Initiation of DNA Synthesis on Single-Stranded DNA Templates in Vitro Promoted by the Bacteriophage λ O and P Replication Proteins

Abstract

The bacteriophage λ O and P protein replication initiators, in conjunction with six purified Escherichia coli replication proteins, replicate the single-stranded chromosomes of phages M13 and ΦX174 to a duplex form. Several discrete steps are involved in this DNA synthesis reaction. In an ATP-dependent step that preceeds priming, the λ O and P proteins interact with the Escherichia coli dnaJ and dnaK proteins to transfer the bacterial dnaB protein onto DNA coated with single-stranded DNA binding protein. This creates a stable prepriming intermediate, isolable by gel filtration, that is rapidly primed and replicated upon the addition of primase and DNA polymerase III holoenzyme. Each of the eight proteins required for this nonspecific single strand replication reaction also have physiological roles in the replication of the bacteriophage λ chromosome in vivo. We propose a scheme for the λ O and P protein-dependent initiation of DNA synthesis that may be relevant to strand initiation events occurring during λ DNA replication.