Meir Aviv, Abdelhai Ahmad, Moskovitz Yoni, Ruthstein Sharon
Department of Chemistry, Faculty of Exact Sciences, Bar Ilan University, Ramat-Gan, Israel.
Department of Chemistry, Faculty of Exact Sciences, Bar Ilan University, Ramat-Gan, Israel.
Biophys J. 2017 Jun 20;112(12):2494-2502. doi: 10.1016/j.bpj.2017.05.013.
Bacterial cells have developed sophisticated systems to deal with the toxicity of metal ions. Escherichia coli CusCFBA is a complex efflux system, responsible for transferring Cu(I) and Ag(I) ions; this system, located in the periplasm, involves four proteins, CusA, CusB, CusC, and CusF. CusA, CusB, and CusC are connected to one another in an oligomerization ratio of 3:6:3 CusA/CusB/CusC to form the CusCBA periplasm membrane transporter. CusB is an adaptor protein that connects the two membrane proteins CusA (inner membrane) and CusC (outer membrane). CusF is a metallochaperone that transfers Cu(I) and Ag(I) to the CusCBA transporter from the periplasm. The crystal structures of CusB, CusC, CusF, and the CusBA complex have been resolved, shedding some light on the efflux mechanism underlying this intriguing system. However, since CusB is an adaptor protein, its role in operating this system is significant, and should be understood in detail. Here, we utilize EPR spectroscopy to target the conformational changes that take place in the full CusB protein upon binding Cu(I). We reveal that CusB is a dimer in solution, and that the orientation of one molecule with respect to the other molecule changes upon Cu(I) coordination, resulting in a more compact CusB structure. These structural and topological changes upon Cu(I) binding probably play the role of a switch for opening the channel and transferring metal ions from CusB to CusC and out of the cell.
细菌细胞已经进化出复杂的系统来应对金属离子的毒性。大肠杆菌的CusCFBA是一个复杂的外排系统，负责转运铜离子（Cu(I)）和银离子（Ag(I)）；该系统位于周质中，涉及四种蛋白质，即CusA、CusB、CusC和CusF。CusA、CusB和CusC以3:6:3的寡聚化比例（CusA/CusB/CusC）相互连接，形成CusCBA周质膜转运体。CusB是一种衔接蛋白，连接着两个膜蛋白CusA（内膜）和CusC（外膜）。CusF是一种金属伴侣蛋白，它将Cu(I)和Ag(I)从周质转运到CusCBA转运体。CusB、CusC、CusF以及CusBA复合物的晶体结构已得到解析，为这个有趣系统的外排机制提供了一些线索。然而，由于CusB是一种衔接蛋白，它在该系统运作中的作用至关重要，需要详细了解。在这里，我们利用电子顺磁共振光谱来研究完整的CusB蛋白在结合Cu(I)时发生的构象变化。我们发现CusB在溶液中是二聚体，并且在Cu(I)配位时，一个分子相对于另一个分子的方向发生变化，导致CusB结构更加紧凑。Cu(I)结合时发生的这些结构和拓扑变化可能起到了打开通道并将金属离子从CusB转运到CusC并排出细胞的开关作用。
