Characterization of the helicase and primase activities of the 63-kDa component of the bacteriophage T7 gene 4 protein

J Biol Chem. 1989 Aug 5;264(22):13066-73.


Leading and lagging strand DNA synthesis at the replication fork of bacteriophage T7 DNA requires the helicase and primase activities of the gene 4 protein. Gene 4 protein consists of two colinear polypeptides of 56- and 63-kDa molecular mass. We have demonstrated previously that the 56-kDa protein possesses helicase but lacks primase activity (Bernstein, J. A., and Richardson, C. C. (1988) Proc. Natl. Acad. Sci. U.S.A. 85, 396-400). The 63-kDa gene 4 protein has now been purified from extracts of T7-infected cells. The preparation contains 5-10% contaminating 56-kDa protein, as shown by Western analysis using polyclonal antibodies to the purified 56-kDa protein. The 63-kDa protein catalyzes DNA-dependent dTTP hydrolysis and has helicase activity; both specific activities are similar to those determined for the 56-kDa protein. The 63-kDa protein efficiently synthesizes sequence-specific di-, tri-, and tetraribonucleotides and stimulates the elongation of tetraribonucleotides by T7 DNA polymerase. Although the 56-kDa protein alone lacks primase activity, it enhances the primase activity of the 63-kDa protein 4-fold. This stimulation can be accounted for by a similar increase in the amount of primers synthesized by the 63-kDa protein in the presence of the 56-kDa protein.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Blotting, Western
  • Catalysis
  • Chromatography, Ion Exchange
  • DNA Helicases / isolation & purification
  • DNA Helicases / metabolism*
  • DNA Helicases / physiology
  • DNA Primase
  • Durapatite
  • Escherichia coli Proteins
  • Hydrolysis
  • Hydroxyapatites
  • Molecular Weight
  • Oligoribonucleotides / chemical synthesis
  • Peptide Chain Elongation, Translational / drug effects
  • RNA Nucleotidyltransferases / isolation & purification
  • RNA Nucleotidyltransferases / metabolism*
  • RNA Nucleotidyltransferases / physiology
  • T-Phages / enzymology*
  • T-Phages / genetics
  • Viral Proteins / isolation & purification
  • Viral Proteins / metabolism*
  • Viral Proteins / physiology


  • Escherichia coli Proteins
  • Hydroxyapatites
  • Oligoribonucleotides
  • Viral Proteins
  • Durapatite
  • DNA Primase
  • RNA Nucleotidyltransferases
  • TraI protein, E coli
  • DNA Helicases