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Nucleic Acids Res. 2013 Sep;41(16):7738-44. doi: 10.1093/nar/gkt570. Epub 2013 Jun 26.

Molecular mechanism of sequence-dependent stability of RecA filament.

Author information

1
Department of Physics and Interdisciplinary Program of Integrated Biotechnology, Sogang University, Seoul 121-742, Korea, Kavli Institute of NanoScience, Department of BioNanoScience, Delft University of Technology, 2628 CJ, Delft, The Netherlands, Department of Physics and Center for the Physics of Living Cells, Institute for Genomic Biology and Center for Biophysics and Computational Biology, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA and Howard Hughes Medical Institute, Urbana, IL 61801, USA.

Abstract

RecA is a DNA-dependent ATPase and mediates homologous recombination by first forming a filament on a single-stranded (ss) DNA. RecA binds preferentially to TGG repeat sequence, which resembles the recombination hot spot Chi (5'-GCTGGTGG-3') and is the most frequent pattern (GTG) of the codon usage in Escherichia coli. Because of the highly dynamic nature of RecA filament formation, which consists of filament nucleation, growth and shrinkage, we need experimental approaches that can resolve each of these processes separately to gain detailed insights into the molecular mechanism of sequence preference. By using a single-molecule fluorescence assay, we examined the effect of sequence on individual stages of nucleation, monomer binding and dissociation. We found that RecA does not recognize the Chi sequence as a nucleation site. In contrast, we observed that it is the reduced monomer dissociation that mainly determines the high filament stability on TGG repeats. This sequence dependence of monomer dissociation is well-correlated with that of ATP hydrolysis, suggesting that DNA sequence dictates filament stability through modulation of ATP hydrolysis.

PMID:
23804763
PMCID:
PMC3763553
DOI:
10.1093/nar/gkt570
[Indexed for MEDLINE]
Free PMC Article

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