Cloning and sequencing of the Candida albicans C-4 sterol methyl oxidase gene (ERG25) and expression of an ERG25 conditional lethal mutation in Saccharomyces cerevisiae

The ERG25 gene encoding the Candida albicans C-4 sterol methyl oxidase was cloned and sequenced by complementing a Saccharomyces cerevisiae erg25 muta nt with a C. albicans genomic library. The Erg25p is comprised of 308 amino acids and shows 65 and 38% homology to the enzymes from S. cerevisiae and Homo sapiens, respectively. The protein contains three histidine clusters c ommon to nonheme iron-binding enzymes and an endoplasmic reticulum retrieva l signal as do the proteins from S. cerevisiae and humans. A temperature-se nsitive (ts) conditional lethal mutation of the C. albicans ERG25 was isola ted and expressed in S. cerevisiae. Sequence analysis of the ts mutant indi cated an amino acid substitution within the region of the protein encompass ed by the histidine clusters involved in iron binding. Results indicate tha t plasmid-borne conditional lethal mutants of target genes have potential u se in the rescue of Candida mutations in genes that are essential for viabi lity.