Oxidation of cysteine S-conjugates by rabbit liver microsomes and cDNA-expressed flavin-containing mono-oxygenases: studies with S-(1,2-dichlorovinyl)-L-cysteine, S-(1,2,2-trichlorovinyl)-L-cysteine, S-allyl-L-cysteine, and S-benzyl-L-cysteine.
Rabbit liver microsomes catalyzed the highly stereoselective, NADPH- and time-dependent S-oxidation of S-benzyl-L-cysteine (SBC), S-allyl-L-cysteine (SAC), S-(1,2-dichlorovinyl)-L-cysteine (DCVC), and S-(1,2,2-trichlorovinyl)-L-cysteine (TCVC) to their respective sulfoxides. Methimazole, a flavin-containing mono-oxygenase (FMO) substrate, inhibited S-oxidation of all four conjugates. The cytochrome P450 inhibitor 1-benzylimidazole did not affect SAC, SBC, or DCVC S-oxidation but inhibited the S-oxidation of TCVC. Solubilization of microsomes also inhibited TCVC activity, whereas SBC, SAC, and DCVC activities were not affected. Because these results suggested that FMOs were the major catalysts of SBC, SAC, and DCVC sulfoxidations, the four conjugates were evaluated as substrates for cDNA-expressed rabbit FMO isoforms FMO1, FMO2, FMO3, and FMO5. At equimolar concentrations (10 mM), FMO1 S-oxidized SBC and SAC, but no sulfoxides were detected with DCVC or TCVC. FMO3 S-oxidized all four conjugates. Km values determined with FMO3 were comparable to Km values from rabbit liver microsomes. S-Oxidation by FMO2 was detected only with SAC, and no sulfoxides were detected in incubations with FMO5. These results show that FMO isoforms can catalyze cysteine conjugate S-oxidation and that the specific isoform involved depends on the structure of the cysteine conjugate. The cysteine conjugates with more nucleophilic sulfur atoms, SAC and SBC, were much better FMO substrates than those having the less nucleophilic sulfur atoms DCVC and TCVC. The sulfoxides of TCVC and DCVC were reactive toward GSH, whereas the sulfoxides of SBC and SAC were not reactive. These results provide evidence for different chemical reactivities of these sulfoxides.
Ripp
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Department of Comparative Biosciences and Environmental Toxicology Center, University of Wisconsin, Madison 53706, USA.
Overby
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LH
Philpot
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RM
Elfarra
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D000818:Animals; D003001:Cloning, Molecular; D003545:Cysteine / Q000031:analogs & derivatives* / Q000737:chemistry; D018076:DNA, Complementary; D008862:Microsomes, Liver / Q000378:metabolism*; D010084:Oxidation-Reduction; D010105:Oxygenases / Q000378:metabolism*; D011817:Rabbits; D013329:Structure-Activity Relationship
D018076:DNA, Complementary; D010105:Oxygenases; C020066:dimethylaniline monooxygenase (N-oxide forming); D003545:Cysteine