Manipulation of malic enzyme in Saccharomyces cerevisiae for increasing NADPH production capacity aerobically in different cellular compartments.
The yeast Saccharomyces cerevisiae is an attractive cell factory, but in many cases there are constraints related with balancing the formation and consumption of redox cofactors. In this work, we studied the effect of having an additional source of NADPH in the cell. In order to do this, two strains were engineered by overexpression of malic enzyme. In one of them, malic enzyme was overexpressed as its wild-type mitochondrial form, and in the other strain a short form lacking the mitochondrial targeting sequence was overexpressed. The recombinant strains were analyzed in aerobic batch and continuous cultivations, and the basic growth characteristics were generally not affected to a great extent, even though pleiotropic effects of the manipulations could be seen by the altered in vitro activities of selected enzymes of the central metabolism. Moreover, the decreased pentose-phosphate pathway flux and the ratios of redox cofactors showed that a net transhydrogenase effect was obtained, which can be directed to the cytosol or the mitochondria. This may find application in redirecting fluxes for improving specific biotechnological applications.
Moreira dos Santos
Margarida
M
Center for Process Biotechnology, Technical University of Denmark, BioCentrum-DTU, Building 223, DK-2800 Lyngby, Denmark.
Raghevendran
Vijayendran
V
Kötter
Peter
P
Olsson
Lisbeth
L
Nielsen
Jens
J
D000332:Aerobiosis / Q000235:genetics / Q000502:physiology; D015966:Gene Expression Regulation, Fungal / Q000235:genetics* / Q000502:physiology; D005800:Genes, Fungal / Q000235:genetics* / Q000502:physiology; D008291:Malate Dehydrogenase / Q000235:genetics / Q000378:metabolism*; D008928:Mitochondria / Q000201:enzymology* / Q000235:genetics; D009249:NADP / Q000096:biosynthesis* / Q000235:genetics; D010084:Oxidation-Reduction; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*
D029701:Saccharomyces cerevisiae Proteins; D009249:NADP; D008291:Malate Dehydrogenase; C030574:malate dehydrogenase (decarboxylating)