Zenser T V, Lakshmi V M, Rustan T D, Doll M A, Deitz A C, Davis B B, Hein D W
Veterans Affairs Medical Center, St. Louis, Missouri 63125-4199, USA.
Cancer Res. 1996 Sep 1;56(17):3941-7.
These studies were designed to assess metabolism of benzidine and N-acetylbenzidine by N-acetyltransferase (NAT) NAT1 and NAT2. Metabolism was assessed using human recombinant NAT1 and NAT2 and human liver slices. For benzidine and N-acetylbenzidine, Km and Vmax values were higher for NAT1 than for NAT2. The clearance ratios (NAT1/NAT2) for benzidine and N-acetylbenzidine were 54 and 535, respectively, suggesting that N-acetylbenzidine is a preferred substrate for NAT1. The much higher NAT1 and NAT2 Km values for N-acetylbenzidine (1380 +/- 90 and 471 +/- 23 microM, respectively) compared to benzidine (254 +/- 38 and 33.3 +/- 1.5 microM, respectively) appear to favor benzidine metabolism over N-acetylbenzidine for low exposures. Determination of these kinetic parameters over a 20-fold range of acetyl-CoA concentrations demonstrated that NAT1 and NAT2 catalyzed N-acetylation of benzidine by a binary ping-pong mechanism. In vitro enzymatic data were correlated to intact liver tissue metabolism using human liver slices. Samples incubated with either [3H]benzidine or [3H]N-acetylbenzidine had a similar ratio of N-acetylated benzidines (N-acetylbenzidine + N',N'-diacetylbenzidine/ benzidine) and produced amounts of N-acetylbenzidine > benzidine > N,N'-diacetylbenzidine. With [3H]benzidine, p-aminobenzoic acid, a NAT1-specific substrate, increased the amount of benzidine and decreased the amount of N-acetylbenzidine produced, resulting in a decreased ratio of acetylated products. This is consistent with benzidine being a NAT1 substrate. N-Acetylation of benzidine or N-acetylbenzidine by human liver slices did not correlate with the NAT2 genotype. However, a higher average acetylation ratio was observed in human liver slices possessing the NAT110 compared to the NAT14 allele. Thus, a combination of human recombinant NAT and liver slice experiments has demonstrated that benzidine and N-acetylbenzidine are both preferred substrates for NAT1. These results also suggest that NAT1 may exhibit a polymorphic expression in human liver.
这些研究旨在评估N - 乙酰转移酶（NAT）NAT1和NAT2对联苯胺和N - 乙酰联苯胺的代谢情况。使用人重组NAT1和NAT2以及人肝切片评估代谢。对于联苯胺和N - 乙酰联苯胺，NAT1的Km和Vmax值高于NAT2。联苯胺和N - 乙酰联苯胺的清除率（NAT1/NAT2）分别为54和535，这表明N - 乙酰联苯胺是NAT1的优选底物。与联苯胺（分别为254±38和33.3±1.5 microM）相比，N - 乙酰联苯胺的NAT1和NAT2 Km值高得多（分别为1380±90和471±23 microM），这似乎有利于低暴露下联苯胺的代谢而非N - 乙酰联苯胺。在20倍范围的乙酰辅酶A浓度下测定这些动力学参数表明，NAT1和NAT2通过双乒乓机制催化联苯胺的N - 乙酰化。使用人肝切片将体外酶学数据与完整肝组织代谢相关联。用[3H]联苯胺或[3H]N - 乙酰联苯胺孵育的样品具有相似的N - 乙酰化联苯胺比例（N - 乙酰联苯胺+ N'，N'-二乙酰联苯胺/联苯胺），并且产生的N - 乙酰联苯胺量>联苯胺> N，N'-二乙酰联苯胺。对于[3H]联苯胺，对氨基苯甲酸是一种NAT1特异性底物，它增加了联苯胺的量并减少了产生的N -乙酰联苯胺的量，导致乙酰化产物的比例降低。这与联苯胺是NAT1底物一致。人肝切片对联苯胺或N - 乙酰联苯胺的N - 乙酰化与NAT2基因型无关。然而，与NAT14等位基因相比，在具有NAT110的人肝切片中观察到更高的平均乙酰化率。因此，人重组NAT和肝切片实验的结合表明，联苯胺和N - 乙酰联苯胺都是NAT1的优选底物。这些结果还表明NAT1可能在人肝脏中表现出多态性表达。
