TARGETED VERSUS NON-TARGETED DNA HELICASE ACTIVITY OF THE RUVA AND RUVB PROTEINS OF ESCHERICHIA-COLI.
J BIOL CHEM
26552 - 26558.
The RuvA and RuvB proteins of Escherichia coil promote the branch migration of Holliday junctions in vitro. To understand the relationship between branch migration and the intrinsic 5' --> 3' DNA helicase activity of RuvAB, the requirements and substrate specificity of the helicase reaction have been studied in more detail. We find that RuvAB-mediated DNA unwinding and branch migration reactions show similar requirements for Mg2+ and ATP and are inhibited to a similar extent by ADP and ATP gamma S (adenosine 5'-O-(3-thiotriphosphate)). The helicase activity, measured by the dissociation of a short fragment from circular single-stranded DNA, requires both RuvA and RuvB and is stimulated by subsaturating concentrations of single-strand binding protein (SSB). In contrast, saturating concentrations of SSB are inhibitory. Using substrates that contain a DNA junction, which permits the specific binding of RuvA, we find that the RuvA and RuvB proteins promote two types of helicase reactions: nonspecific reactions, which are sensitive to inhibition by stoichiometric amounts of SSB, and junction-targeted reactions, which are not inhibited by SSB. Using three-armed structures, we observe that junction-targeted reactions display a polarity and result in asymmetric product formation. Junction-specific binding and the subsequent initiation of DNA unwinding are Likely to represent early steps in the process of branch migration.
|Title:||TARGETED VERSUS NON-TARGETED DNA HELICASE ACTIVITY OF THE RUVA AND RUVB PROTEINS OF ESCHERICHIA-COLI|
|Open access status:||An open access publication|
|Keywords:||TERMINATION FACTOR-RHO, SYNTHETIC HOLLIDAY JUNCTIONS, BRANCH MIGRATION, FUNCTIONAL INTERACTIONS, GENETIC-RECOMBINATION, RECA PROTEIN, K-12 REVEALS, REPAIR, RESOLUTION, INVITRO|
|UCL classification:||UCL > School of Life and Medical Sciences > Faculty of Life Sciences > Biosciences (Division of) > Structural and Molecular Biology|
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