Active site structure and substrate specificity of cytochrome P450 4A1: steric control of ligand approach perpendicular to heme plane.
The active site of an engineered, expressed fusion protein containing the sequences of cytochrome P450 4A1 (lauric acid omega-hydroxylase) and NADPH-cytochrome P450 reductase, has been probed with olefinic, acetylenic, aromatic, and other analogs of its normal substrate. The fusion protein omega-hydroxylates lauric acid, epoxidizes 11-dodocenoic acid, oxidizes 11-dodecynoic acid to 1,12-dodecandioic acid, but does not oxidize 9-phenylnonanoic acid. Nevertheless, the latter is a tight-binding Type I ligand (Ks = 0.77 microM) and a potent inhibitor of lauric acid hydroxylation. These and other observations are used to construct an active site model that accounts for its remarkable substrate and inhibitor specificity.
Bambal
R B
RB
Department of Medicinal Chemistry, University of Kansas, Lawrence 55045-2506, USA.
Hanzlik
R P
RP
D000494:Allosteric Regulation; D001665:Binding Sites; D042926:Cytochrome P-450 CYP4A; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism*; D003998:Dicarboxylic Acids / Q000378:metabolism; D005229:Fatty Acids, Monounsaturated / Q000378:metabolism; D005231:Fatty Acids, Unsaturated / Q000378:metabolism*; D006418:Heme / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D007850:Lauric Acids / Q000378:metabolism; D008024:Ligands; D006899:Mixed Function Oxygenases / Q000737:chemistry* / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
D003998:Dicarboxylic Acids; D005229:Fatty Acids, Monounsaturated; D005231:Fatty Acids, Unsaturated; D007850:Lauric Acids; D008024:Ligands; D011993:Recombinant Fusion Proteins; C030358:lauric acid; C040948:11-dodecynoic acid; D006418:Heme; C054728:11-dodecenoic acid; D003577:Cytochrome P-450 Enzyme System; C036836:dodecanedioic acid; D006899:Mixed Function Oxygenases; D042926:Cytochrome P-450 CYP4A