Structure and Cellular Dynamics of Deinococcus radiodurans Single-stranded DNA (ssDNA)-binding Protein (SSB)-DNA Complexes*
- Nicholas P. George‡,1,
- Khanh V. Ngo§,1,
- Sindhu Chitteni-Pattu§,
- Cédric A. Norais§¶,
- John R. Battista‖,
- Michael M. Cox§,2 and
- James L. Keck‡,3
- From the ‡Department of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, Madison, Wisconsin 53706,
- the §Department of Biochemistry, University of Wisconsin College of Agriculture and Life Sciences, Madison, Wisconsin 53706,
- the ‖Department of Biological Sciences, Louisiana State University, Baton Rouge, Louisiana 70803, and
- the ¶Laboratoire de Biochimie, UMR CNRS 7654, Département de Biologie, Ecole Polytechnique, 91128 Palaiseau Cedex, France
- ↵2 To whom correspondence may be addressed: Dept. of Biochemistry, University of Wisconsin College of Agriculture and Life Sciences, Madison, WI 53706. Tel.: 608-262-1181; E-mail: cox{at}biochem.wisc.edu.
- ↵3 To whom correspondence may be addressed: Dept. of Biomolecular Chemistry, University of Wisconsin School of Medicine and Public Health, 1135 Biochemical Sciences Bldg., 420 Henry Mall, Madison, WI 53706. Tel.: 608-263-1815; E-mail: jlkeck{at}wisc.edu.
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↵1 Both authors contributed equally to this work.
Abstract
The single-stranded DNA (ssDNA)-binding protein from the radiation-resistant bacterium Deinococcus radiodurans (DrSSB) functions as a homodimer in which each monomer contains two oligonucleotide-binding (OB) domains. This arrangement is exceedingly rare among bacterial SSBs, which typically form homotetramers of single-OB domain subunits. To better understand how this unusual structure influences the DNA binding and biological functions of DrSSB in D. radiodurans radiation resistance, we have examined the structure of DrSSB in complex with ssDNA and the DNA damage-dependent cellular dynamics of DrSSB. The x-ray crystal structure of the DrSSB-ssDNA complex shows that ssDNA binds to surfaces of DrSSB that are analogous to those mapped in homotetrameric SSBs, although there are distinct contacts in DrSSB that mediate species-specific ssDNA binding. Observations by electron microscopy reveal two salt-dependent ssDNA-binding modes for DrSSB that strongly resemble those of the homotetrameric Escherichia coli SSB, further supporting a shared overall DNA binding mechanism between the two classes of bacterial SSBs. In vivo, DrSSB levels are heavily induced following exposure to ionizing radiation. This accumulation is accompanied by dramatic time-dependent DrSSB cellular dynamics in which a single nucleoid-centric focus of DrSSB is observed within 1 h of irradiation but is dispersed by 3 h after irradiation. These kinetics parallel those of D. radiodurans postirradiation genome reconstitution, suggesting that DrSSB dynamics could play important organizational roles in DNA repair.
- DNA-binding Protein
- DNA Damage Response
- DNA Recombination
- DNA Repair
- X-ray Crystallography
- Ionizing Radiation Resistance
- Single-stranded DNA-binding Protein
Footnotes
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↵* This work was supported, in whole or in part, by National Institutes of Health Grants GM068061 (to J. L. K.) and GM32335 (to M. M. C.).
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This article contains supplemental Table 1 and Figs. 1 and 2.
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The atomic coordinates and structure factors (code 3UDG) have been deposited in the Protein Data Bank, Research Collaboratory for Structural Bioinformatics, Rutgers University, New Brunswick, NJ (http://www.rcsb.org/).
- Received March 29, 2012.
- Revision received May 3, 2012.
- © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.
