Disruption of the cytochrome cgene in xylose‐utilizing yeast Pichia stipitisleads 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 cgene (PsCYC1, Accession No. AF030426) was cloned from wild‐type P. stipitisCBS 6054 by cross‐hybridization to CYC1from Saccharomyces cerevisiae. The 333 bp open reading frame of PsCYC1showed 74% and 69% identity to ScCYC1and ScCYC7, respectively, at the DNA level. Disruption of PsCYC1resulted in a mutant that uses the salicylhydroxamic acid (SHAM)‐sensitive respiratory pathway for aerobic energy production. Cytochrome spectra revealed that cytochromes cand a·a3both disappeared in the cyc1‐Δ mutant, so no electron flow through the cytochrome coxidase was possible. The cyc1‐Δ mutant showed 50% lower growth rates than the parent when grown on fermentable sugars. The cyc1‐Δ 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. Copyright © 1999 John Wiley & Sons, Ltd.