Coordination between the polymerase and 5'-nuclease components of DNA polymerase I of Escherichia coli

J Biol Chem. 2000 Jul 7;275(27):20949-55. doi: 10.1074/jbc.M909135199.


The polymerase and 5'-nuclease components of DNA polymerase I must collaborate in vivo so as to generate ligatable structures. Footprinting shows that the polymerase and 5'-nuclease cannot bind simultaneously to a DNA substrate and appear to compete with one another, suggesting that the two active sites are physically separate and operate independently. The desired biological end point, a ligatable nick, results from the substrate specificities of the polymerase and 5'-nuclease. The preferred substrate of the 5'-nuclease is a "double-flap" structure having a frayed base at the primer terminus overlapping the displaced strand that is to be cleaved by the 5'-nuclease. Cleavage of this structure occurs almost exclusively between the first two paired bases of the downstream strand, yielding a ligatable nick. In whole DNA polymerase I, the polymerase and 5'-nuclease activities are coupled such that the majority of molecules cleaved by the 5'-nuclease have also undergone polymerase-catalyzed addition to the primer terminus. This implies that the 5'-nuclease can capture a DNA molecule from the polymerase site more efficiently than from the bulk solution.

Publication types

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

MeSH terms

  • Bacterial Proteins / metabolism
  • Binding Sites
  • DNA / metabolism
  • DNA Footprinting
  • DNA Polymerase I / metabolism*
  • Escherichia coli / enzymology*
  • Kinetics
  • Multienzyme Complexes / metabolism*
  • Oligodeoxyribonucleotides / metabolism
  • Substrate Specificity


  • Bacterial Proteins
  • Multienzyme Complexes
  • Oligodeoxyribonucleotides
  • DNA
  • DNA Polymerase I