PMID:7929380
| Citation |
Tsaneva, IR and West, SC (1994) Targeted versus non-targeted DNA helicase activity of the RuvA and RuvB proteins of Escherichia coli. J. Biol. Chem. 269:26552-8 |
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| Abstract |
The RuvA and RuvB proteins of Escherichia coli 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. |
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| Keywords |
Adenosine Triphosphatases/metabolism; Bacterial Proteins/metabolism; Base Sequence; DNA/metabolism; DNA Damage; DNA Helicases/metabolism; DNA-Binding Proteins/metabolism; Escherichia coli/enzymology; Escherichia coli Proteins; Molecular Sequence Data |
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