Cytochrome P450-type hydroxylation and epoxidation in a tyrosine-liganded hemoprotein, catalase-related allene oxide synthase.
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.
Department of Pharmacology and the Vanderbilt Institute of Chemical Biology, Vanderbilt University, Nashville, Tennessee 37232, USA.
William E
WE
Boeglin
Alan R
AR
Brash
D002374:Catalase / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D006420:Hemeproteins / Q000378:metabolism*; D006900:Hydroxylation; D019746:Intramolecular Oxidoreductases / Q000378:metabolism*; D007462:Iodobenzenes / Q000378:metabolism; D008041:Linoleic Acids / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy
D006420:Hemeproteins; D007462:Iodobenzenes; D008041:Linoleic Acids; D003577:Cytochrome P-450 Enzyme System; D002374:Catalase; D019746:Intramolecular Oxidoreductases; C021377:hydroperoxide isomerase; C037276:iodosobenzene