Skip to main content
PMC home page
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
. 1989 Nov;86(22):8877–8881. doi: 10.1073/pnas.86.22.8877

Escherichia coli mutY gene encodes an adenine glycosylase active on G-A mispairs.

K G Au 1, S Clark 1, J H Miller 1, P Modrich 1
PMCID: PMC298393  PMID: 2682664

Abstract

Mutations in the mutY gene of Escherichia coli confer hypermutability reflecting G.C to T.A transversion mutations and result in a deficiency in methyl-independent G-A to G.C mismatch correction. In the present work, the mutY product has been purified to near homogeneity by virtue of its ability to restore G-A to G.C mismatch correction to cell-free extracts of a mutS mutY strain. The 36-kDa protein renders the strand containing the mispaired adenine labile to base-catalyzed cleavage and sensitive to cleavage by several apurinic/apyrimidinic-site endonucleases, with the sites of strand scission by both agents corresponding to the location of the mismatch. These findings indicate that MutY is a DNA glycosylase that hydrolyzes the glycosyl bond linking the mis-paired adenine to deoxyribose. MutY, a 5'-apurinic/apyrimidinic-site endonuclease, DNA polymerase I, and DNA ligase are sufficient to reconstitute MutY-dependent G-A to G.C repair in vitro.

Full text

PDF
8877

Images in this article

8879Image
on p.8879
8880Image
on p.8880
8880Image
on p.8880
8881Image
on p.8881

Selected References

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

  1. Au K. G., Cabrera M., Miller J. H., Modrich P. Escherichia coli mutY gene product is required for specific A-G----C.G mismatch correction. Proc Natl Acad Sci U S A. 1988 Dec;85(23):9163–9166. doi: 10.1073/pnas.85.23.9163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bhatnagar S. K., Bessman M. J. Studies on the mutator gene, mutT of Escherichia coli. Molecular cloning of the gene, purification of the gene product, and identification of a novel nucleoside triphosphatase. J Biol Chem. 1988 Jun 25;263(18):8953–8957. [PubMed] [Google Scholar]
  3. Brown T., Hunter W. N., Kneale G., Kennard O. Molecular structure of the G.A base pair in DNA and its implications for the mechanism of transversion mutations. Proc Natl Acad Sci U S A. 1986 Apr;83(8):2402–2406. doi: 10.1073/pnas.83.8.2402. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cox E. C. Mutator gene studies in Escherichia coli: the mutT gene. Genetics. 1973 Apr;73(Suppl):67–80. [PubMed] [Google Scholar]
  5. Hunter W. N., Brown T., Anand N. N., Kennard O. Structure of an adenine-cytosine base pair in DNA and its implications for mismatch repair. Nature. 1986 Apr 10;320(6062):552–555. doi: 10.1038/320552a0. [DOI] [PubMed] [Google Scholar]
  6. Jones M., Wagner R., Radman M. Repair of a mismatch is influenced by the base composition of the surrounding nucleotide sequence. Genetics. 1987 Apr;115(4):605–610. doi: 10.1093/genetics/115.4.605. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Kan L. S., Chandrasegaran S., Pulford S. M., Miller P. S. Detection of a guanine X adenine base pair in a decadeoxyribonucleotide by proton magnetic resonance spectroscopy. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4263–4265. doi: 10.1073/pnas.80.14.4263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Kane C. M., Linn S. Purification and characterization of an apurinic/apyrimidinic endonuclease from HeLa cells. J Biol Chem. 1981 Apr 10;256(7):3405–3414. [PubMed] [Google Scholar]
