The highly stereoselective oxidation of polyunsaturated fatty acids by cytochrome P450BM-3

Jorge H. Capdevila, Shozou Wei, Christian Helvig, J R Falck, Yuri Belosludtsev, Gilles Truan, Sandra E. Graham-Lorence, Julian A. Peterson

Research output: Contribution to journalArticle

139 Scopus citations

Abstract

Cytochrome P450BM-3 catalyzes NADPH-dependent metabolism of arachidonic acid to nearly enantiomerically pure 18(R)-hydroxyeicosatetraenoic acid and 14(S),15(R)-epoxyeicosatrienoic acid (80 and 20% of total products, respectively). P450BM-3 oxidizes arachidonic acid with a rate of 3.2 ± 0.4 μmol/min/nmol at 30 °C, the fastest ever reported for an NADPH-dependent, P450-catalyzed reaction. Fatty acid, oxygen, and NADPH are utilized in an approximately 1:1:1 molar ratio, demonstrating efficient coupling of electron transport to monooxygenation. Eicosapentaenoic and eicosatrienoic acids, two arachidonic acid analogs that differ in the properties of the C-15-C-18 carbons, are also actively metabolized by P450BM-3 (1.4 ± 0.2 and 2.9 ± 0.1 μmol/min/nmol at 30 °C, respectively). While the 17,18-olefinic bond of eicosapentaenoic acid is epoxidized with nearly absolute regioand stereochemical selectivity to 17(S),18(R)-epoxyeicosatetraenoic acid (≤99% of total products, 97% optical purity), P450BM-3 is only moderately regioselective during hydroxylation of the eicosatrienoic acid ω-1, ω-2, and ω-3 sp3 carbons, with 17-, 18-, and 19-hydroxyeicosatrienoic acid formed in a ratio of 2.4:2.2:1, respectively. Based on the above and on a model of arachidonic acid-bound P450BM-3, we propose: 1) the formation by P450BM-3 of a single oxidant species capable of olefinic bond epoxidation and sp3 carbon hydroxylation and 2) that product chemistry and, thus, catalytic outcome are critically dependent on active site spatial coordinates responsible for substrate binding and productive orientation between heme- bound active oxygen and acceptor carbon bond(s).

Original languageEnglish (US)
Pages (from-to)22663-22671
Number of pages9
JournalJournal of Biological Chemistry
Volume271
Issue number37
DOIs
StatePublished - Sep 25 1996

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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    Capdevila, J. H., Wei, S., Helvig, C., Falck, J. R., Belosludtsev, Y., Truan, G., Graham-Lorence, S. E., & Peterson, J. A. (1996). The highly stereoselective oxidation of polyunsaturated fatty acids by cytochrome P450BM-3. Journal of Biological Chemistry, 271(37), 22663-22671. https://doi.org/10.1074/jbc.271.37.22663