Studies of the interaction of RecA protein with DNA

Dombroski, Dennise; Scraba, Douglas G.; Bradley, R.; Morgan, A R

doi:10.1093/nar/11.21.7487

Cited by 54 publications

(27 citation statements)

References 27 publications

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“…Pre-synapsis involves the assembly of RecA monomers on single stranded (ss) DNA in the presence of ATP with a stoichiometry of 3 nt per monomer (Di Capua et al, 1982; Dombroski et al, 1983). RecA forms a filament which stretches the ssDNA to a length of 1.5 times the length of B-form DNA (Stasiak et al, 1981; Dunn et al, 1982; Stasiak and Egelman, 1986).…”

Section: Introductionmentioning

confidence: 99%

RecA filament sliding on DNA facilitates homology search

Ragunathan

Liu

2012

eLife

100

113

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During homologous recombination, RecA forms a helical filament on a single stranded (ss) DNA that searches for a homologous double stranded (ds) DNA and catalyzes the exchange of complementary base pairs to form a new heteroduplex. Using single molecule fluorescence imaging tools with high spatiotemporal resolution we characterized the encounter complex between the RecA filament and dsDNA. We present evidence in support of the ‘sliding model’ wherein a RecA filament diffuses along a dsDNA track. We further show that homology can be detected during sliding. Sliding occurs with a diffusion coefficient of approximately 8000 bp2/s allowing the filament to sample several hundred base pairs before dissociation. Modeling suggests that sliding can accelerate homology search by as much as 200 fold. Homology recognition can occur for as few as 6 nt of complementary basepairs with the recognition efficiency increasing for higher complementarity. Our data represents the first example of a DNA bound multi-protein complex which can slide along another DNA to facilitate target search.DOI: http://dx.doi.org/10.7554/eLife.00067.001

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Section: Introductionmentioning

confidence: 99%

RecA filament sliding on DNA facilitates homology search

Ragunathan

Liu

2012

eLife

100

113

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“…Pre-synapsis involves the assembly of RecA monomers on ssDNA to form a helical filament in the presence of ATP. RecA monomers assemble on ssDNA with a stoichiometry of 3 nt per RecA monomer (Di Capua et al, 1982; Dombroski et al, 1983) and the resulting filament stretches the DNA to 1.5 times the length of B-form DNA (Dunn et al, 1982; Stasiak et al, 1981). These observations were recently confirmed by crystallographic studies of the RecA filament formed on single and double stranded DNA (Chen et al, 2008).…”

Section: Introductionmentioning

confidence: 99%

Real-Time Observation of Strand Exchange Reaction with High Spatiotemporal Resolution

Ragunathan

Joo

2011

Structure

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RecA binds to single stranded (ss) DNA to form a helical filament that catalyzes strand exchange with a homologous double stranded (ds) DNA. The study of strand exchange in ensemble assays is limited by the diffusion limited homology search process which masks the subsequent strand exchange reaction. We developed a single molecule fluorescence assay with a few basepair and milliseconds resolution which can separate initial docking from the subsequent propagation of joint molecule formation. Our data suggests that propagation occurs in 3 bp increments with destabilization of the incoming dsDNA and concomitant pairing with the reference ssDNA. Unexpectedly, we discovered the formation of a dynamic complex between RecA and the displaced DNA that remains bound transiently after joint molecule formation. This finding could have important implications for the irreversibility of strand exchange. Our model for strand exchange links structural models of RecA to its catalytic function.

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“…Previous work showed that the RecA protein binds dsDNA along its minor groove. 56 Because the heteroduplex DNA in the RecA ⅐ tsDNA complex exhibits an extended and unwound conformation indistinguishable from that of RecA-bound dsDNA in our FRET work, it is likely that RecA protein also binds to the heteroduplex along its minor groove. This inference is based on the theory that similar RecA-dsDNA interactions will produce similar conformational changes in the bound DNA molecule.…”

Section: Implications For the Structure Of The Strand Exchange Intermmentioning

confidence: 86%

“…56 One possible result of the stretched and underwound DNA structure imposed by RecA binding is to widen the minor groove and move its floor closer to the cylindrical surface contour of the DNA molecule. Such structural changes would likely provide for more facile interaction between the interior surface of the RecA protein filament and the exposed floor of the minor groove of DNA, as described for TATA boxbinding protein 57,58 and architectural proteins 59 binding their cognate dsDNA sequences.…”

Section: Implications For Reca-dna Filament Structurementioning

confidence: 99%

The stretched DNA geometry of recombination and repair nucleoprotein filaments

Singleton

Xiao

2002

Biopolymers

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The RecA protein of Escherichia coli plays essential roles in homologous recombination and restarting stalled DNA replication forks. In vitro, the protein mediates DNA strand exchange between single-stranded (ssDNA) and homologous double-stranded DNA (dsDNA) molecules that serves as a model system for the in vivo processes. To date, no high-resolution structure of the key intermediate, comprised of three DNA strands simultaneously bound to a RecA filament (RecA x tsDNA complex), has been elucidated by classical methods. Here we review the systematic characterization of the helical geometries of the three DNA strands of the RecA x tsDNA complex using fluorescence resonance energy transfer (FRET) under physiologically relevant solution conditions. Measurements of the helical parameters for the RecA x tsDNA complex are consistent with the hypothesis that this complex is a late, poststrand-exchange intermediate with the outgoing strand shifted by about three base pairs with respect to its registry with the incoming and complementary strands. All three strands in the RecA x tsDNA complex adopt extended and unwound conformations similar to those of RecA-bound ssDNA and dsDNA.

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Studies of the interaction of RecA protein with DNA

Cited by 54 publications

References 27 publications

RecA filament sliding on DNA facilitates homology search

RecA filament sliding on DNA facilitates homology search

Real-Time Observation of Strand Exchange Reaction with High Spatiotemporal Resolution

The stretched DNA geometry of recombination and repair nucleoprotein filaments

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