This Article
Right arrow Full Text
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrowReprints and Permissions
Right arrow Copyright Information
Right arrow Books from ASM Press
Right arrow MicrobeWorld
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Rebeil, R.
Right arrow Articles by Nicholson, W. L.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Rebeil, R.
Right arrow Articles by Nicholson, W. L.

 Previous Article  |  Next Article 

Journal of Bacteriology, September 1998, p. 4879-4885, Vol. 180, No. 18
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Spore Photoproduct Lyase from Bacillus subtilis Spores Is a Novel Iron-Sulfur DNA Repair Enzyme Which Shares Features with Proteins such as Class III Anaerobic Ribonucleotide Reductases and Pyruvate-Formate Lyases

Roberto Rebeil,1 Yubo Sun,2,dagger Lilian Chooback,2,Dagger Mario Pedraza-Reyes,3 Cynthia Kinsland,4 Tadhg P. Begley,4 and Wayne L. Nicholson1,*

Department of Veterinary Science and Microbiology, University of Arizona, Tucson, Arizona 857211; Department of Microbiology and Immunology, University of North Texas Health Science Center, Fort Worth, Texas 761072; Facultad de Ciencias Químicas, Instituto de Investigación en Biología Experimental, Universidad de Guanajuato, Guanajuato, México3; and Chemistry Department, Cornell University, Ithaca, New York 148534

Received 8 June 1998/Accepted 20 July 1998

The major photoproduct in UV-irradiated spore DNA is the unique thymine dimer 5-thyminyl-5,6-dihydrothymine, commonly referred to as spore photoproduct (SP). An important determinant of the high UV resistance of Bacillus subtilis spores is the accurate in situ reversal of SP during spore germination by the DNA repair enzyme SP lyase. To study the molecular aspects of SP lyase-mediated SP repair, the cloned B. subtilis splB gene was engineered to encode SP lyase with a molecular tag of six histidine residues at its amino terminus. The engineered six-His-tagged SP lyase expressed from the amyE locus restored UV resistance to spores of a UV-sensitive mutant B. subtilis strain carrying a deletion-insertion mutation which removed the entire splAB operon at its natural locus and was shown to repair SP in vivo during spore germination. The engineered SP lyase was purified both from dormant B. subtilis spores and from an Escherichia coli overexpression system by nickel-nitrilotriacetic acid (NTA) agarose affinity chromatography and was shown by Western blotting, UV-visible spectroscopy, and iron and acid-labile sulfide analysis to be a 41-kDa iron-sulfur (Fe-S) protein, consistent with its amino acid sequence homology to the 4Fe-4S clusters in anaerobic ribonucleotide reductases and pyruvate-formate lyases. SP lyase was capable of reversing SP from purified SP-containing DNA in an in vitro reaction either when present in a cell-free extract prepared from dormant spores or after purification on nickel-NTA agarose. SP lyase activity was dependent upon reducing conditions and addition of S-adenosylmethionine as a cofactor.

* Corresponding author. Mailing address: Department of Veterinary Science and Microbiology, Building 90, Room 102, University of Arizona, Tucson, AZ 85721. Phone: (520) 621-2157. Fax: (520) 621-6366. E-mail: WLN{at}u.arizona.edu.

dagger Present address: Department of Medicine, University of Miami, Coral Gables, FL 33124.

Dagger Present address: Department of Chemistry and Biochemistry, University of Oklahoma, Norman, OK 73019.

Journal of Bacteriology, September 1998, p. 4879-4885, Vol. 180, No. 18
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Chandor-Proust, A., Berteau, O., Douki, T., Gasparutto, D., Ollagnier-de-Choudens, S., Fontecave, M., Atta, M. (2008). DNA Repair and Free Radicals, New Insights into the Mechanism of Spore Photoproduct Lyase Revealed by Single Amino Acid Substitution. J. Biol. Chem. 283: 36361-36368 [Abstract] [Full Text]  
  • Vey, J. L., Yang, J., Li, M., Broderick, W. E., Broderick, J. B., Drennan, C. L. (2008). From the Cover: Structural basis for glycyl radical formation by pyruvate formate-lyase activating enzyme. Proc. Natl. Acad. Sci. USA 105: 16137-16141 [Abstract] [Full Text]  
  • Pieck, J. C., Hennecke, U., Pierik, A. J., Friedel, M. G., Carell, T. (2006). Characterization of a New Thermophilic Spore Photoproduct Lyase from Geobacillus stearothermophilus (SplG) with Defined Lesion Containing DNA Substrates. J. Biol. Chem. 281: 36317-36326 [Abstract] [Full Text]  
  • Chandor, A., Berteau, O., Douki, T., Gasparutto, D., Sanakis, Y., Ollagnier-de-Choudens, S., Atta, M., Fontecave, M. (2006). Dinucleotide Spore Photoproduct, a Minimal Substrate of the DNA Repair Spore Photoproduct Lyase Enzyme from Bacillus subtilis. J. Biol. Chem. 281: 26922-26931 [Abstract] [Full Text]  
  • Buis, J. M., Cheek, J., Kalliri, E., Broderick, J. B. (2006). Characterization of an Active Spore Photoproduct Lyase, a DNA Repair Enzyme in the Radical S-Adenosylmethionine Superfamily. J. Biol. Chem. 281: 25994-26003 [Abstract] [Full Text]  
  • Boon, E. M., Livingston, A. L., Chmiel, N. H., David, S. S., Barton, J. K. (2003). DNA-mediated charge transport for DNA repair. Proc. Natl. Acad. Sci. USA 100: 12543-12547 [Abstract] [Full Text]  
  • Rebeil, R., Nicholson, W. L. (2001). The subunit structure and catalytic mechanism of the Bacillus subtilis DNA repair enzyme spore photoproduct lyase. Proc. Natl. Acad. Sci. USA 10.1073/pnas.161278998v1 [Abstract] [Full Text]  
  • Sofia, H. J., Chen, G., Hetzler, B. G., Reyes-Spindola, J. F., Miller, N. E. (2001). Radical SAM, a novel protein superfamily linking unresolved steps in familiar biosynthetic pathways with radical mechanisms: functional characterization using new analysis and information visualization methods. Nucleic Acids Res 29: 1097-1106 [Abstract] [Full Text]  
  • Slieman, T. A., Rebeil, R., Nicholson, W. L. (2000). Spore Photoproduct (SP) Lyase from Bacillus subtilis Specifically Binds to and Cleaves SP (5-Thyminyl-5,6-Dihydrothymine) but Not Cyclobutane Pyrimidine Dimers in UV-Irradiated DNA. J. Bacteriol. 182: 6412-6417 [Abstract] [Full Text]  
  • Nicholson, W. L., Munakata, N., Horneck, G., Melosh, H. J., Setlow, P. (2000). Resistance of Bacillus Endospores to Extreme Terrestrial and Extraterrestrial Environments. Microbiol. Mol. Biol. Rev. 64: 548-572 [Abstract] [Full Text]  
  • Fajardo-Cavazos, P., Nicholson, W. L. (2000). The TRAP-Like SplA Protein Is a trans-Acting Negative Regulator of Spore Photoproduct Lyase Synthesis during Bacillus subtilis Sporulation. J. Bacteriol. 182: 555-560 [Abstract] [Full Text]  
  • Rebeil, R., Nicholson, W. L. (2001). The subunit structure and catalytic mechanism of the Bacillus subtilis DNA repair enzyme spore photoproduct lyase. Proc. Natl. Acad. Sci. USA 98: 9038-9043 [Abstract] [Full Text]  


Copyright © 2009 by the American Society for Microbiology.   For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»