Transmembrane Architecture of a Full-Length Monospanning Cytochrome P450

Abstract

Lanosterol demethylase or Cyp51 catalyzes the first committed post-cyclization step in cholesterol or ergosterol biosynthesis, making it a key clinical antifungal target. Cyp51 is a member of the cytochrome p450 superfamily of monoxygenases that catalyze key reactions in lipid biosynthesis and xenobiotic detoxification. Due to substrate hydrophobicity, many cytochrome p450s catalyze difficult surface reactions near the lipid bilayer/soluble milieu interface. Despite ample structural data, details of how cytochrome p450s orient into the membrane to perform this process are not well understood. Here, we crystallized the bitopic, membrane monospanning Saccharomyces cerevisiae cyp51 with its membrane spanning integral membrane domain. This structure identifies the transmembrane architecture of a membrane spanning cytochrome p450 for the first time, identifying the likely orientation into the lipid bilayer, and is the first structure of an enzyme with a single transmembrane domain intact. This orientation reveals a possible substrate entry channel, and a second unanticipated channel, the positioning of a key antifungal inhibitor, and numerous likely interaction points with the lipid bilayer. These findings have implications for the orientations of the 50% of integral membrane proteins that are single transmembrane spanning, for cytochrome p450 catalysis in general, and for the development of novel antifungal therapeutics.