Isolation of an Ustilago maydis ERG11 gene and its expression in a mutant deficient in sterol 14 alpha-demethylase activity

A gene (ERG11) encoding cytochrome P450 sterol 14 alpha-demethylase (P450(14DM)) was isolated from the maize pathogen, Ustilago maydis, by amplifying part of the coding region of the gene using PCR and by employing the amplified DNA fragment as a hybridization probe to recover the complete gene from an U. maydis lambda EMBL3 genomic library. The deduced amino acid sequence of the U. maydis gene showed homology to P450(14DM)s from other organisms and contained specific motifs which were hallmarks of P450s. Expression of the gene in an U. maydis mutant (A20) deficient in P450(14DM) led to only a partial restoration of P450(14DM) activity. Accumulation of ergosta-7,22-dienol and ergost-7-enol in A20 transformants containing the ERG11 gene implied that an additional mutation affecting sterol Delta(5,6)-desaturase activity accompanied the P450(14DM) lesion.