Involvement of PpiD in Sec-dependent protein translocation.

The periplasmic space in between the inner and outer membrane of Gram-negative bacteria contains numerous chaperones that are involved in the biogenesis and rescue of extra-cytosolic proteins. In contrast to most of those periplasmic chaperones, PpiD is anchored by an N-terminal transmembrane domain within the inner membrane of Escherichia coli. There it is located in close proximity to the SecY subunit of the SecYEG translocon, which is the primary transporter for secretory and membrane proteins. By site-specific cross-linking we now found the periplasmic domain of PpiD also in close vicinity to the SecG subunit of the Sec translocon and we provide the first direct evidence for a functional cooperation between PpiD and the Sec translocon. Thus we demonstrate that PpiD stimulates in a concentration-dependent manner the translocation of two different secretory proteins into proteoliposomes that had been reconstituted with sub-saturating amounts of SecYEG. In addition we found ribosome-associated nascent chains of a secretory protein stalled at SecY also being in close contact to PpiD. Collectively these results suggest that PpiD plays a role in clearing the Sec translocon of newly translocated secretory proteins thereby improving the overall translocation efficiency. Consistent with this conclusion we demonstrate that PpiD contributes to the efficient detachment of newly secreted OmpA from the inner membrane and in doing so, seems to cooperate in a hierarchical manner with other periplasmic chaperones such as SurA, DegP, and Skp.