Manting E H, Kaufmann A, van der Does C, Driessen A J
Department of Microbiology and Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands.
J Biol Chem. 1999 Aug 20;274(34):23868-74. doi: 10.1074/jbc.274.34.23868.
The SecYEG complex constitutes a protein conducting channel across the bacterial cytoplasmic membrane. It binds the peripheral ATPase SecA to form the translocase. When isoleucine 278 in transmembrane segment 7 of the SecY subunit was replaced by a unique cysteine, SecYEG supported an increased preprotein translocation and SecA translocation ATPase activity, and allowed translocation of a preprotein with a defective signal sequence. SecY(I278C)EG binds SecA with a higher affinity than normal SecYEG, in particular in the presence of ATP. The increased translocation activity of SecY(I278C)EG was confirmed in a purified system consisting of SecYEG proteoliposomes, while immunoprecipitation in detergent solution reveal that translocase-preprotein complexes are more stable with SecY(I278C) than with normal SecY. These data imply an important role for SecY transmembrane segment 7 in SecA binding. As improved SecA binding to SecY was also observed with the prlA4 suppressor mutation, it may be a general mechanism underlying signal sequence suppression.
SecYEG复合物构成了一个跨越细菌细胞质膜的蛋白质传导通道。它与外周ATP酶SecA结合形成转位酶。当SecY亚基跨膜区段7中的异亮氨酸278被一个独特的半胱氨酸取代时，SecYEG支持前体蛋白转运增加以及SecA转运ATP酶活性增加，并允许具有缺陷信号序列的前体蛋白进行转运。SecY(I278C)EG与SecA的结合亲和力高于正常的SecYEG，特别是在ATP存在的情况下。SecY(I278C)EG增加的转运活性在由SecYEG蛋白脂质体组成的纯化系统中得到了证实，而在去污剂溶液中的免疫沉淀显示，转位酶-前体蛋白复合物与SecY(I278C)比与正常SecY更稳定。这些数据表明SecY跨膜区段7在SecA结合中起重要作用。由于在prlA4抑制突变中也观察到SecA与SecY的结合得到改善，这可能是信号序列抑制的一种普遍机制。
