Targeting Holliday junctions by the RecG branch migration protein of Escherichia coli

J Biol Chem. 1998 Jul 31;273(31):19729-39. doi: 10.1074/jbc.273.31.19729.


The RecG protein of Escherichia coli is a junction-specific DNA helicase that drives branch migration of Holliday intermediates in genetic recombination and DNA repair. The reaction was investigated using synthetic X-junctions. RecG dissociates X-junctions to flayed duplex products, although DNA unwinding of the heterologous arms is limited to </=30 base pairs. Junction unwinding requires Mg2+ and the hydrolysis of ATP. X-junction DNA stimulates the ATPase activity of RecG. ATPase activity is also stimulated by linear duplex DNA, although to a lesser extent than by X-DNA, but not by linear single-stranded DNA. In situ 1,10-phenanthroline-copper footprinting shows that RecG binds to the strand cross-over point at the center of the X-junction. Substrate recognition by RecG was investigated using DNAs that represented the various component parts of an X-junction. The minimal DNA structure that RecG forms a stable complex with is a flayed duplex, suggesting that this is the critical feature for junction recognition by RecG. Junction binding and unwinding also depend critically on the concentration of free Mg2+, excess free cation dramatically inhibiting both processes. These inhibitory effects are not mediated specifically by Mg2+; e.g. both Ca2+ and hexamminecobalt(III) chloride also inhibit X-junction binding and unwinding by RecG. The relative abilities of these cations to inhibit RecG-junction binding is correlated with their respective abilities to stack X-junction DNA. From this we conclude that RecG is unable to bind or binds very poorly to fully stacked X-junctions.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenosine Triphosphate / metabolism
  • Bacterial Proteins / physiology*
  • Chlorides / metabolism
  • Cobalt / metabolism
  • DNA / chemistry*
  • DNA Footprinting
  • DNA Helicases / metabolism
  • DNA-Binding Proteins / metabolism
  • Escherichia coli / physiology*
  • Escherichia coli Proteins*
  • Magnesium / pharmacology
  • Phenanthrolines / metabolism
  • Recombination, Genetic / genetics*


  • Bacterial Proteins
  • Chlorides
  • DNA-Binding Proteins
  • Escherichia coli Proteins
  • Phenanthrolines
  • hexaminecobalt (III) trichloride
  • RecG protein, E coli
  • Cobalt
  • Adenosine Triphosphate
  • DNA
  • DNA Helicases
  • Magnesium
  • 1,10-phenanthroline