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

Abstract

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 14K K-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.z 1999 Federation of European Biochemical Societies.

Association of Multiple Developmental Defects and Embryonic Lethality with Loss of Microsomal NADPH-Cytochrome P450 Oxidoreductase

“…These results confirm the critical role of CYPOR in embryonic development and are consistent with the report that RNA-mediated interference of CYPOR expression in the nematode Caenorhabditis elegans produces embryonic lethality and undefined structural abnormalities (26). Although yeast are able to survive in the absence of CYPOR, with cytochrome b 5 serving as an alternate electron donor to the cytochromes P450 (9,10), it is clear that the functions of CYPOR are essential in higher organisms.…”

“…Electron transfer to the cytochromes P450 as well as membrane anchoring requires the hydrophobic, N-terminal membrane-binding domain of this microsomal flavoprotein (6,7). No other physiological electron donor to the cytochromes P450 has been identified in vertebrates, although plants contain multiple CYPOR genes (8) and electron transfer from the cytochrome b 5 /cytochrome b 5 system to P450 has been reported in yeast lacking CYPOR (9,10). Development of the mammalian embryo requires temporally and spatially regulated biosynthesis and degradation of signaling factors, many of which, such as retinoic acid, sterols, prostaglandins, and steroids, are dependent upon cytochrome P450 or other CYPOR-dependent pathways.…”

The P450–1 gene of Gibberella fujikuroi encodes a multifunctional enzyme in gibberellin biosynthesis

“…Sequential oxidations at a single carbon atom that are mediated by cytochrome P450s have been described as, for example, in the loss of C-14 in steroid biosynthesis (30) and the three-step oxidation of ent-kaurene to ent-kaurenoic acid in G. fujikuroi (8). However, the reactions catalyzed by P450-1 involve oxidations The mass spectrometric data are provided as Table 2, which is published as supplemental data on the PNAS web site, www.pnas.org.…”

Role of the non‐respiratory pathways in the utilization of molecular oxygen by Saccharomyces cerevisiae

“…Enzymes: ERG1, squalene epoxidase; ERG7, lanosterol synthase; ERG11 (CYP51), lanosterol C-14 demethylase; ERG24, sterol C-14 reductase; ERG25, sterol C-4 methyloxydase; ERG26, sterol C-3 dehydrogenase (C4-decarboxylase); ERG27, sterol C-3 ketoreductase; ERG6, sterol C-24 methyltransferase; ERG2, sterol C-8 isomerase; ERG3, sterol C-5 desaturase; ERG5, sterol C-22 desaturase; ERG4, sterol C-24 reductase; ACAT (ARE1,2), acyl-CoA sterol acyl transferases. 113 However, by using various physiological modulations of the cellular NADH level during transitions from anaerobic (classical laboratory and enological) to aerobic conditions, it has been shown that the oxygen uptake due to sterol synthesis (i.e. terbinafine-and fenpropimorph-sensitive O 2 consumption) does not correlate with the cellular NADH redox state 168 (Rosenfeld et al, unpublished data).…”