Force and ATP hydrolysis dependent regulation of RecA nucleoprotein filament by single-stranded DNA binding protein |
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| Authors: | Hongxia Fu Shimin Le Hu Chen K. Muniyappa Jie Yan |
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| Affiliation: | 1Mechanobiology Institute, National University of Singapore, 117411 Singapore, 2Department of Physics, National University of Singapore, 117542 Singapore, 3Centre for Bioimaging Sciences, National University of Singapore, 117546 Singapore and 4Department of Biochemistry, Indian Institute of Science, Bangalore 560012, India |
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| Abstract: | In Escherichia coli, the filament of RecA formed on single-stranded DNA (ssDNA) is essential for recombinational DNA repair. Although ssDNA-binding protein (SSB) plays a complicated role in RecA reactions in vivo, much of our understanding of the mechanism is based on RecA binding directly to ssDNA. Here we investigate the role of SSB in the regulation of RecA polymerization on ssDNA, based on the differential force responses of a single 576-nucleotide-long ssDNA associated with RecA and SSB. We find that SSB outcompetes higher concentrations of RecA, resulting in inhibition of RecA nucleation. In addition, we find that pre-formed RecA filaments de-polymerize at low force in an ATP hydrolysis- and SSB-dependent manner. At higher forces, re-polymerization takes place, which displaces SSB from ssDNA. These findings provide a physical picture of the competition between RecA and SSB under tension on the scale of the entire nucleoprotein SSB array, which have broad biological implications particularly with regard to competitive molecular binding. |
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