Helicase-DNA polymerase interaction is critical to initiate leading-strand DNA synthesis

Proc Natl Acad Sci U S A. 2011 Jun 7;108(23):9372-7. doi: 10.1073/pnas.1106678108. Epub 2011 May 23.

Abstract

Interactions between gene 4 helicase and gene 5 DNA polymerase (gp5) are crucial for leading-strand DNA synthesis mediated by the replisome of bacteriophage T7. Interactions between the two proteins that assure high processivity are known but the interactions essential to initiate the leading-strand DNA synthesis remain unidentified. Replacement of solution-exposed basic residues (K587, K589, R590, and R591) located on the front surface of gp5 with neutral asparagines abolishes the ability of gp5 and the helicase to mediate strand-displacement synthesis. This front basic patch in gp5 contributes to physical interactions with the acidic C-terminal tail of the helicase. Nonetheless, the altered polymerase is able to replace gp5 and continue ongoing strand-displacement synthesis. The results suggest that the interaction between the C-terminal tail of the helicase and the basic patch of gp5 is critical for initiation of strand-displacement synthesis. Multiple interactions of T7 DNA polymerase and helicase coordinate replisome movement.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Bacteriophage T7 / genetics
  • Bacteriophage T7 / metabolism*
  • Binding Sites / genetics
  • DNA Helicases / chemistry
  • DNA Helicases / genetics
  • DNA Helicases / metabolism*
  • DNA Replication*
  • DNA, Single-Stranded / chemistry
  • DNA, Single-Stranded / genetics
  • DNA, Single-Stranded / metabolism
  • DNA, Viral / chemistry
  • DNA, Viral / genetics
  • DNA, Viral / metabolism
  • DNA-Directed DNA Polymerase / chemistry
  • DNA-Directed DNA Polymerase / genetics
  • DNA-Directed DNA Polymerase / metabolism
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Escherichia coli / virology
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / genetics
  • Escherichia coli Proteins / metabolism
  • Models, Molecular
  • Mutation
  • Nucleic Acid Conformation
  • Protein Binding
  • Protein Structure, Secondary
  • Protein Structure, Tertiary
  • Surface Plasmon Resonance
  • Thioredoxins / chemistry
  • Thioredoxins / genetics
  • Thioredoxins / metabolism
  • Viral Proteins / chemistry
  • Viral Proteins / genetics
  • Viral Proteins / metabolism*

Substances

  • DNA, Single-Stranded
  • DNA, Viral
  • Escherichia coli Proteins
  • Viral Proteins
  • Thioredoxins
  • DNA-Directed DNA Polymerase
  • DNA Helicases