Disruption of the cytochrome c gene in xylose-utilizing yeast Pichia stipitis leads to higher ethanol production.
The xylose-utilizing yeast, Pichia stipitis, has a complex respiratory system that contains cytochrome and non-cytochrome alternative electron transport chains in its mitochondria. To gain primary insights into the alternative respiratory pathway, a cytochrome c gene (PsCYC1, Accession No. AF030426) was cloned from wild-type P. stipitis CBS 6054 by cross-hybridization to CYC1 from Saccharomyces cerevisiae. The 333 bp open reading frame of PsCYC1 showed 74% and 69% identity to ScCYC1 and ScCYC7, respectively, at the DNA level. Disruption of PsCYC1 resulted in a mutant that uses the salicylhydroxamic acid (SHAM)-sensitive respiratory pathway for aerobic energy production. Cytochrome spectra revealed that cytochromes c and a.a(3) both disappeared in the cyc1-Delta mutant, so no electron flow through the cytochrome c oxidase was possible. The cyc1-Delta mutant showed 50% lower growth rates than the parent when grown on fermentable sugars. The cyc1-Delta mutant was also found to be unable to grow on glycerol. Interestingly, the mutant produced 0.46 g/g ethanol from 8% xylose, which was 21% higher in yield than the parental strain (0.38 g/g). These results suggested that the alternative pathway might play an important role in supporting xylose conversion to ethanol under oxygen-limiting conditions.
Shi
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Department of Bacteriology, University of Wisconsin, 1550 Linden Drive, Madison, WI 53706, USA.
Davis
B
B
Sherman
F
F
Cruz
J
J
Jeffries
T W
TW
D000935:Antifungal Agents / Q000737:chemistry / Q000378:metabolism; D001483:Base Sequence; D015139:Blotting, Southern; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics*; D017931:DNA Primers; D004271:DNA, Fungal / Q000737:chemistry; D004579:Electron Transport / Q000235:genetics; D000431:Ethanol / Q000378:metabolism*; D005285:Fermentation; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D010802:Phylogeny; D010843:Pichia / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D012457:Salicylamides / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D014994:Xylose / Q000378:metabolism*
D000935:Antifungal Agents; D003574:Cytochrome c Group; D017931:DNA Primers; D004271:DNA, Fungal; D012457:Salicylamides; D000431:Ethanol; C005703:salicylhydroxamic acid; D014994:Xylose