Small-Molecule Inhibitors of Staphylococcus aureus RnpA-Mediated RNA Turnover and tRNA Processing
Small-Molecule Inhibitors of Staphylococcus aureus RnpA-Mediated RNA Turnover and tRNA Processing
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DOI：
10.1128/aac.04352-14
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发表时间：
2015-04-01
影响因子：
4.9
通讯作者：
Dunman, Paul M.
中科院分区：
文献类型：
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作者：
Eidem, Tess M.;Lounsbury, Nicole;Dunman, Paul M.
治疗金黄色葡萄球菌感染迫切需要新的药物。在这方面，金黄色葡萄球菌核糖核酸酶RnpA可能是一个很有前途的新型双功能抗菌靶点，它参与了两个重要的细胞过程：RNA降解和tRNA成熟。因此，我们之前使用高通量筛选来鉴定该酶RNA降解活性的小分子抑制剂，并表明RnpA抑制剂RNPA1000是一个有吸引力的抗微生物开发候选药物。在这项研究中，我们使用了一系列体外和细胞测试来表征第二个RnpA抑制剂RNPA2000，它是在我们的初步筛选活动中确定的，在结构上与RNPA1000不同。在这样做的过程中，发现金黄色葡萄球菌RnpA确实如先前假设的那样参与了5‘-前体tRNA的加工。此外，我们证明RNPA2000是一种杀菌剂，在体外和金黄色葡萄球菌内都能抑制RnpA相关的RNA降解和tRNA成熟活性。该化合物表现出对RnpA的特异性，因为它不会显著影响无关细菌或真核核糖核酸酶的体外活性，也没有表现出可测量的人类细胞毒性。最后，我们证明了RNPA2000具有抗菌活性，并抑制外排缺陷的革兰氏阴性病原体的tRNA加工。综上所述，这些数据支持将RnpA用于抗菌药物开发的靶向，确立了可以识别具有两种酶功能的小分子抑制剂，并提供了RnpA抑制剂可能具有广谱抗菌活性的证据。
New agents are urgently needed for the therapeutic treatment of Staphylococcus aureus infections. In that regard, S. aureus RNase RnpA may represent a promising novel dual-function antimicrobial target that participates in two essential cellular processes, RNA degradation and tRNA maturation. Accordingly, we previously used a high-throughput screen to identify small-molecule inhibitors of the RNA-degrading activity of the enzyme and showed that the RnpA inhibitor RNPA1000 is an attractive antimicrobial development candidate. In this study, we used a series of in vitro and cellular assays to characterize a second RnpA inhibitor, RNPA2000, which was identified in our initial screening campaign and is structurally distinct from RNPA1000. In doing so, it was found that S. aureus RnpA does indeed participate in 5'-precursor tRNA processing, as was previously hypothesized. Further, we show that RNPA2000 is a bactericidal agent that inhibits both RnpA-associated RNA degradation and tRNA maturation activities both in vitro and within S. aureus. The compound appears to display specificity for RnpA, as it did not significantly affect the in vitro activities of unrelated bacterial or eukaryotic ribonucleases and did not display measurable human cytotoxicity. Finally, we show that RNPA2000 exhibits antimicrobial activity and inhibits tRNA processing in efflux-deficient Gram-negative pathogens. Taken together, these data support the targeting of RnpA for antimicrobial development purposes, establish that small-molecule inhibitors of both of the functions of the enzyme can be identified, and lend evidence that RnpA inhibitors may have broad-spectrum antimicrobial activities.