Evidence for the involvement of N-hydroxylation of 3-amino-1-methyl-5H-pyrido[4,3-b]indole by cytochrome P-450 in the covalent binding to DNA.

The involvement of N-hydroxylation of 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) by cytochrome P-450 in the formation of covalent binding of Trp-P-2 to DNA, the induction of his+ revertant in the Ames test, and the formation of the active metabolite were confirmed. Among four cytochrome P-450 preparations, PCB-P-448 and MC-P-448 purified from liver microsomes of polychlorinated biphenyl (PCB)- and 3-methylcholanthrene-treated rats, respectively, showed higher activities for induction of mutation by Trp-P-2 than did the other two preparations, PCB-P-450 and PB-P-450 purified from PCB-and phenobarbital (PB)-treated rats, respectively. PCB-P-448 was more active than was PB-P-450 in metabolizing Trp-P-2 to N-hydroxylated Trp-P-2 (N-hydroxy-Trp-P-2). Cytochrome P-450 with higher capacity to form the N-hydroxylated metabolite induced a larger number of his+ revertants. Larger amounts of [1-14C]Trp-P-2 bound covalently to DNA were also seen when PCB-P-448 was incubated with calf thymus DNA and reduced nicotinamide adenine dinucleotide phosphate than with PCB-P-450 and PB-P-450. The direct binding of N-[ring-3H]hydroxy-Trp-P-2 isolated by high-performance liquid chromatography to calf thymus DNA was also demonstrated. These results indicate that N-hydroxylation of Trp-P-2 is an obligatory step for the covalent binding to DNA and mutagenesis of Trp-P-2. Based on these results, we propose that N-hydroxy-Trp-P-2 produced by cytochrome P-450 is important in the exertion of the mutagenicity of Trp-P-2 as it binds to DNA.