Multicopy suppressors for novel antibacterial compounds reveal targets and drug efflux susceptibility
Multicopy suppressors for novel antibacterial compounds reveal targets and drug efflux susceptibility
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DOI：
10.1016/j.chembiol.2004.08.014
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发表时间：
2004-10-01
影响因子：
--
通讯作者：
Brown, ED
中科院分区：
文献类型：
--
作者：
Li, XM;Zolli-Juran, M;Brown, ED
基因剂量经常被用来选择特定突变体和来自高拷贝随机基因组文库的克隆之间的遗传相互作用。我们在此报告首次使用多拷贝抑制作为正向遗传方法来确定通过高通量筛选鉴定的新型抗菌化合物的细胞靶点和潜在耐药机制。对 8640 个用于抑制高渗透性大肠杆菌菌株生长的小分子进行筛选，从含有大肠杆菌基因组文库的克隆中鉴定出 49 个先导物，用于选择抑制子。发现大多数抑制剂编码多药外排泵 AcrB，表明这些化合物是外排的底物。两个先导物产生了含有编码二氢叶酸还原酶（DHFR）的基因 folA 的克隆，被证明在体内靶向 DHFR，并且在体外是竞争性抑制剂。
Gene dosage has frequently been exploited to select for genetic interactions between a particular mutant and clones from a random genomic library at high copy. We report here the first use of multicopy suppression as a forward genetic method to determine cellular targets and potential resistance mechanisms for novel antibacterial compounds identified through high-throughput screening. A screen of 8640 small molecules for growth inhibition of a hyperpermeable strain of Escherichia coli led to the identification of 49 leads for suppressor selection from clones harboring an E. coli genomic library. The majority of suppressors were found to encode the multidrug efflux pump AcrB, indicating that those compounds were substrates for efflux. Two leads, which produced clones containing the gene folA, encoding dihydrofolate reductase (DHFR), proved to target DHFR in vivo and were competitive inhibitors in vitro.