Cumene Hydroperoxide-Supported Demethylation Reactions Catalyzed by Cytochrome P450 2B4 Lacking the NH2-Terminal Sequence

• Published in: Biochemical and biophysical research communications 1999-04, Vol.258 (1), p.32-38
• Main Authors: Zhang, Yiqun, Pernecky, Steven J.
• Format: Article
• Language: English
• Subjects: Aniline Compounds - metabolism; Animals; Aryl Hydrocarbon Hydroxylases; Benzene Derivatives - metabolism; Catalysis; Cytochrome P-450 Enzyme System - chemistry; Cytochrome P-450 Enzyme System - metabolism; cytochrome P450; heterologous expression; hydroperoxides; Hydroxylation; Methylation; Microsomes, Liver - enzymology; Phosphatidylcholines - metabolism; Rabbits; Sequence Deletion; Steroid Hydroxylases - chemistry; Steroid Hydroxylases - metabolism; Substrate Specificity
• ISSN: 0006-291X; 1090-2104
• DOI: 10.1006/bbrc.1999.0569

Catalytic activities of cytochrome P450 2B4 lacking NH2-terminal amino acids 2-27 (wt Δ2B4) and that of truncated 2B4 containing a Pro to Ser mutation at position 221 were examined in a system supported by cumene hydroperoxide. Demethylation activities of either truncated 2B4 with N-methylaniline, N,N-dimethylaniline, andd-benzphetamine were lower than those of liver microsomal 2B4, whereas the rate of 1-phenylethanol oxidation to acetophenone catalyzed by liver microsomal and truncated 2B4 enzymes was nearly the same. TheKmandVmaxvalues for cumene hydroperoxide in the demethylation of N-methylaniline by wt Δ2B4 were 20% and 28%, respectively, of those obtained for 2B4. The reaction with wt Δ2B4 displayed a lesser dependence on phospholipid than did that with 2B4, and a complex relationship between activity and substrate concentration. The results suggest that the NH2-terminal region contributes to interaction of oxidant, substrate, and phospholipid in cumene hydroperoxide-supported reactions catalyzed by cytochrome P450 2B4.