Cytochrome P450-type hydroxylation and epoxidation in a tyrosine-liganded hemoprotein, catalase-related allene oxide synthase

J Biol Chem. 2012 Jul 13;287(29):24139-47. doi: 10.1074/jbc.M112.364216. Epub 2012 May 24.

Abstract

The ability of hemoproteins to catalyze epoxidation or hydroxylation reactions is usually associated with a cysteine as the proximal ligand to the heme, as in cytochrome P450 or nitric oxide synthase. Catalase-related allene oxide synthase (cAOS) from the coral Plexaura homomalla, like catalase itself, has tyrosine as the proximal heme ligand. Its natural reaction is to convert 8R-hydroperoxy-eicosatetraenoic acid (8R-HPETE) to an allene epoxide, a reaction activated by the ferric heme, forming product via the Fe(IV)-OH intermediate, Compound II. Here we oxidized cAOS to Compound I (Fe(V)=O) using the oxygen donor iodosylbenzene and investigated the catalytic competence of the enzyme. 8R-hydroxyeicosatetraenoic acid (8R-HETE), the hydroxy analog of the natural substrate, normally unreactive with cAOS, was thereby epoxidized stereospecifically on the 9,10 double bond to form 8R-hydroxy-9R,10R-trans-epoxy-eicosa-5Z,11Z,14Z-trienoic acid as the predominant product; the turnover was 1/s using 100 μm iodosylbenzene. The enantiomer, 8S-HETE, was epoxidized stereospecifically, although with less regiospecificity, and was hydroxylated on the 13- and 16-carbons. Arachidonic acid was converted to two major products, 8R-HETE and 8R,9S-eicosatrienoic acid (8R,9S-EET), plus other chiral monoepoxides and bis-allylic 10S-HETE. Linoleic acid was epoxidized, whereas stearic acid was not metabolized. We conclude that when cAOS is charged with an oxygen donor, it can act as a stereospecific monooxygenase. Our results indicate that in the tyrosine-liganded cAOS, a catalase-related hemoprotein in which a polyunsaturated fatty acid can enter the active site, the enzyme has the potential to mimic the activities of typical P450 epoxygenases and some capabilities of P450 hydroxylases.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Catalase / metabolism*
  • Chromatography, High Pressure Liquid
  • Cytochrome P-450 Enzyme System / metabolism*
  • Gas Chromatography-Mass Spectrometry
  • Hemeproteins / metabolism*
  • Hydroxylation
  • Intramolecular Oxidoreductases / metabolism*
  • Iodobenzenes / metabolism
  • Linoleic Acids / metabolism
  • Magnetic Resonance Spectroscopy

Substances

  • Hemeproteins
  • Iodobenzenes
  • Linoleic Acids
  • Cytochrome P-450 Enzyme System
  • Catalase
  • Intramolecular Oxidoreductases
  • hydroperoxide isomerase
  • iodosobenzene