KID, A SMALL PROTEIN OF THE PARD STABILITY SYSTEM OF PLASMID R1, IS AN INHIBITOR OF DNA-REPLICATION ACTING AT THE INITIATION OF DNA-SYNTHESIS

The Kid and Ks proteins are the killer component and the antagonist be longing to parD, a killer stability system of plasmid R1. The Kid and Ks proteins have been purified, the second one as a C-LYT-Kis fusion t hat conserves the antagonistic activity of the Kis protein, but not it s auto-regulatory potential. Kid inhibits in vitro replication of CoEl to a basal level without altering the superhelicity of the template b ut it does not substantially affect in vitro replication of P4, a DnaA , DnaB, DnaC and DnaG-independent replicon. Kid inhibits lytic inducti on of a lambda, prophage, but this inhibition can be neutralized by ex cess DnaB. In addition, a multicopy dnaB recombinant, but not a multic opy dnaG recombinant, prevents the toxicity associated with this prote in. Inhibition of ColE1 replication by Kid in vitro is prevented by th e C-LYT-Kis protein. Functional analysis indicates that the antagonist ic activity of Ks is independent of its activity as a co-regulator of the parD promoter. It is also;shown that C-LYT-Kis and Kid interact, f orming a tight complex. These results strongly suggest that the toxici ty of the kid protein is due to inhibition of DnaB-dependent DNA repli cation, and that direct protein-protein interactions are involved in t he neutralization of the activity of the killer protein by the antagon ist.