CYTOCHROME-C PEROXIDASE FROM A METHYLOTROPHIC YEAST - PHYSIOLOGICAL-ROLE AND ISOLATION

Mutant strains of the methylotrophic yeast Hansenula polymorpha defect ive in catalase (car) and in glucose repression of alcohol oxidase syn thesis (gcr1) have been isolated following multiple UV mutagenesis ste ps. One representative gcr1 cat mutant C-105 grows during batch cultiv ation in a glucose/methanol medium. However, growth is preceded by a p rolonged lag period. C-105 and other gcr1 cat mutants do not grow on m ethanol medium without an alternative carbon source, A large collectio n of second-site suppressor catalase-defective (scd) revertants were i solated with restored ability for methylotrophic growth (Mth(+)) in th e absence of catalase activity. These Mth(+) Ro I car scd strains util ize methanol as a sole source of carbon and energy, although biomass y ields are reduced relative to the wildtype strain. In contrast to the parental C-105 strain, H2O2 does not accumulate in the methanol medium of the revertants. We show that restoration of methylotrophic growth in the suppressor strains is strongly correlated with increased levels of the alternative H2O2-destroying enzyme, cytochrome c peroxidase, C ytochrome c peroxidase from cell-free extracts of one of the scn rever tants has been purified to homogeneity and crystallized.