Molecular basis for the omega-regiospecificity of the CYP4A2 and CYP4A3 fatty acid hydroxylases.
The CYP4A fatty acid omega-hydroxylases are involved in important physiological processes such as the regulation of vascular pressure. A previous study of chimeras of the rat CYP4A2 and CYP4A3 enzymes established that the regiochemistry of fatty acid hydroxylation is determined by the first 119 amino acid residues (Hoch, U., Zhang. Z. P., Kroetz, D. L., and Ortiz de Montellano, P. R. (2000) Arch. Biochem. Biophys. 373, 63-71). The role of the individual amino acid differences in this region has therefore been examined by site-specific mutagenesis to determine which residues actually control the omega- versus (omega-1)-regiospecificity. The results indicate that regiospecificity is controlled by the presence or absence of a three-residue insert (Ser-114, Gly-115, Ile-116) in CYP4A3 and by the residue at position 119 (CYP4A3 numbering). Furthermore, analysis of the absolute stereochemistry of the 11-hydroxylauric acid product indicates that this stereochemistry is not very sensitive to changes in the residues that line the substrate access channel. These results define a model of the specificity determinants of an important class of cytochrome P450 enzymes.
Hoch
U
U
Department of Pharmaceutical Chemistry, University of California, San Francisco, California 94143-0446, USA.
Falck
J R
JR
de Montellano
P R
PR
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D042926:Cytochrome P-450 CYP4A; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D007850:Lauric Acids / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D051381:Rats; D017386:Sequence Homology, Amino Acid; D013237:Stereoisomerism; D013379:Substrate Specificity
D017931:DNA Primers; D007850:Lauric Acids; C030358:lauric acid; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D042926:Cytochrome P-450 CYP4A