A single Sec61-complex functions as a protein-conducting channel
A single Sec61-complex functions as a protein-conducting channel
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DOI：
10.1016/j.bbamcr.2008.08.005
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发表时间：
2008-12-01
影响因子：
5.1
通讯作者：
Hartmann, Enno
中科院分区：
文献类型：
--
作者：
Kalies, Kai-Uwe;Stokes, Vivica;Hartmann, Enno
在蛋白质共翻译易位到内质网（ER）的翻译核糖体结合Sec 61-复合物。目前存在两种模型，这些膜蛋白复合物如何形成蛋白传导通道。虽然电子显微镜数据表明，由四个Sec 61复合物组成的环状结构构建了通道，但最近解决的同源细菌蛋白质复合物的晶体结构导致推测实际的隧道仅由一个单独的Sec 61复合物形成。使用蛋白酶保护试验与定量免疫印迹，我们直接研究了哺乳动物蛋白传导通道的结构。我们发现，在天然ER膜中，一个单一的Sec 61 α-分子优先受到膜结合核糖体的保护，无论是在新生多肽的存在和不存在下。此外，我们提出的证据表明，新生的多肽不稳定的环状易位装置形成的四个Sec 61-复合物。此外，我们发现，ER-膜的溶解后，一个单一的Sec 61-复合物足以保护新生的多肽链对添加的蛋白酶。最后，我们可以证明，这个单一的Sec 61复合物允许新生链的运动，当它已被释放的核糖体嘌呤霉素处理。总的来说，我们的数据表明，ER中的活性蛋白传导通道是由单个Sec 61复合物形成的。(C)2008 Elsevier B. V.保留所有权利。
During cotranslational translocation of proteins into the endoplasmic reticulum (ER) translating ribosomes bind to Sec61-complexes. Presently two models exist how these membrane protein complexes might form protein-conducting channels. While electron microscopic data suggest that a ring-like structure consisting of four Sec61-complexes build the channel, the recently solved crystal Structure of a homologous bacterial protein complex led to the speculation that the actual tunnel is formed by just one individual Sec61-complex. Using protease protection assays together with quantitative immunoblotting we directly examined the structure of mammalian protein-conducting channels. We found that in native ER-membranes one single Sec61 alpha-molecule is preferentially protected by a membrane bound ribosome, both, in the presence and absence of nascent polypeptides. In addition we present evidence that the nascent polypeptide destabilizes the ring-like translocation apparatus formed by four Sec61-complexes. Moreover, we found that after solubilization of ER-membranese a single Sec61-complex is sufficient to protect the nascent polypeptide chain against added proteases. Finally, we could show that this single Sec61-complex allows the movement of the nascent chain, when it has been released from the ribosome by puromycin treatment. Collectively, our data suggest that the active protein-conducting channel in the ER is formed by a single Sec61-complex. (C) 2008 Elsevier B.V. All rights reserved.