Mechanistic studies on the cytochrome P450-catalyzed dehydrogenation of 3-methylindole.
The mechanism of 3-methyleneindolenine (3MEI) formation from 3-methylindole (3MI) in goat lung microsomes was examined using stable isotope techniques. 3MEI is highly electrophilic, and its production is a principal factor in the systemic pneumotoxicity of 3MI. Noncompetitive intermolecular isotope effects of DV = 3.3 and D(V/K) = 1.1 obtained after deuterium substitution at the 3-methyl position indicated either that hydrogen abstraction from the methyl group was not the initial rate-limiting step or that this step was rate-limiting and was masked by a high forward commitment and low reverse commitment to catalysis. An intramolecular isotope effect of 5.5 demonstrated that hydrogen atom abstraction was probably the initial oxidative and rate-limiting step of 3MI bioactivation or that deprotonation of an aminium cation radical, produced by one-electron oxidation of the indole nitrogen, was rate-limiting. However, a mechanism which requires deprotonation of the aminium cation radical is probably precluded by an unusual requirement for specific base catalysis at a site in the cytochrome P450 enzyme other than the heme iron. The pattern of 18O incorporation into indole-3-carbinol from 18O2 and H(2)18O indicated that approximately 80% of the indole-3-carbinol was formed in goat lung microsomes by hydration of 3MEI. However, the inverse reaction, dehydration of indole-3-carbinol, did not significantly contribute to the formation of 3MEI. These results show that 3MEI was formed in a cytochrome P450-catalyzed dehydrogenation reaction in which the rate-limiting step was presumably hydrogen atom abstraction from the 3-methyl position. The ratio of the amounts of 3MEI to indole-3-carbinol formed (50:1) indicated that dehydrogenation of 3MI is an unusually facile process when compared to the dehydrogenation of other substrates catalyzed by cytochrome P450 enzymes.
Skiles
G L
GL
Department of Pharmacology and Toxicology, University of Utah, Salt Lake City 84112, USA.
Yost
G S
GS
D000818:Animals; D001711:Biotransformation; D002384:Catalysis; D003577:Cytochrome P-450 Enzyme System / Q000494:pharmacology*; D003903:Deuterium / Q000493:pharmacokinetics; D006041:Goats; D006865:Hydrogenation; D007211:Indoles / Q000493:pharmacokinetics; D008168:Lung / Q000201:enzymology / Q000378:metabolism; D051379:Mice; D008862:Microsomes, Liver / Q000201:enzymology / Q000378:metabolism; D010103:Oxygen Isotopes; D012862:Skatole / Q000378:metabolism* / Q000633:toxicity
D007211:Indoles; D010103:Oxygen Isotopes; D003577:Cytochrome P-450 Enzyme System; D012862:Skatole; D003903:Deuterium; C016517:indole-3-carbinol