Université Paris Diderot, Sorbonne Paris Cité, Unité de Biologie Fonctionnelle et Adaptative, CNRS EAC4413, 75013 Paris, France.
J Biol Chem. 2013 Aug 2;288(31):22493-505. doi: 10.1074/jbc.M113.468595. Epub 2013 Jun 16.
Arylamine N-acetyltransferases (NATs), a class of xenobiotic-metabolizing enzymes, catalyze the acetylation of aromatic amine compounds through a strictly conserved Cys-His-Asp catalytic triad. Each residue is essential for catalysis in both prokaryotic and eukaryotic NATs. Indeed, in (HUMAN)NAT2 variants, mutation of the Asp residue to Asn, Gln, or Glu dramatically impairs enzyme activity. However, a putative atypical NAT harboring a catalytic triad Glu residue was recently identified in Bacillus cereus ((BACCR)NAT3) but has not yet been characterized. We report here the crystal structure and functional characterization of this atypical NAT. The overall fold of (BACCR)NAT3 and the geometry of its Cys-His-Glu catalytic triad are similar to those present in functional NATs. Importantly, the enzyme was found to be active and to acetylate prototypic arylamine NAT substrates. In contrast to (HUMAN) NAT2, the presence of a Glu or Asp in the triad of (BACCR)NAT3 did not significantly affect enzyme structure or function. Computational analysis identified differences in residue packing and steric constraints in the active site of (BACCR)NAT3 that allow it to accommodate a Cys-His-Glu triad. These findings overturn the conventional view, demonstrating that the catalytic triad of this family of acetyltransferases is plastic. Moreover, they highlight the need for further study of the evolutionary history of NATs and the functional significance of the predominant Cys-His-Asp triad in both prokaryotic and eukaryotic forms.
芳香胺 N-乙酰基转移酶（NATs）是一类外源性代谢酶，通过严格保守的 Cys-His-Asp 催化三联体催化芳香胺化合物的乙酰化。每个残基对于原核和真核 NATs 的催化都是必不可少的。事实上，在（HUMAN）NAT2 变体中，将 Asp 残基突变为 Asn、Gln 或 Glu 会显著降低酶活性。然而，最近在蜡状芽孢杆菌（（BACCR）NAT3）中发现了一种具有催化三联体 Glu 残基的假定非典型 NAT，但尚未对其进行表征。我们在此报告该非典型 NAT 的晶体结构和功能特征。（BACCR）NAT3 的整体折叠和其 Cys-His-Glu 催化三联体的几何形状与功能性 NAT 中的相似。重要的是，发现该酶具有活性并能乙酰化典型的芳基胺 NAT 底物。与（HUMAN）NAT2 不同，（BACCR）NAT3 三联体中的 Glu 或 Asp 的存在并未显著影响酶的结构或功能。计算分析鉴定出（BACCR）NAT3 活性位点中残基堆积和空间限制的差异，使其能够容纳 Cys-His-Glu 三联体。这些发现推翻了传统观点，证明了该乙酰转移酶家族的催化三联体具有可塑。此外，它们强调需要进一步研究 NAT 的进化历史以及在原核和真核形式中普遍存在的 Cys-His-Asp 三联体的功能意义。
