Heme-thiolate sulfenylation of human cytochrome P450 4A11 functions as a redox switch for catalytic inhibition.

Albertolle ME, Kim D, Nagy LD, Yun CH, Pozzi A, Savas Ü, Johnson EF, Guengerich FP
J Biol Chem. 2017 292 (27): 11230-11242

PMID: 28533430 · PMCID: PMC5500791 · DOI:10.1074/jbc.M117.792200

Cytochrome P450 (P450, CYP) 4A11 is a human fatty acid ω-hydroxylase that catalyzes the oxidation of arachidonic acid to the eicosanoid 20-hydroxyeicosatetraenoic acid (20-HETE), which plays important roles in regulating blood pressure regulation. Variants of P450 4A11 have been associated with high blood pressure and resistance to anti-hypertensive drugs, and 20-HETE has both pro- and antihypertensive properties relating to increased vasoconstriction and natriuresis, respectively. These physiological activities are likely influenced by the redox environment, but the mechanisms are unclear. Here, we found that reducing agents ( dithiothreitol and tris(2-carboxyethyl)phosphine) strongly enhanced the catalytic activity of P450 4A11, but not of 10 other human P450s tested. Conversely, added HO attenuated P450 4A11 catalytic activity. Catalytic roles of five of the potentially eight implicated Cys residues of P450 4A11 were eliminated by site-directed mutagenesis. Using an isotope-coded dimedone/iododimedone-labeling strategy and mass spectrometry of peptides, we demonstrated that the heme-thiolate cysteine (Cys-457) is selectively sulfenylated in an HO concentration-dependent manner. This sulfenylation could be reversed by reducing agents, including dithiothreitol and dithionite. Of note, we observed heme ligand cysteine sulfenylation of P450 4A11 e in kidneys and livers derived from transgenic mice. We also detected sulfenylation of murine P450 4a12 and 4b1 heme peptides in kidneys. To our knowledge, reversible oxidation of the heme thiolate has not previously been observed in P450s and may have relevance for 20-HETE-mediated functions.

© 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

MeSH Terms (14)

Animals Catalysis Cytochrome P-450 CYP4A Dithiothreitol Heme Humans Hydrogen Peroxide Hydroxyeicosatetraenoic Acids Kidney Liver Mice Mice, Transgenic Oxidation-Reduction Rats

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