CONJUGATION-INDEPENDENT, SITE-SPECIFIC RECOMBINATION AT THE ORIT OF THE INCW PLASMID R388 MEDIATED BY TRWC

Plasmids containing a direct repeat of plasmid R388 oriT are capable o f site-specific recombination, which results in deletion of the interv ening DNA. This reaction occurs in the presence, but not in the absenc e, of the region of R388 implicated in DNA processing during conjugati on. This region contains three genes, trwA, trwB, and trwC. By using m utants of each of the three genes, it was demonstrated that only trwC is required for the oriT-specific recombination. Further analysis show ed that the N-terminal 272 amino acids of the protein are sufficient t o catalyze recombination. TrwC is also capable of promoting intermolec ular recombination between two plasmids containing oriT, suggesting th at double-strand breaks in both plasmid DNAs are involved in the proce ss. Additionally, intramolecular recombination between R388 oriT and R 46 oriT did not occur in the presence of both nickases. This suggests that the half-reactions at each oriT are not productive if they occur separately; therefore, an interaction between the recombination comple xes formed at each recombining site is required. This is the first rep ort in which a nicking-closing enzyme involved in conjugal DNA transfe r promotes oriT-specific recombination of double-stranded DNA in the a bsence of conjugation.