On the nature of the cytochrome P450scc "ultimate oxidant": characterization of a productive radical intermediate.
The electron paramagnetic (EPR) properties of a transient species detected during a cytochrome P450 (P450)-mediated peroxidative reaction have been compared with those of peroxidases Compound I and model metalloporphyrins. The reaction which was studied with cholesterol-specific P450scc and (20R)-20-hydroperoxycholesterol, occurred without enzyme denaturation. The resulting transient species, which reached its maximum after 50 s reaction, was characterized by a one-line EPR spectrum, g = 2.0035, delta 1/2 = 1.08 mT. The decay of this radical was concomitant with the production of (20R)-20,21-dihydroxycholesterol. The reaction (almost 100% yield) conserved the stereo-specificity of the natural pathway. We suggest this intermediate is a candidate for the in vivo ultimate oxidant in the P450-mediated hydroxylation process. The comparison of the observed EPR spectrum with those of peroxidase Compound I and related synthetic models allows us to propose a FeIV porphyrin II cation radical structure for the intermediate.
Larroque
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Institut National de la Santé et de la Recherche Médicale, Unité 128, Montpellier, France.
Lange
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Maurin
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Bienvenue
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A
van Lier
J E
JE
D000302:Adrenal Cortex / Q000201:enzymology*; D000818:Animals; D002248:Carbon Monoxide / Q000378:metabolism; D002417:Cattle; D002784:Cholesterol / Q000031:analogs & derivatives / Q000378:metabolism; D002786:Cholesterol Side-Chain Cleavage Enzyme / Q000378:metabolism*; D004578:Electron Spin Resonance Spectroscopy; D005609:Free Radicals; D006888:Hydroxycholesterols / Q000378:metabolism; D006900:Hydroxylation; D007700:Kinetics; D010084:Oxidation-Reduction; D011485:Protein Binding
D005609:Free Radicals; D006888:Hydroxycholesterols; C038027:20-hydroperoxycholesterol; C020890:20,21-dihydroxycholesterol; D002248:Carbon Monoxide; D002784:Cholesterol; D002786:Cholesterol Side-Chain Cleavage Enzyme