  9. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  10. Laengle-Rouault F., Maenhaut-Michel G., Radman M. GATC sequence and mismatch repair in Escherichia coli. EMBO J. 1986 Aug;5(8):2009–2013. doi: 10.1002/j.1460-2075.1986.tb04457.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Lu A. L., Chang D. Y. A novel nucleotide excision repair for the conversion of an A/G mismatch to C/G base pair in E. coli. Cell. 1988 Sep 9;54(6):805–812. doi: 10.1016/s0092-8674(88)91109-9. [DOI] [PubMed] [Google Scholar]
  12. Lu A. L., Chang D. Y. Repair of single base-pair transversion mismatches of Escherichia coli in vitro: correction of certain A/G mismatches is independent of dam methylation and host mutHLS gene functions. Genetics. 1988 Apr;118(4):593–600. doi: 10.1093/genetics/118.4.593. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Lu A. L., Clark S., Modrich P. Methyl-directed repair of DNA base-pair mismatches in vitro. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4639–4643. doi: 10.1073/pnas.80.15.4639. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  15. Modrich P. DNA mismatch correction. Annu Rev Biochem. 1987;56:435–466. doi: 10.1146/annurev.bi.56.070187.002251. [DOI] [PubMed] [Google Scholar]
  16. Modrich P. Methyl-directed DNA mismatch correction. J Biol Chem. 1989 Apr 25;264(12):6597–6600. [PubMed] [Google Scholar]
  17. Nghiem Y., Cabrera M., Cupples C. G., Miller J. H. The mutY gene: a mutator locus in Escherichia coli that generates G.C----T.A transversions. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2709–2713. doi: 10.1073/pnas.85.8.2709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Patel D. J., Kozlowski S. A., Ikuta S., Itakura K. Deoxyguanosine-deoxyadenosine pairing in the d(C-G-A-G-A-A-T-T-C-G-C-G) duplex: conformation and dynamics at and adjacent to the dG X dA mismatch site. Biochemistry. 1984 Jul 3;23(14):3207–3217. doi: 10.1021/bi00309a015. [DOI] [PubMed] [Google Scholar]
  19. Radicella J. P., Clark E. A., Fox M. S. Some mismatch repair activities in Escherichia coli. Proc Natl Acad Sci U S A. 1988 Dec;85(24):9674–9678. doi: 10.1073/pnas.85.24.9674. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Radman M., Wagner R. Mismatch repair in Escherichia coli. Annu Rev Genet. 1986;20:523–538. doi: 10.1146/annurev.ge.20.120186.002515. [DOI] [PubMed] [Google Scholar]
  21. Schaaper R. M., Dunn R. L. Escherichia coli mutT mutator effect during in vitro DNA synthesis. Enhanced A.G replicational errors. J Biol Chem. 1987 Dec 5;262(34):16267–16270. [PubMed] [Google Scholar]
  22. Sowers L. C., Fazakerley G. V., Kim H., Dalton L., Goodman M. F. Variation of nonexchangeable proton resonance chemical shifts as a probe of aberrant base pair formation in DNA. Biochemistry. 1986 Jul 15;25(14):3983–3988. doi: 10.1021/bi00362a002. [DOI] [PubMed] [Google Scholar]
  23. Studier F. W. Analysis of bacteriophage T7 early RNAs and proteins on slab gels. J Mol Biol. 1973 Sep 15;79(2):237–248. doi: 10.1016/0022-2836(73)90003-x. [DOI] [PubMed] [Google Scholar]
  24. Su S. S., Lahue R. S., Au K. G., Modrich P. Mispair specificity of methyl-directed DNA mismatch correction in vitro. J Biol Chem. 1988 May 15;263(14):6829–6835. [PubMed] [Google Scholar]
  25. Su S. S., Modrich P. Escherichia coli mutS-encoded protein binds to mismatched DNA base pairs. Proc Natl Acad Sci U S A. 1986 Jul;83(14):5057–5061. doi: 10.1073/pnas.83.14.5057. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Wiebauer K., Jiricny J. In vitro correction of G.T mispairs to G.C pairs in nuclear extracts from human cells. Nature. 1989 May 18;339(6221):234–236. doi: 10.1038/339234a0. [DOI] [PubMed] [Google Scholar]
  27. Yajko D. M., Weiss B. Mutations simultaneously affecting endonuclease II and exonuclease III in Escherichia coli. Proc Natl Acad Sci U S A. 1975 Feb;72(2):688–692. doi: 10.1073/pnas.72.2.688. [DOI] [PMC free article] [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