van der Sluis Eli O, Nouwen Nico, Koch Joachim, de Keyzer Jeanine, van der Does Chris, Tampé Robert, Driessen Arnold J M
Department of Molecular Microbiology, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, 9751 NN Haren, The Netherlands.
J Mol Biol. 2006 Sep 1;361(5):839-49. doi: 10.1016/j.jmb.2006.07.017. Epub 2006 Jul 15.
The motor protein SecA drives the translocation of (pre-)proteins across the SecYEG channel in the bacterial cytoplasmic membrane by nucleotide-dependent cycles of conformational changes often referred to as membrane insertion/de-insertion. Despite structural data on SecA and an archaeal homolog of SecYEG, the identity of the sites of interaction between SecA and SecYEG are unknown. Here, we show that SecA can be cross-linked to several residues in cytoplasmic loop 5 (C5) of SecY, and that SecA directly interacts with a part of transmembrane segment 4 (TMS4) of SecY that is buried in the membrane region of SecYEG. Mutagenesis of either the conserved Arg357 in C5 or Glu176 in TMS4 interferes with the catalytic activity of SecA but not with binding of SecA to SecYEG. Our data explain how conformational changes in SecA could be directly coupled to the previously proposed opening mechanism of the SecYEG channel.
马达蛋白SecA通过通常被称为膜插入/去插入的核苷酸依赖性构象变化循环，驱动（前体）蛋白质穿过细菌细胞质膜中的SecYEG通道。尽管有关于SecA和SecYEG的古细菌同源物的结构数据，但SecA与SecYEG之间相互作用位点的身份尚不清楚。在这里，我们表明SecA可以与SecY的细胞质环5（C5）中的几个残基交联，并且SecA直接与SecY的跨膜片段4（TMS4）中埋在SecYEG膜区域的一部分相互作用。C5中保守的Arg357或TMS4中的Glu176的诱变会干扰SecA的催化活性，但不会干扰SecA与SecYEG的结合。我们的数据解释了SecA中的构象变化如何能够直接与先前提出的SecYEG通道的开放机制相耦合。
