The cytochromes P450 and b5 and their reductases--promising targets for structural studies by advanced solid-state NMR spectroscopy

Abstract

Members of the cytochrome P450 (cyt P450) superfamily of enzymes oxidize a wide array of endogenous and xenobiotic substances to prepare them for excretion. Most of the drugs in use today are metabolized in part by a small set of human cyt P450 isozymes. Consequently, cyt P450s have for a long time received a lot of attention in biochemical and pharmacological research. Cytochrome P450 receives electrons from cytochrome P450 reductase and in selected cases from cytochrome b5 (cyt b5). Numerous structural studies of cyt P450s, cyt b5, and their reductases have given considerable insight into fundamental structure-function relationships. However, structural studies so far have had to rely on truncated variants of the enzymes to make conventional X-ray crystallographic and solution-state NMR techniques applicable. In spite of significant efforts it has not yet been possible to crystallize any of these proteins in their full-length membrane bound forms. The truncated parts of the enzymes are assumed to be alpha-helical membrane anchors that are essential for some key properties of cyt P450s. In the present contribution we set out with a basic overview on the current status of functional and structural studies. Our main aim is to demonstrate how advanced modern solid-state NMR spectroscopic techniques will be able to make substantial progress in cyt P450 research. Solid-state NMR spectroscopy has sufficiently matured over the last decade to be fully applicable to any membrane protein system. Recent years have seen a remarkable increase in studies on membrane protein structure using a host of solid-state NMR techniques. Solid-state NMR is the only technique available today for structural studies on full-length cyt P450 and full-length cyt b5. We aim to give a detailed account of modern techniques as applicable to cyt P450 and cyt b5, to show what has already been possible and what seems to be viable in the very near future.