The T341C (lle114Thr) polymorphism of N-acetyltransferase 2 yields slow acetylator phenotype by enhanced protein degradation

Objectives Human N-acetyltransferase 2 (NAT2) plays a significant role in the clearance and biotransformation of many drugs and carcinogens. A (TC)-C-341 (Ile1 14Thr) single nucleotide polymorphism (SNP) of NAT2 is commonly found in slow acetylators, leading to altered drug response and toxicity and possibly cancer susceptibility from carcinogens. The objective of this study was to investigate the mechanism by which this SNP causes slow acetylator phenotype. Methods A cDNA expression system was used to compare the NAT2*4 reference allele with an identical one possessing the (TC)-C-341 SNP in COS-1 cells. The recombinant human NAT2 enzymes were compared in regard to catalytic activity, kinetic parameters, thermostability, immunoreactive protein level, mRNA level and in-vivo protein degradation. Results The (TC)-C-341 (Ile1 14Thr) SNP significantly reduced enzyme activity without changing the apparent kinetic parameters K-m and V-max (normalized for NAT2 protein), indicating that Ile1 14Thr did not change substrate or cofactor binding affinities or catalytic efficiency. Furthermore, no significant difference in NAT2 mRNA level was observed, indicating no impairment of transcription. The (TC)-C-341 (Ile1 14Thr) SNP did not alter thermostability of NAT2 at either 37 or 50degreesC. However, this SNP significantly reduced cytosolic NAT2 immunoreactive protein through enhanced protein degradation. Conclusion This is the first report indicating that protein degradation is an important mechanism of human NAT2 slow acetylator phenotype. (C) 2004 Lippincott Williams Wilkins.