Inhibition of protein-mediated homologous pairing by a DNA helicase.
Protein-mediated exchange of homologous DNA strands is a central reaction in general genetic recombination and the mechanism by which proteins mediate this process in vivo is a topic of keen interest. The dda protein of the bacteriophage T4 is a DNA helicase that has been shown to accelerate branch migration catalyzed by the phage uvsX and gene 32 proteins in vitro (Kodadek, T., and Alberts, B.M. (1987) Nature 326, 312-314). This study did not address the potential role of the helicase in protein-mediated homologous pairing, the first phase of the overall strand-exchange reaction. It is shown here that the dda protein inhibits uvsX protein-mediated pairing between homologous single and double-stranded DNAs. Experiments using deproteinized heteroduplex joints demonstrate that the dda helicase is capable of unwinding these structures to some extent and suggests that this activity may be responsible for the observed inhibition of pairing. It is found that the helicase also reduces the level of uvsX protein-mediated, single-stranded DNA-dependent ATP hydrolysis in the strand-exchange reactions, suggesting that the helicase may also act to destabilize the uvsX protein-DNA filaments that are important intermediates in the pairing reaction. Three other helicases are found to have no effect on the uvsX protein-mediated pairing reaction. A model rationalizing the ability of the dda protein to both inhibit homologous pairing and stimulate branch migration is presented and possible in vivo roles for this interesting activity are discussed.
Kodadek
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Department of Chemistry and Biochemistry, University of Texas, Austin 78712.
D000255:Adenosine Triphosphate / Q000378:metabolism; D002384:Catalysis; D004265:DNA Helicases / Q000037:antagonists & inhibitors*; D004264:DNA Topoisomerases, Type I / Q000235:genetics* / Q000378:metabolism; D004277:DNA, Single-Stranded / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D011938:Rec A Recombinases / Q000378:metabolism; D011995:Recombination, Genetic
D004277:DNA, Single-Stranded; D000255:Adenosine Triphosphate; D011938:Rec A Recombinases; D004265:DNA Helicases; D004264:DNA Topoisomerases, Type I