4.3 Article

Function and biochemical characterization of RecJ in Deinococcus radiodurans

Journal

DNA REPAIR
Volume 11, Issue 4, Pages 349-356

Publisher

ELSEVIER
DOI: 10.1016/j.dnarep.2011.11.008

Keywords

recJ; Deinococcus radiodurans; Manganese ion; Exonuclease

Funding

  1. National Natural Science Foundation of China [30830006, 30900031]
  2. Special Fund for Agroscientific Research in the Public Interest [201103007]
  3. China Postdoctoral Science Foundation [20090451465]
  4. [2009ZXJ09001-034]

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The single-stranded DNA-specific nuclease RecJ is found in most bacteria where it is involved in the Rec-FOR double-stranded break (DSBs) repair pathway. DSBs repair mainly occurs via the RecFOR pathway in Deinococcus radiodurans, a well-known radiation-resistant bacterium. A recJ null mutant was constructed to investigate the role of recJ in D. radiodurans. recJ inactivation caused growth defects and sensitivity to high temperatures. However, the radiation resistance of the recJ mutant was only moderately decreased. The full-length D. radiodurans RecJ (DrRecJ) protein was expressed and purified to further characterize its biochemical properties. DrRecJ possessed a Mn2+ concentration-dependent nuclease activity where the optimal Mn2+ concentration was 0.1 mM. DrRecJ had a similar activity profile after adding 10 mM Mg2+ to reactions with different Mn2+ concentrations, indicating that Mn2+ is a RecJ regulator. Escherichia coli RecJ has no activity on 5' ssDNA tails shorter than 6-nt, but DrRecJ could effectively degrade DNA with a 4-nt 5' ssDNA tail, suggesting that DrRecJ may have a wider range of DNA substrates. Moreover, SSB in D. radiodurans stimulated the DrRecJ exonuclease activity, whereas DdrB inhibited it and provided protection to ssDNA. Overall, our results indicate that recJ is a nonessential gene in D. radiodurans and that the activity of DrRecJ is regulated by Mn2+ and SSB-DdrB. (C) 2012 Elsevier B.V. All rights reserved.

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