Metabolic activation of heterocyclic aromatic amines catalyzed by human arylamine N-acetyltransferase isozymes (NAT1 and NAT2) expressed in Salmonella typhimurium

Abstract

Heterocyclic aromatic amines formed during the cooking of meat and meat-derived products can be activated to reactive metabolites which bind to DNA, induce mutations and cause tumors in animals. A principal route of metabolic activation is N-oxidation to hydroxylamines, and their subsequent activation by acetyltransferase-catalyzed O-acetylation. We have used mutagenicity assays to study O-acetylation of heterocyclic arylhydroxylamines by the two isozymes of human N-acetyltransferase, NAT1 and NAT2, expressed in Salmonella typhimurium. N-Acetylation was also examined, using an HPLC method. In addition, Salmonella strains with endogenous acetyltransferase and lacking this activating activity were used. Hydroxylamines of nine heterocyclic aromatic amines, IQ, isoIQ, MeIQ, MeIQx, NI, PhIP, Glu-P-1, Glu-P-2, and Trp-P-2 were generated in situ by rat liver S9 mix. The strains expressing human NAT1 and lacking acetyltransferase activity showing little or no ability to activate these substrates. The strains expressing human NAT2 and Salmonella acetyltransferase supported to different extents the activation of all the compounds except PhIP and Trp-P-2. N-Acetylation of IQ, MeIQx and PhIP was slow or not detectable. In conclusion, human NAT2 but not NAT1 can O-acetylate heterocyclic hydroxylamines. NAT2 probably plays a key role in the genotoxic effects of the above heterocyclic amines except for PhIP and Trp-P-2, which have NAT2-independent mutagenic activity.

Insights into the O-Acetylation Reaction of Hydroxylated Heterocyclic Amines by Human Arylamine N-Acetyltransferases: A Computational Study

“…51 Josephy and coworkers have shown that mutagenic activation of PhIP in Salmonella is not promoted by hNAT2, but MeIQx genotoxcity is linked to hNAT2 activation. 53 These docking results are not consistent with this result because both molecules are able to bind in the active site of hNAT2.…”

2-Amino-3,8-Dimethylimidazo-[4,5-f]Quinoxaline–Induced DNA Adduct Formation and Mutagenesis in DNA Repair–Deficient Chinese Hamster Ovary Cells Expressing Human Cytochrome P4501A1 and Rapid or Slow Acetylator N-Acetyltransferase 2

“…Metabolic activation of MeIQx by extrahepatic cytochrome P450 isoforms, such as cytochrome P4501A1 (CYP1A1), has also been reported (13). N-hydroxyMeIQx can be further O-acetylated by N-acetyltransferase 2 (NAT2) to the acetoxy derivative that is highly unstable, leading to electrophilic intermediates that are mutagenic (14) and form DNA adducts (15).…”

Protection against Trp-P-2 mutagenicity by purpurin: mechanism of in vitro antimutagenesis

“…Purpurin is one of several hydroxyanthraquinone constituents found in madder root (Kawasaki et al, 1992), extracts of which are used both as herbal remedies for bladder and kidney stones (Gebhardt, 1979) and as food colorings. Previously, we have demonstrated that purpurin is protective against a number of food-derived heterocyclic amines in bacterial mutagenicity assays (Marczylo et al, 1999).…”

“…Unlike other NIntroduction hydroxylated heterocyclic amines, acetylation of Trp-P-2(NHOH) does not increase the mutagenicity (Nagao et al, The cooking of meat and fish forms bacterial mutagens 1983; Saito et al, 1983a;Wild et al, 1995;Marczylo et al, (Sugimura, 1982), which have been identified as rodent carcino-1999). Purpurin inhibits Trp-P-2(NHOH) mutagenicity to the gens (Sugimura, 1985) and are suspected human carcinogens same extent in TA98 and O-acetyltransferase-deficient TA98- (Weisburger, 1993;Adamson et al, 1995;Schut and 1,8-DNP 6 (Marczylo et al, 1999).…”