dashboard Cite

The gene integration method is an important tool to stably express desirable genes in bacteria. To avoid heavy workload and cost, we constructed a rapid and efficient method for genome modification. This method depended on a mobilizable plasmid, which contains a P tac promoter, an introduced multiple cloning site (iMCS), and rrnBT1T2 terminator. Briefly, the mobilizable plasmid pK18-MBPMT with the P tac-iMCS-rrnBT1T2 cartridge derived from pK18mobsacB was prepared to directly integrate hetero-/homologous DNA into the Corynebacterium glutamicum genome. Like our previous method, this method was based on insertional inactivation and double-crossover homologous recombination, which simultaneously achieved gene overexpression and inactivation in the genome without the use of genetic markers. Compared to the previous method, this protocol omitted the construction of a recombinant expression plasmid and clone of the target gene(s) cassette, which significantly decreased the workload, cost, and operational time. Using this method, the heterologous gene amy and the homologous gene lysC (T311I) were successfully integrated into the C. glutamicum genome at alaT and avtA loci, respectively. Moreover, the operation time of this method was shorter than that of the previous method, especially for repeated integration. This method, which is based on the mobilizable plasmid pK18-MBPMT, thus represents a potentially attractive protocol for the integration of genes in the course of genetic modification of C. glutamicum.

show abstract

“…Biomass was determined by measuring OD 562 with UV-1800 spectrophotometer (Shimadzu, Tokyo, Japan). The dry cell weight (DCW) per liter was calculated according to the formula: DCW (g/L) = 0.36 × OD 562 [24] …”

Section: Fermentation Ofmentioning

confidence: 99%

Enhancing pentose phosphate pathway in Corynebacterium glutamicum to improve l‐isoleucine production

Wang

et al. 2016

Biotech and App Biochem

View full text Add to dashboard Cite

Three genes, gnd, pgl, and fbp, relevant to the pentose phosphate pathway (PPP) were overexpressed in Corynebacterium glutamicum IWJ001, leading to increase of l-isoleucine production. The transcriptional levels of gnd, pgl, and fbp significantly increased in IWJ001/pDXW-8-gnd-fbp-pgl. Compared with the control strain IWJ001/pDXW-8, intracellular NADPH/NADP ratios in IWJ001/pDXW-8-gnd and IWJ001/pDXW-8-gnd-fbp cells grown for 36 H increased threefold and fourfold, respectively, indicating that overexpression of gnd and fbp redirected the carbon flux to PPP. Intracellular NADPH/NADP ratio in IWJ001/pDXW-8-gnd-fbp-pgl grown for 36 H was similar to IWJ001/pDXW-8, suggesting that the NADPH produced by PPP could be quickly consumed for l-isoleucine production. 10.9 and 28.96 g/L of l-isoleucine was produced in IWJ001/pDXW-8-gnd-fbp-pgl in shake flask cultivation and fed-batch fermentation, respectively. In addition, IWJ001/pDXW-8-gnd-fbp-pgl grew fast, its dry cell weight reached 49 g/L after 48 H, whereas the start strain IWJ001/pDXW-8 reached only 40 g/L. After 96 H fermentation, l-isoleucine yield on glucose in IWJ001/pDXW-8-gnd-fbp-pgl reached 0.138 g/g. The results demonstrate that carbon flux redirection to PPP is an efficient approach to enhance l-isoleucine production in C. glutamicum.

show abstract

l-Valine production with minimization of by-products’ synthesis in Corynebacterium glutamicum and Brevibacterium flavum

Cited by 46 publications

References 25 publications

Engineering of Corynebacterium glutamicum for High-Yieldl-Valine Production under Oxygen Deprivation Conditions

Engineering of Corynebacterium glutamicum for High-Yieldl-Valine Production under Oxygen Deprivation Conditions

A method for simultaneous gene overexpression and inactivation in the Corynebacterium glutamicum genome

Enhancing pentose phosphate pathway in Corynebacterium glutamicum to improve l‐isoleucine production

Contact Info

Product

Resources

About