title: Oxidation of cycloalkylamines by cytochrome P-450. Mechanism-based inactivation, adduct formation, ring expansion, and nitrone formation.
abstract: Mechanism-based destruction of cytochrome P-450 (P-450) and P-450 heme is observed during the oxidation of N-cyclopropyl and N-cyclobutyl benzylamines. The slower inactivation by the cyclobutylamines relative to cyclopropylamines is consistent with known relative rates of ring opening of cycloalkyl-substituted aminium radicals. Evidence was found that porphyrin meso adducts of the type reported for horseradish peroxidase and cyclopropanone hydrate (Wiseman, J. S., Nichols, J. S., and Kolpak, M. X. (1982) J. Biol. Chem. 257, 6328-6332) were not formed. Radiolabels from cyclopropylamine substrates were covalently attached to protein but essentially only from the cyclopropyl portion and not the benzylic portion. Neither label appeared to be bound to extractable heme; however, during oxidations with cyclopropylamines, labeled P-450 heme became covalently attached to protein. Oxidation of 1-phenylcyclobutylamine by P-450 yielded 2-phenyl-1-pyrroline and 2-phenylpyrrolidine, and the ring expansion is interpreted as evidence for the existence of aminium radicals based on precedents with monoamine oxidase (Silverman, R. B., and Zieske, P. A. (1985) Biochemistry 24, 2128-2138). In addition, purified P-450PB-B oxidized N-(1-phenylcyclobutyl)-benzylamine to N-(1-phenyl)cyclobutyl phenyl nitrone, identified using spectral techniques. This transformation involves two sequential oxidations with either a hydroxylamine or benzylidene intermediate. While P-450 oxidized the amine to both compounds, only the hydroxylamine was rapidly oxidized to give the nitrone. The ring expansion and nitrone products are interpreted in the context of aminium radical intermediates involved in the mechanism of P-450-catalyzed amine oxidation.
authors.affiliation: Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, Tennessee 37232.
authors.forename: A
authors.initials: A
authors.lastname: Bondon
authors.forename: T L
authors.initials: TL
authors.lastname: Macdonald
authors.forename: T M
authors.initials: TM
authors.lastname: Harris
authors.forename: F P
authors.initials: FP
authors.lastname: Guengerich
mesh_terms: D000818:Animals; D001596:Benzylamines / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism; D015394:Molecular Structure; D051381:Rats; D013379:Substrate Specificity
chemical_list: D001596:Benzylamines; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System