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

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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.