Tus prevents overreplication of oriC plasmid DNA

Abstract

Minichromosome plasmid DNA templates containing oriC and two Ter sites, oriented as they are on the Escherichia coli chromosome, have been used to study the role of Tus in termination of bidirectional replication. In replication reactions reconstituted with purified proteins where it could be demonstrated that each active template was replicating bidirectionally, Tus was required to prevent extensive overreplication. In the presence of Tus, both replication forks terminated DNA synthesis at one or the other Ter site in an apparent stepwise manner. First, the progress of one replication fork was arrested by a properly oriented Tus-Ter complex. Then, either because of steric hindrance resulting from the stalled replication machinery of the first fork or because of the formation of a branched DNA structure, the progression of the second opposing fork was halted at the same site on the DNA template. In the absence of Tus, overreplication required DNA ligase and arose via a template strand-switching mechanism. Thus, the role of Tus in E. coli is more likely to prevent overreplication rather than to ensure accurate termination.