Redox regulation of the human xenobiotic metabolizing enzyme arylamine N-acetyltransferase 1 (NAT1). Reversible inactivation by hydrogen peroxide.
Oxidative stress is increasingly recognized as a key mechanism in the biotransformation and/or toxicity of many xenobiotics. Human arylamine N-acetyltransferase 1 (NAT1) is a polymorphic ubiquitous phase II xenobiotic metabolizing enzyme that catalyzes the biotransformation of primary aromatic amine or hydrazine drugs and carcinogens. Functional and structural studies have shown that NAT1 catalytic activity is based on a cysteine protease-like catalytic triad, containing a reactive cysteine residue. Reactive protein cysteine residues are highly susceptible to oxidation by hydrogen peroxide (H2O2) generated within the cell. We, therefore, investigated whether human NAT1 activity was regulated by this cellular oxidant. Using purified recombinant NAT1, we show here that NAT1 is rapidly (kinact = 420 m-1.min-1) inactivated by physiological concentrations of H2O2. Reducing agents, such as reduced glutathione (GSH), reverse the H2O2-dependent inactivation of NAT1. Kinetic analysis and protection experiments with acetyl-CoA, the physiological acetyl-donor substrate of the enzyme, suggested that the H2O2-dependent inactivation reaction targets the active-site cysteine residue. Finally, we show that the reversible inactivation of NAT1 by H2O2 is due to the formation of a stable sulfenic acid group at the active-site cysteine. Our results suggest that, in addition to known genetically controlled interindividual variations in NAT1 activity, oxidative stress and cellular redox status may also regulate NAT1 activity. This may have important consequences with regard to drug biotransformation and cancer risk.
Atmane
Noureddine
N
CNRS-UMR 7000, Faculté de Médecine Pitié-Salpêtrière, 75013 Paris, France.
Dairou
Julien
J
Paul
Angela
A
Dupret
Jean-Marie
JM
Rodrigues-Lima
Fernando
F
D000105:Acetyl Coenzyme A / Q000378:metabolism; D001191:Arylamine N-Acetyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001711:Biotransformation; D003065:Coenzyme A / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005982:Glutathione Transferase / Q000235:genetics; D006706:Homeostasis; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007700:Kinetics; D010084:Oxidation-Reduction; D011993:Recombinant Fusion Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D014162:Transfection; D015262:Xenobiotics / Q000493:pharmacokinetics*
D004791:Enzyme Inhibitors; D011993:Recombinant Fusion Proteins; D015262:Xenobiotics; D000105:Acetyl Coenzyme A; D006861:Hydrogen Peroxide; D001191:Arylamine N-Acetyltransferase; D005982:Glutathione Transferase; D003065:Coenzyme A; D004229:Dithiothreitol