Efficient bypass of a thymine-thymine dimer by yeast DNA polymerase, Polη

Science

Volumn 283, Issue 5404, 1999, Pages 1001-1004

  Johnson, Robert E ^a   Prakash, Satya ^a   Prakash, Louise ^a  

^a UNIVERSITY OF TEXAS MEDICAL BRANCH   (United States)

Author keywords

[No Author keywords available]

Indexed keywords

DIMER; DNA DIRECTED DNA POLYMERASE ETA; DNA POLYMERASE; NUCLEOTIDE; THYMINE DERIVATIVE; UNCLASSIFIED DRUG;

ARTICLE; DNA DAMAGE; DNA REPAIR; DNA REPLICATION; DNA SEQUENCE; GENE; NONHUMAN; PRIORITY JOURNAL; PROTEIN PROCESSING; SACCHAROMYCES CEREVISIAE; ULTRAVIOLET IRRADIATION;

ADENINE; DEOXYRIBONUCLEOTIDES; DNA DAMAGE; DNA REPAIR; DNA, FUNGAL; DNA-DIRECTED DNA POLYMERASE; ENZYME INHIBITORS; PYRIMIDINE DIMERS; RECOMBINANT FUSION PROTEINS; SACCHAROMYCES CEREVISIAE; TEMPLATES, GENETIC; ULTRAVIOLET RAYS;

EID: 0033548231     PISSN: 00368075     EISSN: None     Source Type: Journal    
DOI: 10.1126/science.283.5404.1001     Document Type: Article

References (23)

