Nigel J. Manning

Resistance to fluconazole and cross‐resistance to amphotericin B in Candida albicans from AIDS patients caused by defective sterol Δ5,6‐desaturation

Fluconazole resistance occurs in >10% of cases of candidosis during the late stages of AIDS. We show here in two clinical isolates that resistance was caused by defective sterol Δ5,6‐desaturation. This altered the type of sterol accumulating under fluconazole treatment from 14α‐methylergosta‐8,24(28)‐dien‐3β,6α‐diol to 14α‐methylfecosterol which is capable of supporting growth. A consequence of this mechanism of azole resistance is that an absence of ergosterol causes cross‐resistance to the other major antifungal agent available, amphotericin B. The results also show that growth arrest after fluconazole treatment of C. albicans in clinical conditions is caused by 14α‐methylergosta‐8,24(28)‐dien‐3β,6α‐diol accumulation.

Biodiversity of the P450 catalytic cycle: yeast cytochrome b5/NADH cytochrome b5 reductase complex efficiently drives the entire sterol 14-demethylation (CYP51) reaction.

The widely accepted catalytic cycle of cytochromes P450 (CYP) involves the electron transfer from NADPH cytochrome P450 reductase (CPR), with a potential for second electron donation from the microsomal cytochrome b 5/NADH cytochrome b 5 reductase system. The latter system only supported CYP reactions inefficiently. Using purified proteins including Candida albicans CYP51 and yeast NADPH cytochrome P450 reductase, cytochrome b 5 and NADH cytochrome b 5 reductase, we show here that fungal CYP51 mediated sterol 14α‐demethylation can be wholly and efficiently supported by the cytochrome b 5/NADH cytochrome b 5 reductase electron transport system. This alternative catalytic cycle, where both the first and second electrons were donated via the NADH cytochrome b 5 electron transport system, can account for the continued ergosterol production seen in yeast strains containing a disruption of the gene encoding CPR.