An array of Escherichia coli clones over-expressing essential proteins:: A new strategy of identifying cellular targets of potent antibacterial compounds
An array of Escherichia coli clones over-expressing essential proteins:: A new strategy of identifying cellular targets of potent antibacterial compounds
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DOI：
10.1016/j.bbrc.2006.08.166
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发表时间：
2006-11-03
影响因子：
3.1
通讯作者：
Bailey, Melissa Wu
中科院分区：
文献类型：
--
作者：
Xu, H. Howard;Real, Lilian;Bailey, Melissa Wu
随着高通量筛选技术的发展，发现抑制细菌细胞增殖的目标化合物变得越来越容易和快捷。然而，用于鉴定有效抗菌抑制剂的细胞靶点的技术的发展已经落后。在这里，我们描述了一种新的策略，目标识别抗菌抑制剂使用一个阵列的大肠杆菌克隆每个过表达一个必需的蛋白质。在概念验证研究中，将8个必需基因克隆到pLex5BA载体中，置于诱导型启动子的控制下。证实了靶蛋白的过表达。对于两个克隆，一个过度表达FabI和其他过度表达MurA酶，宿主细胞成为17倍和139倍更耐特异性抑制剂三氯生和磷霉素，分别，而其他克隆对这些抑制剂的敏感性在诱导基因表达后保持不变。使用混合克隆和单个克隆测定形式证明了通过靶蛋白过表达的靶鉴定。(c)2006年爱思唯尔公司All rights reserved.
With the advancement of high throughput screening, it has become easier and faster to discover hit compounds that inhibit proliferation of bacterial cells. However, development in technologies used to identify cellular targets of potent antibacterial inhibitors has lagged behind. Here, we describe a novel strategy of target identification for antibacterial inhibitors using an array of Escherichia coli clones each over-expressing one essential protein. In a proof-of-concept study, eight essential genes were cloned into pLex5BA vector under the control of an inducible promoter. Over-expression of target proteins was confirmed. For two clones, one over-expressing FabI and the other over-expressing MurA enzymes, the host cells became 17- and 139-fold more resistant to the specific inhibitors triclosan and phosphomycin, respectively, while the susceptibility of other clones towards these inhibitors remained unchanged after induction of gene expression. Target identification via target protein over-expression was demonstrated using both mixed clone and individual clone assay formats. (c) 2006 Elsevier Inc. All rights reserved.