The linker region between the helicase and primase domains of the bacteriophage T7 gene 4 protein is critical for hexamer formation

J Biol Chem. 1999 Oct 15;274(42):30303-9. doi: 10.1074/jbc.274.42.30303.


The gene 4 protein of bacteriophage T7, a functional hexamer, comprises DNA helicase and primase activities. Both activities depend on the unidirectional movement of the protein along single-stranded DNA in a reaction coupled to the hydrolysis of dTTP. We have characterized dTTPase activity and hexamer formation for the full-length gene 4 protein (gp4) as well as for three carboxyl-terminal fragments starting at residues 219 (gp4-C219), 241 (gp4-C241), and 272 (gp4-C272). The region between residues 242 and 271, residing between the primase and helicase domains, is critical for oligomerization of the gene 4 protein. A functional TPase active site is dependent on oligomerization. During native gel electrophoresis, gp4, gp4-C219, and gp4-C241 migrate as oligomers, whereas gp4-C272 is monomeric. The steady-state k(cat) for dTTPase activity of gp4-C272 increases sharply with protein concentration, indicating that it forms oligomers only at high concentrations. gp4-C219 and gp4-C241 both form a stable complex with gp4, whereas gp4-C272 interacts only weakly with gp4. Measurements of surface plasmon resonance indicate that a monomer of T7 DNA polymerase binds to a dimer of gp4, gp4-C219, or gp4-C241 but to a monomer of gp4-C272. Like the homologous RecA and F(1)-ATPase proteins, the oligomerization domain of the gene 4 protein is adjacent to the amino terminus of the NTP-binding domain.

Publication types

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

MeSH terms

  • Bacteriophage T7 / enzymology
  • Base Sequence
  • Biopolymers
  • Catalysis
  • Cloning, Molecular
  • DNA Helicases / metabolism*
  • DNA Primase / genetics
  • DNA Primase / metabolism*
  • DNA Primers
  • DNA-Directed DNA Polymerase / metabolism
  • Hydrolysis
  • Peptide Fragments / isolation & purification
  • Pyrophosphatases / metabolism
  • Surface Plasmon Resonance
  • Thymine Nucleotides / metabolism


  • Biopolymers
  • DNA Primers
  • Peptide Fragments
  • Thymine Nucleotides
  • DNA Primase
  • DNA-Directed DNA Polymerase
  • Pyrophosphatases
  • thymidine-triphosphatase
  • DNA Helicases
  • thymidine 5'-triphosphate