1. E. C. Friedberg, G. C. Walker, W. Siede, DNA Repair and Mutagenesis (American Society for Microbiology Press, Washington, DC, 1995).
2. S. Prakash, P. Sung, L. Prakash, Annu. Rev. Genet. 27, 33 (1993).
3. R. E. Johnson et al., Mol. Cell. Biol. 12, 3807 (1992).
4. J. P. McDonald, A. S. Levine, R. Woodgate, Genetics 147, 1557 (1997).
5. J. R. Nelson, C. W. Lawrence, D. C. Hinkle, Science 272, 1646 (1996).
6. _, Nature 382, 729 (1996).
7. R. E. Johnson, S. Prakash, L. Prakash, J. Biol. Chem. 269, 28259 (1994).
8. 4 (pH 7.4), 10% glycerol, and 10 mM β-mercaptoethanol] containing 1 M NaCl followed by a wash with 10 volumes of K buffer containing 50 mM KCl. GST-Rad30 was batch eluted twice with one column volume of 100 mM tris-HCl (pH 7.5) containing 100 mM NaCl, 0.01% NP-40, and 20 mM glutathione The eluate was then pooled and loaded directly onto a Mini-Q column (Pharmacia), washed with 10 column volumes of K buffer containing 50 mM KCl, and eluted with 10 column volumes of 50 to 500 mM KCl gradient. The CST-Rad30 fractions were equilibrated in K buffer containing 50 mM KCI by microconcentration in a Microcon-30 microconcentrator (Amicon, Beverly, MA). Aliqouts were frozen at -70°C.
9. 32P-labeled 44-nt oligomer primer N430ft 5′-CTTTTCCCAGTCACGACGATGCTCCGGTACTCCAGTGTAGGCAT-3′ or the 30-nt oligomer primer that contains the first 30 nt of N4309 was annealed to the 75-nt oligomer template 5′- AGCTACCATGCCTGCACGMGAGTTCGTATTATGCCTACACTGGAGTACCGGAGCATCGTCGTGACTGGGAAAAC- 3′, which contained or did not contain a cis-syn T-T dimer at the underlined position. A Molecular Dynamics STORM phosphoimager and the ImageQuant software were used for quantitation. Percent activity was determined from the number of nucleotides incorporated.
10. P. M. J. Burgers and G. A. Bauer, J. Biol. Chem. 263, 925 (1988).
11. A. F. Wahl et al., Biochemistry 25, 7821 (1986).
12. To generate the carboxyl-terminal truncation of Rad30, we introduced a + 1 frameshift mutation into the RAD30 gene at the unique Bgl II site at position +681 in relation to +1 A of the ATG start codon. This mutation interrupts the RAD30 reading frame at amino acid position 226 and adds 23 amino acids from an alternate reading frame, yielding a 55-kD CST-Rad30 carboxyl-terminus deleted protein.
13. Single-letter abbreviations for amino acids are as follows: D, Asp; E, Glu; F. Phe; I, Ile; S, Ser; and V, Val.
14. R. E. Johnson, S. Prakash, L. Prakash, unpublished results.
15. For sequencing, reactions were carried out as described (9) except that oligonucleotide primers were not labeled, the DNA substrate concentration was increased to 100 nM, and Rad30 concentration was increased to 40 nM. Reactions were carried out for 30 min and then stopped by the addition of 50 mM EDTA. DNA products were extracted with phenol: chloroform and precipitated with ethanol and 300 mM NaOAc. The DNA was resuspended in 10 mM tris-HCl (pH 7.5) and sequenced by the dideoxynudeotide method with sequenase (U.S. Biochemical Corporation, Cleveland, OH) and the radiolabeled oligonucleotide N4494: 5′-AGCTACCATGCCTGC-3′ that would anneal to the DNA strand synthesized by Rad30. This analysis allows for the detection of mutations with ∼95% accuracy.
16. M. Tang et al., Proc. Natl. Acad. Sci. U.S.A. 95, 9755 (1998); N. B. Reuven, G. Tomer, Z. Livneh, Mol. Cell 2, 191 (1998); M. F. Goodman, Nature 395, 221 (1998); G. C. Walker, Proc. Natl. Acad. Sci. U.S.A. 95, 10348 (1998).
17. M. Tang et al., Proc. Natl. Acad. Sci. U.S.A. 95, 9755 (1998); N. B. Reuven, G. Tomer, Z. Livneh, Mol. Cell 2, 191 (1998); M. F. Goodman, Nature 395, 221 (1998); G. C. Walker, Proc. Natl. Acad. Sci. U.S.A. 95, 10348 (1998).
18. M. Tang et al., Proc. Natl. Acad. Sci. U.S.A. 95, 9755 (1998); N. B. Reuven, G. Tomer, Z. Livneh, Mol. Cell 2, 191 (1998); M. F. Goodman, Nature 395, 221 (1998); G. C. Walker, Proc. Natl. Acad. Sci. U.S.A. 95, 10348 (1998).
19. M. Tang et al., Proc. Natl. Acad. Sci. U.S.A. 95, 9755 (1998); N. B. Reuven, G. Tomer, Z. Livneh, Mol. Cell 2, 191 (1998); M. F. Goodman, Nature 395, 221 (1998); G. C. Walker, Proc. Natl. Acad. Sci. U.S.A. 95, 10348 (1998).
20. D. L. Svoboda, L. P. Briley, J.-M. H. Vos, Cancer Res. 58, 2445 (1998).
21. Y.-C. Wang, V. M. Maher, J. J. McCormick, Proc. Natl. Acad. Sci. U.S.A. 88, 7810 (1991); Y.-C. Wang, V. M. Maher, D. L. Mitchell, J. J. McCormick, Mol. Cell. Biol. 13, 4276 (1993).
22. Y.-C. Wang, V. M. Maher, J. J. McCormick, Proc. Natl. Acad. Sci. U.S.A. 88, 7810 (1991); Y.-C. Wang, V. M. Maher, D. L. Mitchell, J. J. McCormick, Mol. Cell. Biol. 13, 4276 (1993).
23. We thank J. Stephen-Taylor for the cis-syn T-T dimer and P. Burgers for yeast Polδ and accessory proteins. Supported by NIH grant GM19261.