Charged residues render pro-OmpA potential dependent for initiation of membrane translocation

We have examined the effects of positively and negatively charged residues on the translocation of outer membrane protein A precursor (pro-OmpA) across the bacterial inner membrane. Pro-OmpA does not translocate across the membrane when 2 positively charged residues are inserted immediately after the leader peptide, whereas it does insert when 2 neutral or negatively charged residues are introduced. Using a cell-free translocation system, we show that the membrane potential stimulated the rate of initial insertion of pro-OmpA with negatively charged residues, inhibited pro-OmpA with positively charged residues, and had no effect on neutral pro-OmpA. Thus, acidic residues render pro-OmpA potential-dependent for loop formation, which then initiates the translocation process.