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. 1997 Feb 15;25(4):719–726. doi: 10.1093/nar/25.4.719

Characterization of a RecA/RAD51 homologue from the hyperthermophilic archaeon Pyrococcus sp. KOD1.

N Rashid 1, M Morikawa 1, K Nagahisa 1, S Kanaya 1, T Imanaka 1
PMCID: PMC146504  PMID: 9016620

Abstract

The Pk-rec gene, encoding a RecA/RAD51 homologue from the hyperthermophilic archaeon Pyrococcussp. KOD1, was expressed in Escherichia coli. The recombinant Pk-REC was purified to homogeneity and was shown to be in a dimeric form. A striking property of the purified recombinant Pk-REC was the unusual DNase activity on both single- and double-stranded DNAs along with the ATPase activity. The reaction product of this DNase activity was mononucleotides. The optimum temperature and pH for the DNase activity were 60 degrees C and 8-8.5, respectively. In addition, the metal ion requirement for DNase activity was different from that for the ATPase activity. The protein exhibited no DNase activity in the presence of Zn2+ion, which was one of the most preferable divalent cations for ATPase activity. Another unique characteristic of the recombinant protein was that the reaction product of ATPase activity was AMP instead of ADP.Pk-REC may represent a common prototype of the RecA family proteins with high RecA-like activity.

