HIGHLY EFFICIENT DNA-SYNTHESIS BY THE PHAGE PHI-29 DNA-POLYMERASE - SYMMETRICAL MODE OF DNA-REPLICATION

  • 25 May 1989
    • journal article
    • research article
    • Vol. 264  (15) , 8935-8940
Abstract
The results presented in this paper indicate that the .vphi.29 DNA polymerase is the only enzyme required for efficient synthesis of full length .vphi.29 DNA with the .vphi.29 terminal protein, the initiation primer, as the only additional proein requirement. Analysis of .vphi.29 DNA polymerase activity in various in vitro DNA replication systems indicates that two main reasons are responsible for the efficiency of this minimal system: 1) the .vphi.29 DNA polymerase is highly processive in the absence of any accessory protein; 2) the polymerase itself is able to produce strand displacement coupled to the polymerization process. Using primed M13 DNA as template, the .vphi.29 DAN polymerase is able to synthesize DNA chains greater than 70 kilobase pairs. Furthermore, conditions that increase the stability of secondary structure in the template do not affect the processivity and strand displacement ability of the enzyme. Thus, the catalytic properties of the .vphi.29 DNA polymeras are appropriate for a .vphi.29 DNA replication mechanism involving two replication origins, strand displacement and continuous synthesis of both strands. The enzymology of .vphi.29 DNA replication would support a symmetrical model of DNA replication.