Allosteric regulation of arylamine N-acetyltransferase 1 by adenosine triphosphate.

In the present study, a screen of adenosine analogs as potential modulators of arylamine-N-acetyltransferase 1 activity identified ATP as an inhibitor within its range of physiological concentrations. Kinetically, ATP was a non-competitive inhibitor with respect to the acetyl acceptor but a competitive inhibitor with respect to the acetyl donor (acetyl-coenzyme A). In silico modelling predicted that ATP bound within the active site cleft arranged with the triphosphate group in close proximity to arginine 127. Since lysine 100 has previously been implicated in the binding of acetyl-coenzyme A to the enzyme, this amino acid was mutated to either an arginine or a glutamine. Both substitutions significantly changed the affinity of ATP for the enzyme, as well as the nature of the interaction to one with a large Hill coefficient (>3). Under these conditions, ATP was a strong allosteric modulator of arylamine-N-acetyltransferase 1 activity. Western blot analysis identified lysine 100 as a site of post-translational modification by acetylation. The results suggest that acetylation of lysine 100 converts arylamine-N-acetyltransferase 1 into a switch modulated by ATP. This observation provides important understanding of the molecular regulation of NAT1 activity and may reveal possible insight into the endogenous role of the enzyme.