title: Second-site RecA-DNA interactions: lack of identical recognition.
abstract: The RecA protein plays crucial roles in recombination and repair. In vitro, it polymerizes on single-stranded DNA and promotes homologous recognition of duplex DNA and subsequent strand exchange. How the RecA filament recognizes homologous duplex DNA is not yet clear. Recent research has indicated the possibility of recognition between identical DNA strands in the RecA filament which may be involved in a triple-stranded structure prior to strand exchange. Here we address this type of recognition by the RecA filament with a variety of physical techniques. By a gel retardation assay, we find interaction of identical DNAs in RecA filaments to be strongly dependent on the DNA length. Fluorescence measurements (emission quenching and resonance energy transfer) show that two identical DNA strands do not make tight contacts in the RecA complex and are similar in magnitude to heterologous interactions. This conclusion is supported by caloriometric measurements, which show a large exothermic enthalpy change upon the recognition of complementary strands by the RecA filament, but not for binding of identical strands. Spectroscopic techniques, linear and circular dichroism, indicate that the complexes between RecA and pairs of either identical or complementary DNA strands still have rather similar overall structures. The present study thus reveals no significant interactions between identical single strands of DNA in the RecA filament in vitro.
authors.affiliation: Department of Physical Chemistry, Chalmers University of Technology, Gothenburg, Sweden. wittung@phc.chalmers.se
authors.forename: P
authors.initials: P
authors.lastname: Wittung
authors.forename: L R
authors.initials: LR
authors.lastname: Bazemore
authors.forename: M
authors.initials: M
authors.lastname: Takahashi
authors.forename: B
authors.initials: B
authors.lastname: Nordén
authors.forename: C
authors.initials: C
authors.lastname: Radding
mesh_terms: D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D011938:Rec A Recombinases / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013329:Structure-Activity Relationship
chemical_list: D004247:DNA; D011938:Rec A Recombinases