Skip to main content
Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1987 Sep;84(17):6045–6049. doi: 10.1073/pnas.84.17.6045

The RAD3 gene of Saccharomyces cerevisiae encodes a DNA-dependent ATPase.

P Sung, L Prakash, S Weber, S Prakash
PMCID: PMC299004  PMID: 2957691

Abstract

The RAD3 gene of the yeast Saccharomyces cerevisiae is required for excision repair of damaged DNA and for cell viability. A protein of approximately equal to 89 kDa was purified to near homogeneity from yeast strains harboring multicopy plasmids that overproduce RAD3 protein; this protein corresponds closely to the expected size of the RAD3 protein and cross-reacts with the antiserum raised against a truncated RAD3 protein produced in Escherichia coli. The purified RAD3 protein shows a single-stranded DNA-dependent ATPase activity that catalyzes hydrolysis of ATP to ADP and Pi. The ATPase activity was coincident with the RAD3 protein during purification and is inhibited by anti-RAD3 antibodies, indicating that the RAD3 gene encodes this activity.

Full text

PDF
6045

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Arai K., Kornberg A. Mechanism of dnaB protein action. III. Allosteric role of ATP in the alteration of DNA structure by dnaB protein in priming replication. J Biol Chem. 1981 May 25;256(10):5260–5266. [PubMed] [Google Scholar]
  2. Arai N., Arai K., Kornberg A. Complexes of Rep protein with ATP and DNA as a basis for helicase action. J Biol Chem. 1981 May 25;256(10):5287–5293. [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Higgins D. R., Prakash S., Reynolds P., Polakowska R., Weber S., Prakash L. Isolation and characterization of the RAD3 gene of Saccharomyces cerevisiae and inviability of rad3 deletion mutants. Proc Natl Acad Sci U S A. 1983 Sep;80(18):5680–5684. doi: 10.1073/pnas.80.18.5680. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Ito H., Fukuda Y., Murata K., Kimura A. Transformation of intact yeast cells treated with alkali cations. J Bacteriol. 1983 Jan;153(1):163–168. doi: 10.1128/jb.153.1.163-168.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  7. Madura K., Prakash S. Nucleotide sequence, transcript mapping, and regulation of the RAD2 gene of Saccharomyces cerevisiae. J Bacteriol. 1986 Jun;166(3):914–923. doi: 10.1128/jb.166.3.914-923.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Miller R. D., Prakash L., Prakash S. Defective excision of pyrimidine dimers and interstrand DNA crosslinks in rad7 and rad23 mutants of Saccharomyces cerevisiae. Mol Gen Genet. 1982;188(2):235–239. doi: 10.1007/BF00332681. [DOI] [PubMed] [Google Scholar]
  9. Miller R. D., Prakash L., Prakash S. Genetic control of excision of Saccharomyces cerevisiae interstrand DNA cross-links induced by psoralen plus near-UV light. Mol Cell Biol. 1982 Aug;2(8):939–948. doi: 10.1128/mcb.2.8.939. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Naumovski L., Chu G., Berg P., Friedberg E. C. RAD3 gene of Saccharomyces cerevisiae: nucleotide sequence of wild-type and mutant alleles, transcript mapping, and aspects of gene regulation. Mol Cell Biol. 1985 Jan;5(1):17–26. doi: 10.1128/mcb.5.1.17. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Naumovski L., Friedberg E. C. A DNA repair gene required for the incision of damaged DNA is essential for viability in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4818–4821. doi: 10.1073/pnas.80.15.4818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Perozzi G., Prakash S. RAD7 gene of Saccharomyces cerevisiae: transcripts, nucleotide sequence analysis, and functional relationship between the RAD7 and RAD23 gene products. Mol Cell Biol. 1986 May;6(5):1497–1507. doi: 10.1128/mcb.6.5.1497. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Plevani P., Badaracco G., Chang L. M. Purification and characterization of two forms of DNA-dependent ATPase from yeast. J Biol Chem. 1980 May 25;255(10):4957–4963. [PubMed] [Google Scholar]
  14. RANDERATH K., RANDERATH E. ION-EXCHANGE CHROMATOGRAPHY OF NUCLEOTIDES ON POLY-(ETHYLENEIMINE)-CELLULOSE THIN LAYERS. J Chromatogr. 1964 Oct;16:111–125. doi: 10.1016/s0021-9673(01)82445-6. [DOI] [PubMed] [Google Scholar]
  15. Reynolds P., Higgins D. R., Prakash L., Prakash S. The nucleotide sequence of the RAD3 gene of Saccharomyces cerevisiae: a potential adenine nucleotide binding amino acid sequence and a nonessential acidic carboxyl terminal region. Nucleic Acids Res. 1985 Apr 11;13(7):2357–2372. doi: 10.1093/nar/13.7.2357. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Reynolds P., Prakash L., Dumais D., Perozzi G., Prakash S. Nucleotide sequence of the RAD10 gene of Saccharomyces cerevisiae. EMBO J. 1985 Dec 16;4(13A):3549–3552. doi: 10.1002/j.1460-2075.1985.tb04115.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Reynolds R. J., Friedberg E. C. Molecular mechanisms of pyrimidine dimer excision in Saccharomyces cerevisiae: incision of ultraviolet-irradiated deoxyribonucleic acid in vivo. J Bacteriol. 1981 May;146(2):692–704. doi: 10.1128/jb.146.2.692-704.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Seeberg E., Steinum A. L. Purification and properties of the uvrA protein from Escherichia coli. Proc Natl Acad Sci U S A. 1982 Feb;79(4):988–992. doi: 10.1073/pnas.79.4.988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Sugino A., Higgins N. P., Brown P. O., Peebles C. L., Cozzarelli N. R. Energy coupling in DNA gyrase and the mechanism of action of novobiocin. Proc Natl Acad Sci U S A. 1978 Oct;75(10):4838–4842. doi: 10.1073/pnas.75.10.4838. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Sugino A., Ryu B. H., Sugino T., Naumovski L., Friedberg E. C. A new DNA-dependent ATPase which stimulates yeast DNA polymerase I and has DNA-unwinding activity. J Biol Chem. 1986 Sep 5;261(25):11744–11750. [PubMed] [Google Scholar]
  21. Wilcox D. R., Prakash L. Incision and postincision steps of pyrimidine dimer removal in excision-defective mutants of Saccharomyces cerevisiae. J Bacteriol. 1981 Nov;148(2):618–623. doi: 10.1128/jb.148.2.618-623.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wray W., Boulikas T., Wray V. P., Hancock R. Silver staining of proteins in polyacrylamide gels. Anal Biochem. 1981 Nov 15;118(1):197–203. doi: 10.1016/0003-2697(81)90179-2. [DOI] [PubMed] [Google Scholar]

Articles from Proceedings of the National Academy of Sciences of the United States of America are provided here courtesy of National Academy of Sciences

RESOURCES