Related Experiment Video
Updated: Aug 7, 2026

Visualization of UV-induced Replication Intermediates in E. coli using Two-dimensional Agarose-gel Analysis
Published on: December 21, 2010
Targeting Holliday junctions by the RecG branch migration protein of Escherichia coli
1Microbiology Unit, Department of Biochemistry, University of Oxford, South Parks Road, Oxford OX1 3QU, United Kingdom. whitby@bioch.ox.ac.uk
Abstract:
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.
More Related Videos
14:06Mapping Bacterial Functional Networks and Pathways in Escherichia Coli using Synthetic Genetic Arrays
Published on: November 12, 2012
09:29Analysis of Nonhomologous End Joining and Homologous Recombination Efficiency in HEK-293T Cells Using GFP-Based Reporter Systems
Published on: February 2, 2024
Related Concept Videos
Mismatch Repair
Homologous Recombination
Conservative Site-specific Recombination and Phase Variation
The recognition sites for Cre recombinase called LoxP...
Nucleotide Excision Repair
Cells are regularly exposed to mutagens—factors in the environment that can damage DNA and generate mutations. UV radiation is one of the most common mutagens and is estimated to introduce a significant number of changes in DNA. These include bends or kinks in the structure, which can block DNA replication or transcription. If these errors are not fixed, the damage can cause mutations, which in turn can result in cancer or disease depending on which sequences are...
Mismatch Repair
The Mutator Protein Family Plays a Key Role in DNA Mismatch Repair
The human genome has more than 3 billion base pairs of DNA per cell. Prior to cell division, that vast amount of genetic...
Chemotaxis in E. coli