Life Sciences Institute, 210 Washtenaw Avenue, University of Michigan, Ann Arbor, Michigan 48109-2216, USA.
Biochemistry. 2012 Jun 19;51(24):4959-67. doi: 10.1021/bi3004473. Epub 2012 Jun 5.
The enhanced intracellular survival (Eis) protein improves the survival of Mycobacterium smegmatis (Msm) in macrophages and functions as the acetyltransferase responsible for kanamycin A resistance, a hallmark of extensively drug-resistant (XDR) tuberculosis, in a large number of Mycobacterium tuberculosis (Mtb) clinical isolates. We recently demonstrated that Eis from Mtb (Eis_Mtb) efficiently multiacetylates a variety of aminoglycoside (AG) antibiotics. Here, to gain insight into the origin of substrate selectivity of AG multiacetylation by Eis, we analyzed AG acetylation by Eis_Msm, investigated its inhibition, and compared these functions to those of Eis_Mtb. Even though for several AGs the multiacetylation properties of Eis_Msm and Eis_Mtb are similar, there are three major differences. (i) Eis_Msm diacetylates apramycin, a conformationally constrained AG, which Eis_Mtb cannot modify. (ii) Eis_Msm triacetylates paromomycin, which can be only diacetylated by Eis_Mtb. (iii) Eis_Msm only monoacetylates hygromycin, a structurally unique AG that is diacetylated by Eis_Mtb. Several nonconserved amino acid residues lining the AG-binding pocket of Eis are likely responsible for these differences between the two Eis homologues. Specifically, we propose that because the AG-binding pocket of Eis_Msm is more open than that of Eis_Mtb, it accommodates apramycin for acetylation in Eis_Msm, but not in Eis_Mtb. We also demonstrate that inhibitors of Eis_Mtb that we recently discovered can inhibit Eis_Msm activity. These observations help define the structural origins of substrate preference among Eis homologues and suggest that Eis_Mtb inhibitors may be applied against all pathogenic mycobacteria to overcome AG resistance caused by Eis upregulation.
增强型细胞内生存 (Eis) 蛋白可提高耻垢分枝杆菌 (Msm) 在巨噬细胞中的生存能力，并作为乙酰转移酶发挥作用，负责耐多种药物（XDR）结核病的标志性药物卡那霉素 A 的耐药性，这在大量结核分枝杆菌 (Mtb) 临床分离株中均如此。我们最近证明，来自 Mtb 的 Eis（Eis_Mtb）可有效地对多种氨基糖苷 (AG) 抗生素进行多乙酰化。在这里，为了深入了解 Eis 对 AG 多乙酰化的底物选择性的起源，我们分析了 Eis_Msm 对 AG 的乙酰化作用，研究了其抑制作用，并将这些功能与 Eis_Mtb 的功能进行了比较。尽管对于几种 AG，Eis_Msm 和 Eis_Mtb 的多乙酰化性质相似，但仍存在三个主要差异。(i)Eis_Msm 二乙酰化了阿米卡星，这是一种构象受限的 AG，Eis_Mtb 无法修饰。(ii)Eis_Msm 三乙酰化了巴龙霉素，Eis_Mtb 只能将其二乙酰化。(iii)Eis_Msm 仅单乙酰化了潮霉素，这是一种结构独特的 AG，Eis_Mtb 将其二乙酰化。Eis 中 AG 结合口袋周围的几个非保守氨基酸残基可能是这两种 Eis 同系物之间差异的原因。具体而言，我们提出，由于 Eis_Msm 的 AG 结合口袋比 Eis_Mtb 的更开放，因此它可以容纳阿米卡星进行乙酰化，但在 Eis_Mtb 中则不行。我们还证明了我们最近发现的 Eis_Mtb 的抑制剂可以抑制 Eis_Msm 的活性。这些观察结果有助于定义 Eis 同系物之间底物偏好的结构起源，并表明 Eis_Mtb 抑制剂可能被应用于所有致病性分枝杆菌，以克服 Eis 上调引起的 AG 耐药性。
