Howard Hughes Medical Institute and Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA.
J Cell Biol. 2012 Sep 3;198(5):881-93. doi: 10.1083/jcb.201205140. Epub 2012 Aug 27.
The transport of proteins across the plasma membrane in bacteria requires a channel formed from the SecY complex, which cooperates with either a translating ribosome in cotranslational translocation or the SecA ATPase in post-translational translocation. Whether translocation requires oligomers of the SecY complex is an important but controversial issue: it determines channel size, how the permeation of small molecules is prevented, and how the channel interacts with the ribosome and SecA. Here, we probe in vivo the oligomeric state of SecY by cross-linking, using defined co- and post-translational translocation intermediates in intact Escherichia coli cells. We show that nontranslocating SecY associated transiently through different interaction surfaces with other SecY molecules inside the membrane. These interactions were significantly reduced when a translocating polypeptide inserted into the SecY channel co- or post-translationally. Mutations that abolish the interaction between SecY molecules still supported viability of E. coli. These results show that a single SecY molecule is sufficient for protein translocation.
细菌跨膜蛋白运输需要由 SecY 复合物形成的通道，该通道与共翻译易位中的翻译核糖体或翻译后易位中的 SecA ATP 酶合作。易位是否需要 SecY 复合物的寡聚体是一个重要但有争议的问题：它决定了通道的大小，如何防止小分子的渗透，以及通道如何与核糖体和 SecA 相互作用。在这里，我们通过交联在体内探测 SecY 的寡聚状态，使用完整的大肠杆菌细胞中的定义的共翻译和翻译后易位中间体。我们表明，非易位的 SecY 通过不同的相互作用表面与膜内的其他 SecY 分子短暂相关。当易位多肽共翻译或翻译后插入 SecY 通道时，这些相互作用显著减少。消除 SecY 分子之间相互作用的突变仍然支持大肠杆菌的存活。这些结果表明，单个 SecY 分子足以进行蛋白质易位。
