Factors affecting the efficiency of protein secretion in E.coli.

大肠杆菌前蛋白转位酶由外周成分SecA和完整的膜成分SecY、SecE和SecG组成，SecA被认为是通过经历ATP驱动的膜插入和去插入循环将前蛋白运送到由SecY和SecE形成的可能的蛋白质传导通道。SecG通过未知的机制使转位酶变得高效。外加的针对SecG C-末端区域的抗体抑制了前蛋白转运到外翻的膜小泡中，但我们发现，在没有前蛋白质易位的情况下，这一区域暴露在膜小泡的内侧(周质侧)，从而不受外部蛋白酶K的影响。令人惊讶的是，当前蛋白易位开始时，C-末端区域暴露在膜小泡的外部(细胞质侧)，从而被蛋白酶K消化。这种拓扑倒置与SecG功能紧密耦合，并与SecA的插入-去插入循环联系在一起。我们认为，SecG拓扑的倒置周期促进了SecA的插入-去插入循环，从而导致了有效的前蛋白转位。

Preprotein translocase of E.coli comprises a peripheral component, SecA,and integral membrane components, SecY,SecE and SecG.SecA is thought to deliver the preprotein to the putative protein-conducting channel formed by SecY and SecE by undergoing ATP-driven cycles of membrane insertion and deinsertion. SecG renders the translocase highly efficient by unknown mechanism. Preprotein translocation into everted membrane vesicles is inhibited by an externally added antibody raised against the C-terminal region of SecG.However, we found that this region is exposed to the inside (periplasmic side) of membrane vesicles in the absence of preprotein translocation, thereby being protected from external proteinase K.Surprisingly, when preprotein translocation was started with ATP hydrolysis, the C-terminal region was exposed to the outside (cytoplasmic side) of membrane vesicles and thus digested by proteinase K.Another region of SecG showed a change in membrane sidedness, from the cytoplasmic to the periplasmic side, upon preprotein translocation, indicating that SecG undergoes topology inversion. This topology inversion was tightly coupled to the SecG function, and linked with the insertion-deinsertion cycle of SecA.We propose here that the inversion cycle of the SecG topology facilitates the insertion-deinsertion cycle of SecA,thereby causing efficient preprotein translocation.