Function of YidC for the insertion of M13 Procoat protein in Escherichia coli - Translocation of mutants that show differences in their membrane potential dependence and Sec requirement

The membrane insertion of the Sec-independent M13 Procoat protein in bacter ia requires the membrane electrochemical potential and the integral membran e protein YidC. We show here that YidC is involved in the translocation but not in the targeting of the Procoat protein, because we found the protein was partitioned into the membrane in the absence of YidC. YidC can function also to promote membrane insertion of Procoat mutants that insert independ ently of the membrane potential, proving that the effect of YidC depletion is not due to a dissipation of the membrane potential. We also found that Y idC is absolutely required for Sec-dependent translocation of a long peripl asmic loop of a mutant Procoat in which the periplasmic region has been ext ended from 20 to 194 residues. Furthermore, when Sec-dependent membrane pro teins with large periplasmic domains were overproduced under YidC-limited c onditions, we found that the exported proteins pro-OmpA and pre-peptidoglyc an-associated lipoprotein accumulated in the cytoplasm. This suggests for S ec-dependent proteins that YidC functions at a late stage in membrane inser tion, after the See translocase interacts with the translocating membrane p rotein. These studies are consistent with the understanding that YidC coope rates with the See translocase for membrane translocation and that YidC is required for clearing the protein-conducting channel.