Mechanism of termination of DNA replication of Escherichia coli involves helicase-contrahelicase interaction

Using yeast forward and reverse two-hybrid analyses, we have discovered tha t the replication terminator protein Tus of Escherichia coli physically int eracts with DnaB helicase in vivo. We have confirmed this protein-protein i nteraction in vitro. We show further that replication termination involves protein-protein interaction between Tus and DnaB at a critical region of Tu s protein, called the L1 loop. Several mutations located in the L1 loop reg ion not only reduced the protein-protein interaction but also eliminated or reduced the ability of the mutant forms of Tus to arrest DnaB at a Ter sit e. At least one mutation, E49K, significantly reduced Tus-DnaB interaction and almost completely eliminated the contrahelicase activity of Tus protein in vitro without significantly reducing the affinity of the mutant form of Tus for Ter DNA, in comparison with the wild-type protein. The results, co nsidered along with the crystal structure of Tus-Ter complex, not only eluc idate further the mechanism of helicase arrest but also explain the molecul ar basis of polarity of replication fork arrest at Ter sites.