Relevance of NADH Dehydrogenase and Alternative Two-Enzyme Systems for Growth of Corynebacterium glutamicum With Glucose, Lactate, and Acetate

Abstract

The oxidation of NADH with the concomitant reduction of a quinone is a crucial step in the metabolism of respiring cells. In this study, we analyzed the relevance of three different NADH oxidation systems in the actinobacterial model organism Corynebacterium glutamicum by characterizing defined mutants lacking the non-proton-pumping NADH dehydrogenase Ndh (Δndh) and/or one of the alternative NADH-oxidizing enzymes, L-lactate dehydrogenase LdhA (ΔldhA) and malate dehydrogenase Mdh (Δmdh). Together with the menaquinone-dependent L-lactate dehydrogenase LldD and malate:quinone oxidoreductase Mqo, the LdhA-LldD and Mdh-Mqo couples can functionally replace Ndh activity. In glucose minimal medium the Δndh mutant, but not the ΔldhA and Δmdh strains, showed reduced growth and a lowered NAD+/NADH ratio, in line with Ndh being the major enzyme for NADH oxidation. Growth of the double mutants ΔndhΔmdh and ΔndhΔldhA, but not of strain ΔmdhΔldhA, in glucose medium was stronger impaired than that of the Δndh mutant, supporting an active role of the alternative Mdh-Mqo and LdhA-LldD systems in NADH oxidation and menaquinone reduction. In L-lactate minimal medium the Δndh mutant grew better than the wild type, probably due to a higher activity of the menaquinone-dependent L-lactate dehydrogenase LldD. The ΔndhΔmdh mutant failed to grow in L-lactate medium and acetate medium. Growth with L-lactate could be restored by additional deletion of sugR, suggesting that ldhA repression by the transcriptional regulator SugR prevented growth on L-lactate medium. Attempts to construct a ΔndhΔmdhΔldhA triple mutant were not successful, suggesting that Ndh, Mdh and LdhA cannot be replaced by other NADH-oxidizing enzymes in C. glutamicum.

Transcriptome profiles of high-lysine adaptation reveal insights into osmotic stress response in Corynebacterium glutamicum

“…RNA-seq analysis from the ST and HL samples revealed that the high-lysine stresses did not affect the expression levels of ndh genes in C. glutamicum . Further physiological experiments showed that deletion of ndh gene did not cause serious effects on cell growth under the tested no-stress condition, which was consistent with previous reports ( Nantapong et al, 2004 ; Maeda et al, 2021 ) ( Figure 6B ). However, under high-lysine stress conditions, the growth curve analysis suggested that the ∆ndh mutant showed clear growth defects as compared with wild-type and complemented strains.…”

A Hybrid Process for Valorization of d-Fructose to 2,5-Bis(hydroxymethyl)furan by Bridging Chemocatalysis and Biocatalysis in a Betaine:Benzenesulfonic Acid System

“…54 The selection of an appropriate biocatalyst and strategy can be employed for efficient bioreduction. 55,56 Recombinant E. coli DCF harboring CgCR and FDH was constructed and chosen for whole-cell biocatalysis of 5-HMF using a sodium formate (HCOONa) cosubstrate. HCOONa is nearly neutral in a water system, which might provide a biocompatible environment for E. coli DCF.…”

Review on Microreactors for Photo-Electrocatalysis Artificial Photosynthesis Regeneration of Coenzymes

“…It is widely distributed in nature and participates in catalyzing various types of reactions to prepare a variety of chiral pharmaceutical intermediates, which promote the progress and development of the pharmaceutical industry [ 18 , 19 , 20 ]. At the same time, oxidoreductase also participates in the key steps of biological cell metabolism and energy conversion [ 21 , 22 ], and has considerable research potential in the preparation of sor and environmental treatment fields [ 23 , 24 , 25 ]. However, in the application of oxidoreductase, a coenzyme such as NADH is often needed as a hydride donor to promote the reaction towards catalysis [ 26 ].…”