The pharmacogenetics of NAT: structural aspects

Abstract

Arylamine N-acetyltransferases (NATs) catalyze the transfer of an acetyl group from acetyl-CoA to arylhydrazines and to arylamine drugs and carcinogens or to their N-hydroxylated metabolites. NAT plays an important role in detoxification and metabolic activation of xenobiotics and was first identified as the enzyme responsible for inactivation of the antitubercular drug isoniazid, an arylhydrazine. The rate of inactivation was polymorphically distributed in the population: the first example of interindividual pharmacogenetic variation. Polymorphism in NAT activity is primarily due to single nucleotide polymorphisms (SNPs) in the coding region of NAT genes. NAT enzymes are widely distributed in eukaryotes and genome sequences have revealed many homologous members of this enzyme family in prokaryotes. The structures of S almonella typhimurium and Mycobacterium smegmatis NATs have been determined, revealing a unique fold in which a catalytic triad (Cys-His-Asp) forms the active site. Determination of prokaryotic and eukaryotic NAT structures could lead to a better understanding of their role in xenobiotics and endogenous metabolism.

Peroxynitrite Irreversibly Inactivates the Human Xenobioticmetabolizing Enzyme Arylamine N-Acetyltransferase 1 (NAT1) in Human Breast Cancer Cells

“…Crystallographic determination of the structure of the NATs from Salmonella typhimurium and Mycobacterium smegmatis, and the subsequent construction of theoretical models of human NAT1 and NAT2 revealed structural similarities to cysteine proteases (19 -22). These data demonstrated the existence of a conserved cysteine protease-like catalytic triad (Cys, His, and Asp) in NATs (19 -22), confirming the fundamental role in catalysis of a conserved active site cysteine residue (11,23,24). Thus, NATs may be prone to inactivation by oxidative reactions as shown recently for NAT1 (25) and for other reactive cysteine-containing enzymes (26 -29).…”

“…However, it can also modify the tyrosine residues of proteins to yield 3-nitrotyrosine, and this modification may lead to the irreversible inactivation of enzymes (39,49,52). Treatment of human NAT1 with N-acetylimidazole (20 mM), a tyrosine-modifying agent, did not affect the activity of the enzyme (data not shown), confirming that the tyrosine residues of NAT1 are not involved in its catalytic activity (11). We therefore investigated whether peroxynitrite modified NAT1 cysteine residues.…”

“…This raised the possibility that the absence of NATs or their inactivation may contribute to carcinogenesis and/or tumor progression (6,8,9). Despite their high degrees of sequence identity, NAT1 and NAT2 differ markedly in terms of amine-containing acceptor substrates (10,11) and tissue distribution (2,12). Indeed, NAT2 is primarily located in the liver and colon epithelium (13,14), whereas NAT1 seems ubiquitous (15,16).…”

“…NAT isoforms have been detected in a number of species, from bacteria to mammals (11,18). Crystallographic determination of the structure of the NATs from Salmonella typhimurium and Mycobacterium smegmatis, and the subsequent construction of theoretical models of human NAT1 and NAT2 revealed structural similarities to cysteine proteases (19 -22).…”

“…The catalytic activity of NAT enzymes depends on a reactive cysteine residue present in a cysteine protease-like triad. This cysteine residue is known to form a covalent acetylcysteinyl enzyme with the acetyl group of AcCoA, the physiological acetyl donor substrate of NATs (11). We therefore investigated whether AcCoA and CoA (a product resulting from the hydrolysis of AcCoA, which does not form an acetyl enzyme) protected human NAT1 against peroxynitrite-dependent inactivation.…”