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Selected References

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  1. Adzuma K., Ogawa T., Ogawa H. Primary structure of the RAD52 gene in Saccharomyces cerevisiae. Mol Cell Biol. 1984 Dec;4(12):2735–2744. doi: 10.1128/mcb.4.12.2735. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Alani E., Padmore R., Kleckner N. Analysis of wild-type and rad50 mutants of yeast suggests an intimate relationship between meiotic chromosome synapsis and recombination. Cell. 1990 May 4;61(3):419–436. doi: 10.1016/0092-8674(90)90524-i. [DOI] [PubMed] [Google Scholar]
  3. BOMAN H. G., KALETTA U. Chromatography of rattlesnake venom; a separation of three phosphodiesterases. Biochim Biophys Acta. 1957 Jun;24(3):619–631. doi: 10.1016/0006-3002(57)90256-1. [DOI] [PubMed] [Google Scholar]
  4. Basile G., Aker M., Mortimer R. K. Nucleotide sequence and transcriptional regulation of the yeast recombinational repair gene RAD51. Mol Cell Biol. 1992 Jul;12(7):3235–3246. doi: 10.1128/mcb.12.7.3235. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brown J. R., Doolittle W. F. Root of the universal tree of life based on ancient aminoacyl-tRNA synthetase gene duplications. Proc Natl Acad Sci U S A. 1995 Mar 28;92(7):2441–2445. doi: 10.1073/pnas.92.7.2441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bult C. J., White O., Olsen G. J., Zhou L., Fleischmann R. D., Sutton G. G., Blake J. A., FitzGerald L. M., Clayton R. A., Gocayne J. D. Complete genome sequence of the methanogenic archaeon, Methanococcus jannaschii. Science. 1996 Aug 23;273(5278):1058–1073. doi: 10.1126/science.273.5278.1058. [DOI] [PubMed] [Google Scholar]
  7. Clark A. J., Sandler S. J. Homologous genetic recombination: the pieces begin to fall into place. Crit Rev Microbiol. 1994;20(2):125–142. doi: 10.3109/10408419409113552. [DOI] [PubMed] [Google Scholar]
  8. Dutreix M., Burnett B., Bailone A., Radding C. M., Devoret R. A partially deficient mutant, recA1730, that fails to form normal nucleoprotein filaments. Mol Gen Genet. 1992 Apr;232(3):489–497. doi: 10.1007/BF00266254. [DOI] [PubMed] [Google Scholar]
  9. Friedberg E. C. Deoxyribonucleic acid repair in the yeast Saccharomyces cerevisiae. Microbiol Rev. 1988 Mar;52(1):70–102. doi: 10.1128/mr.52.1.70-102.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gogarten J. P., Kibak H., Dittrich P., Taiz L., Bowman E. J., Bowman B. J., Manolson M. F., Poole R. J., Date T., Oshima T. Evolution of the vacuolar H+-ATPase: implications for the origin of eukaryotes. Proc Natl Acad Sci U S A. 1989 Sep;86(17):6661–6665. doi: 10.1073/pnas.86.17.6661. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Iwabe N., Kuma K., Hasegawa M., Osawa S., Miyata T. Evolutionary relationship of archaebacteria, eubacteria, and eukaryotes inferred from phylogenetic trees of duplicated genes. Proc Natl Acad Sci U S A. 1989 Dec;86(23):9355–9359. doi: 10.1073/pnas.86.23.9355. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kowalczykowski S. C., Dixon D. A., Eggleston A. K., Lauder S. D., Rehrauer W. M. Biochemistry of homologous recombination in Escherichia coli. Microbiol Rev. 1994 Sep;58(3):401–465. doi: 10.1128/mr.58.3.401-465.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kowalczykowski S. C., Eggleston A. K. Homologous pairing and DNA strand-exchange proteins. Annu Rev Biochem. 1994;63:991–1043. doi: 10.1146/annurev.bi.63.070194.005015. [DOI] [PubMed] [Google Scholar]
  14. Marsh T. L., Reich C. I., Whitelock R. B., Olsen G. J. Transcription factor IID in the Archaea: sequences in the Thermococcus celer genome would encode a product closely related to the TATA-binding protein of eukaryotes. Proc Natl Acad Sci U S A. 1994 May 10;91(10):4180–4184. doi: 10.1073/pnas.91.10.4180. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Mikawa T., Masui R., Ogawa T., Ogawa H., Kuramitsu S. N-terminal 33 amino acid residues of Escherichia coli RecA protein contribute to its self-assembly. J Mol Biol. 1995 Jul 21;250(4):471–483. doi: 10.1006/jmbi.1995.0391. [DOI] [PubMed] [Google Scholar]
  16. Morikawa M., Izawa Y., Rashid N., Hoaki T., Imanaka T. Purification and characterization of a thermostable thiol protease from a newly isolated hyperthermophilic Pyrococcus sp. Appl Environ Microbiol. 1994 Dec;60(12):4559–4566. doi: 10.1128/aem.60.12.4559-4566.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Rashid N., Morikawa M., Imanaka T. A RecA/RAD51 homologue from a hyperthermophilic archaeon retains the major RecA domain only. Mol Gen Genet. 1996 Dec 13;253(3):397–400. doi: 10.1007/s004380050337. [DOI] [PubMed] [Google Scholar]
  18. Rashid N., Morikawa M., Imanaka T. An abnormally acidic TATA-binding protein from a hyperthermophilic archaeon. Gene. 1995 Dec 1;166(1):139–143. doi: 10.1016/0378-1119(95)00603-2. [DOI] [PubMed] [Google Scholar]
  19. Roca A. I., Cox M. M. The RecA protein: structure and function. Crit Rev Biochem Mol Biol. 1990;25(6):415–456. doi: 10.3109/10409239009090617. [DOI] [PubMed] [Google Scholar]
  20. Rowlands T., Baumann P., Jackson S. P. The TATA-binding protein: a general transcription factor in eukaryotes and archaebacteria. Science. 1994 May 27;264(5163):1326–1329. doi: 10.1126/science.8191287. [DOI] [PubMed] [Google Scholar]
  21. Sandler S. J., Satin L. H., Samra H. S., Clark A. J. recA-like genes from three archaean species with putative protein products similar to Rad51 and Dmc1 proteins of the yeast Saccharomyces cerevisiae. Nucleic Acids Res. 1996 Jun 1;24(11):2125–2132. doi: 10.1093/nar/24.11.2125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Shinohara A., Ogawa H., Ogawa T. Rad51 protein involved in repair and recombination in S. cerevisiae is a RecA-like protein. Cell. 1992 May 1;69(3):457–470. doi: 10.1016/0092-8674(92)90447-k. [DOI] [PubMed] [Google Scholar]
  23. Sung P. Catalysis of ATP-dependent homologous DNA pairing and strand exchange by yeast RAD51 protein. Science. 1994 Aug 26;265(5176):1241–1243. doi: 10.1126/science.8066464. [DOI] [PubMed] [Google Scholar]
  24. Tateishi S., Horii T., Ogawa T., Ogawa H. C-terminal truncated Escherichia coli RecA protein RecA5327 has enhanced binding affinities to single- and double-stranded DNAs. J Mol Biol. 1992 Jan 5;223(1):115–129. doi: 10.1016/0022-2836(92)90720-5. [DOI] [PubMed] [Google Scholar]

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