title: Redox-linked domain movements in the catalytic cycle of cytochrome p450 reductase.
abstract: NADPH-cytochrome P450 reductase is a key component of the P450 mono-oxygenase drug-metabolizing system. There is evidence for a conformational equilibrium involving large-scale domain motions in this enzyme. We now show, using small-angle X-ray scattering (SAXS) and small-angle neutron scattering, that delivery of two electrons to cytochrome P450 reductase leads to a shift in this equilibrium from a compact form, similar to the crystal structure, toward an extended form, while coenzyme binding favors the compact form. We present a model for the extended form of the enzyme based on nuclear magnetic resonance and SAXS data. Using the effects of changes in solution conditions and of site-directed mutagenesis, we demonstrate that the conversion to the extended form leads to an enhanced ability to transfer electrons to cytochrome c. This structural evidence shows that domain motion is linked closely to the individual steps of the catalytic cycle of cytochrome P450 reductase, and we propose a mechanism for this.
authors.affiliation: Henry Wellcome Laboratories for Structural Biology, Department of Biochemistry, University of Leicester, Henry Wellcome Building, Leicester LE1 9HN, UK.
authors.forename: Wei-Cheng
authors.initials: WC
authors.lastname: Huang
authors.forename: Jacqueline
authors.initials: J
authors.lastname: Ellis
authors.forename: Peter C E
authors.initials: PC
authors.lastname: Moody
authors.forename: Emma L
authors.initials: EL
authors.lastname: Raven
authors.forename: Gordon C K
authors.initials: GC
authors.lastname: Roberts
mesh_terms: D055162:Biocatalysis; D020134:Catalytic Domain; D005486:Flavin Mononucleotide / Q000737:chemistry; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D009249:NADP / Q000737:chemistry; D009251:NADPH-Ferrihemoprotein Reductase / Q000737:chemistry*; D010084:Oxidation-Reduction; D011485:Protein Binding; D017433:Protein Structure, Secondary; D053838:Scattering, Small Angle; D014961:X-Ray Diffraction
chemical_list: D009249:NADP; D005486:Flavin Mononucleotide; D009251:NADPH-Ferrihemoprotein Reductase
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