Dynamic Interaction of the Sec Translocon with the Chaperone PpiD*
Dynamic Interaction of the Sec Translocon with the Chaperone PpiD*
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DOI：
10.1074/jbc.m114.577916
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发表时间：
2014-06
期刊：
影响因子：
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通讯作者：
Ilie Sachelaru;Narcis-Adrian Petriman;R. Kudva;H. Koch
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文献类型：
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作者：
Ilie Sachelaru;Narcis-Adrian Petriman;R. Kudva;H. Koch
背景：SEC转位蛋白是一种普遍保守的蛋白转运通道，与多种辅助蛋白相互作用。结果：PPID结合于SecY的侧门，可被新生细胞膜蛋白分离，但不能被SecA分离。结论：PPID是SEC易位子的一个新的瞬时亚基。意义：这是首次描述了SecY-伴侣的直接相互作用。SEC转运子构成了细菌中普遍存在的蛋白质转运通道，由SecY、SecE和SecG三个核心成分组成。更多的蛋白质在蛋白质运输的不同阶段与SecYEG相互作用。在靶向过程中，SecYEG与SecA、SRP受体或核糖体相互作用。然后，通过SecYEG与YidC和SecDFYajC复合体的相互作用，促进蛋白质进入或跨膜运输。在蛋白质运输过程中，SecYEG可能还会与蛋白质质量控制机制相互作用，但关于这种相互作用的细节尚不清楚。通过体内和体外定点定向交联，我们发现周质伴侣PPID位于SecY的侧门前面，跨膜结构域通过该门退出SecY通道。最强的接触是螺旋2b的SecY。蓝色天然PAGE分析证实了天然大肠杆菌膜中存在SecYEG-PPID复合体。PPID-SecY相互作用不受SecA的影响，仅受SecYEG与非翻译核糖体结合的影响较小。相反，PPID在膜蛋白插入过程中失去了与SecY侧门的接触。这些数据表明，PPID是细菌SecYEG易位子的一个额外的瞬时亚基。这些数据进一步证明了SEC转运子的高度模块化和多功能性，这可能是其在细菌和真核生物中跨膜运输各种底物的能力所必需的。
Background: The Sec translocon is a universally conserved protein transport channel that interacts with many auxiliary proteins. Results: PpiD binds to the lateral gate of SecY and is detached by nascent membrane proteins but not by SecA. Conclusion: PpiD is a novel transient subunit of the Sec translocon. Significance: This is the first description of a direct SecY-chaperone interaction. The Sec translocon constitutes a ubiquitous protein transport channel that consists in bacteria of the three core components: SecY, SecE, and SecG. Additional proteins interact with SecYEG during different stages of protein transport. During targeting, SecYEG interacts with SecA, the SRP receptor, or the ribosome. Protein transport into or across the membrane is then facilitated by the interaction of SecYEG with YidC and the SecDFYajC complex. During protein transport, SecYEG is likely to interact also with the protein quality control machinery, but details about this interaction are missing. By in vivo and in vitro site-directed cross-linking, we show here that the periplasmic chaperone PpiD is located in front of the lateral gate of SecY, through which transmembrane domains exit the SecY channel. The strongest contacts were found to helix 2b of SecY. Blue native PAGE analyses verify the presence of a SecYEG-PpiD complex in native Escherichia coli membranes. The PpiD-SecY interaction was not influenced by the addition of SecA and only weakly influenced by binding of nontranslating ribosomes to SecYEG. In contrast, PpiD lost contact to the lateral gate of SecY during membrane protein insertion. These data identify PpiD as an additional and transient subunit of the bacterial SecYEG translocon. The data furthermore demonstrate the highly modular and versatile composition of the Sec translocon, which is probably essential for its ability to transport a wide range of substrates across membranes in bacteria and eukaryotes.