Entry exclusion in F-like plasmids requires intact TraG in the donor that recognizes its cognate TraS in the recipient.

The mating pair stabilization (Mps) protein of the F plasmid, TraG, is unique to F-like type IV secretion systems. TraG is a polytopic inner-membrane protein with a large C-terminal periplasmic domain that is required for piliation and Mps, whereas the N-terminal region is sufficient for pilus synthesis. The C-terminal region of TraG is thought to be cleaved by the host signal peptidase I to give a fragment called TraG* that is responsible for Mps. Using mutational analysis and cell localization studies, it was shown that TraG* is most probably an artifact caused by non-specific degradation. TraS (173 aa in F), which is involved in entry exclusion (Eex), blocks redundant conjugative DNA synthesis and transport between donor cells, suggesting that it interferes with a signalling pathway required to trigger DNA transfer. Using the F and R100 plasmids, TraG in the donor cell was found to recognize TraS in the recipient cell inner membrane, in a plasmid-specific manner. This activity mapped to aa 610-673 in F TraG, the only region that differs significantly from R100 TraG. Expression of traG or traG* in a recipient cell did not affect mating ability or Eex. These results suggest that TraG may be translocated to the recipient cell, where it contacts the inner membrane, initiating transfer, a process that is blocked by TraS.