Enhanced intracellular survival (Eis) proteins belonging to the superfamily of the GCN5‐related N‐acetyltransferases play important functions in mycobacterial pathogenesis. In Mycobacterium tuberculosis, Eis enhances the intracellular survival of the bacilli in macrophages by modulating the host immune response and is capable to chemically modify and inactivate aminoglycosides. In nontuberculous mycobacteria (NTM), Eis shares similar functions. However, Mycobacterium abscessus, a multidrug resistant NTM, possesses two functionally distinct Eis homologues, Eis1_Mab and Eis2_Mab. While Eis2_Mab participates in virulence and aminoglycosides resistance, this is not the case for Eis1_Mab, whose exact biological function remains to be determined. Herein, we show that overexpression of Eis1_Mab in M. abscessus fails to induce resistance to aminoglycosides. To clarify why Eis1_Mab is unable to modify this class of antibiotics, we solved its crystal structure bound to its cofactor, acetyl‐CoA. The structure revealed that Eis1_Mab has a typical homohexameric Eis‐like organization. The structural analysis supported by biochemical approaches demonstrated that while Eis1_Mab can acetylate small substrates, its active site is too narrow to accommodate aminoglycosides. Comparison with other Eis structures showed that an extended loop between strands 9 and 10 is blocking the access of large substrates to the active site and movement of helices 4 and 5 reduces the volume of the substrate‐binding pocket to these compounds in Eis1_Mab. Overall, this study underscores the molecular determinants explaining functional differences between Eis1_Mab and Eis2_Mab, especially those inherent to their capacity to modify aminoglycosides.

中文翻译：

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脓肿分枝杆菌 N-乙酰转移酶 Eis1 的结构分析揭示了其无法修饰氨基糖苷类的分子决定因素。

属于 GCN5 相关N-乙酰转移酶超家族的增强细胞内存活 (Eis) 蛋白在分枝杆菌发病机制中起重要作用。在结核分枝杆菌中，Eis 通过调节宿主免疫反应来增强巨噬细胞中杆菌的细胞内存活，并且能够化学修饰和灭活氨基糖苷类。在非结核分枝杆菌 (NTM) 中，Eis 具有相似的功能。然而，脓肿分枝杆菌是一种耐多药 NTM，具有两种功能不同的 Eis 同源物，即 Eis1 _Mab和 Eis2 _Mab。虽然 Eis2 _Mab参与毒力和氨基糖苷类抗性，但 Eis1 并非如此_Mab，其确切的生物学功能仍有待确定。在这里，我们表明 Eis1 _Mab在脓肿分枝杆菌中的过表达不能诱导对氨基糖苷类药物的抗性。为了阐明为什么 Eis1 _Mab无法修饰这类抗生素，我们解决了与其辅因子乙酰辅酶 A 结合的晶体结构。该结构表明 Eis1 _Mab具有典型的同六聚体 Eis 样组织。生化方法支持的结构分析表明，虽然 Eis1 _Mab能乙酰化小底物，其活性位点太窄而不能容纳氨基糖苷类。与其他 Eis 结构的比较表明，链 9 和 10 之间的延伸环阻止大底物进入活性位点，螺旋 4 和 5 的移动减少了 Eis1 _Mab中这些化合物的底物结合袋的体积。_{总的来说，这项研究强调了解释 Eis1 Mab}和 Eis2 Mab之间功能差异的分子决定因素_，尤其是它们修饰氨基糖苷类的能力所固有的那些。