Impact of overexpressing NADH kinase on glucose and xylose metabolism in recombinant xylose-utilizing Saccharomyces cerevisiae.
During growth of Saccharomyces cerevisiae on glucose, the redox cofactors NADH and NADPH are predominantly involved in catabolism and biosynthesis, respectively. A deviation from the optimal level of these cofactors often results in major changes in the substrate uptake and biomass formation. However, the metabolism of xylose by recombinant S. cerevisiae carrying xylose reductase and xylitol dehydrogenase from the fungal pathway requires both NADH and NADPH and creates cofactor imbalance during growth on xylose. As one possible solution to overcoming this imbalance, the effect of overexpressing the native NADH kinase (encoded by the POS5 gene) in xylose-consuming recombinant S. cerevisiae directed either into the cytosol or to the mitochondria was evaluated. The physiology of the NADH kinase containing strains was also evaluated during growth on glucose. Overexpressing NADH kinase in the cytosol redirected carbon flow from CO(2) to ethanol during aerobic growth on glucose and to ethanol and acetate during anaerobic growth on glucose. However, cytosolic NADH kinase has an opposite effect during anaerobic metabolism of xylose consumption by channeling carbon flow from ethanol to xylitol. In contrast, overexpressing NADH kinase in the mitochondria did not affect the physiology to a large extent. Overall, although NADH kinase did not increase the rate of xylose consumption, we believe that it can provide an important source of NADPH in yeast, which can be useful for metabolic engineering strategies where the redox fluxes are manipulated.
Hou
Jin
J
Center for Microbial Biotechnology, Technical University of Denmark, Building 223, 2800 Kgs., Lyngby, Denmark.
Vemuri
Goutham N
GN
Bao
Xiaoming
X
Olsson
Lisbeth
L
D000332:Aerobiosis; D000449:Aldehyde Reductase / Q000378:metabolism; D000693:Anaerobiosis; D003600:Cytosol / Q000201:enzymology; D050542:D-Xylulose Reductase / Q000378:metabolism; D000431:Ethanol / Q000378:metabolism; D005947:Glucose / Q000378:metabolism*; D007218:Industrial Microbiology / Q000379:methods; D008928:Mitochondria / Q000201:enzymology; D024101:Mitochondrial Proteins; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D054730:Protein Interaction Domains and Motifs; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D014993:Xylitol / Q000378:metabolism; D014994:Xylose / Q000378:metabolism*
D024101:Mitochondrial Proteins; D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D009243:NAD; D000431:Ethanol; D009249:NADP; D014994:Xylose; D000449:Aldehyde Reductase; D050542:D-Xylulose Reductase; D017853:Phosphotransferases (Alcohol Group Acceptor); C060670:POS5 protein, S cerevisiae; D005947:Glucose; D014993:Xylitol