Mutagenesis mapping of the protein-protein interaction underlying FusB-type fusidic acid resistance.
Mutagenesis mapping of the protein-protein interaction underlying FusB-type fusidic acid resistance.
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DOI：
10.1128/aac.00198-13
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发表时间：
2013-10
影响因子：
4.9
通讯作者：
O'Neill AJ
中科院分区：
文献类型：
--
作者：
Cox G;Edwards TA;O'Neill AJ
FusB型蛋白代表葡萄球菌对夫西地酸耐药的主要机制，通过结合药物靶点（延伸因子G [EF-G]）并调节其功能发挥作用。为了进一步深入了解这种抗生素耐药性机制，我们试图鉴定对FusB与EF-G相互作用重要的残基，从而描绘FusB-EF-G复合物内的结合界面。用丙氨酸替换FusB C-末端结构域内的四个保守残基（F156、K184、Y187和F208）中的任何一个，废除了该蛋白赋予对梭链孢酸的抗性的能力;纯化的突变蛋白也失去了结合S.金黄色葡萄球菌EF-G。E. coli EF-G通常不能结合FusB型蛋白质，但在从蛋白质结构域IV中删除3个残基的片段（529 SNP 531）后，使其具有与FusB结合的能力。这项研究已经确定了FuSB和EF-G的关键区域，这些区域对于蛋白质之间的相互作用非常重要，这些发现证实了我们之前对耐药蛋白和药物靶点之间形成的复合物结构的计算机模拟预测（G.考克斯，G. S. Thompson，H. T. Jenkins，F. Peske，A. Savelsbergh，M. V. Rodnina，W. Wintermeyer，S. W. Homans，T. A. Edwards和A. J.奥尼尔，美国国家科学院院刊。Acad. Sci.联合S. A. 109：2102-2107，2012）。
FusB-type proteins represent the predominant mechanism of resistance to fusidic acid in staphylococci and act by binding to and modulating the function of the drug target (elongation factor G [EF-G]). To gain further insight into this antibiotic resistance mechanism, we sought to identify residues important for the interaction of FusB with EF-G and thereby delineate the binding interface within the FusB–EF-G complex. Replacement with alanine of any one of four conserved residues within the C-terminal domain of FusB (F156, K184, Y187, and F208) abrogated the ability of the protein to confer resistance to fusidic acid; the purified mutant proteins also lost the ability to bind S. aureus EF-G in vitro. E. coli EF-G, which is not ordinarily able to bind FusB-type proteins, was rendered competent for binding to FusB following deletion of a 3-residue tract (529SNP531) from domain IV of the protein. This study has identified key regions of both FusB and EF-G that are important for the interaction between the proteins, findings which corroborate our previous in silico prediction for the architecture of the complex formed between the resistance protein and the drug target (G. Cox, G. S. Thompson, H. T. Jenkins, F. Peske, A. Savelsbergh, M. V. Rodnina, W. Wintermeyer, S. W. Homans, T. A. Edwards, and A. J. O'Neill, Proc. Natl. Acad. Sci. U. S. A. 109:2102-2107, 2012).