CoChat
Sign Up
Cited by 22DNA Repair MechanismsDNA and Nucleic Acid ChemistryBiochemical and Molecular Research

Significant contribution of the 3′→5′ exonuclease activity to the high fidelity of nucleotide incorporation catalyzed by human DNA polymerase ϵ

Open access
Zahurancik, Walter J., Klein, Seth J., Suo, Zucai·2014·Nucleic Acids Research
DOI 10.1093/nar/gku1184PMC PMC4267634
Open this paper in CoChat — free account

Abstract

Most eukaryotic DNA replication is performed by A- and B-family DNA polymerases which possess a faithful polymerase activity that preferentially incorporates correct over incorrect nucleotides. Additionally, many replicative polymerases have an efficient 3'→5' exonuclease activity that excises misincorporated nucleotides. Together, these activities contribute to overall low polymerase error frequency (one error per 10(6)-10(8) incorporations) and support faithful eukaryotic genome replication. Eukaryotic DNA polymerase ϵ (Polϵ) is one of three main replicative DNA polymerases for nuclear genomic replication and is responsible for leading strand synthesis. Here, we employed pre-steady-state kinetic methods and determined the overall fidelity of human Polϵ (hPolϵ) by measuring the individual contributions of its polymerase and 3'→5' exonuclease activities. The polymerase activity of hPolϵ has a high base substitution fidelity (10(-4)-10(-7)) resulting from large decreases in both nucleotide incorporation rate constants and ground-state binding affinities for incorrect relative to correct nucleotides. The 3'→5' exonuclease activity of hPolϵ further enhances polymerization fidelity by an unprecedented 3.5 × 10(2) to 1.2 × 10(4)-fold. The resulting overall fidelity of hPolϵ (10(-6)-10(-11)) justifies hPolϵ to be a primary enzyme to replicate human nuclear genome (0.1-1.0 error per round). Consistently, somatic mutations in hPolϵ, which decrease its exonuclease activity, are connected with mutator phenotypes and cancer formation.

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1093/nar/gku1184

Cites

4 of 4 references verified0 citation errors · 0 not checkable

Statistics

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Cites

Flagged first, then by citations

Research works printed but not linked: 43

  1. [1]Davies J.F. 2nd Almassy R.J. Hostomska Z. Ferre R.A. Hostomsky Z. 2.3 A crystal structure of the catalytic domain of DNA polymerase beta Cell 1994 76 1123 1133 8137427
  2. [3]Wang J. Sattar A.K. Wang C.C. Karam J.D. Konigsberg W.H. Steitz T.A. Crystal structure of a pol alpha family replication DNA polymerase from bacteriophage RB69 Cell 1997 89 1087 1099 9215631
  3. [4]Doublie S. Tabor S. Long A.M. Richardson C.C. Ellenberger T. Crystal structure of a bacteriophage T7 DNA replication complex at 2.2 A resolution Nature 1998 391 251 258 9440688
  4. [5]Freemont P.S. Friedman J.M. Beese L.S. Sanderson M.R. Steitz T.A. Cocrystal structure of an editing complex of Klenow fragment with DNA Proc. Natl. Acad. Sci. U.S.A. 1988 85 8924 8928 3194400 PMC282619
  5. [6]Savino C. Federici L. Johnson K.A. Vallone B. Nastopoulos V. Rossi M. Pisani F.M. Tsernoglou D. Insights into DNA replication: the crystal structure of DNA polymerase B1 from the archaeon Sulfolobus solfataricus Structure 2004 12 2001 2008 15530364
  6. [7]Beese L.S. Steitz T.A. Structural basis for the 3’-5’ exonuclease activity of Escherichia coli DNA polymerase I: a two metal ion mechanism EMBO J. 1991 10 25 33 1989886 PMC452607
  7. [8]Capson T.L. Peliska J.A. Kaboord B.F. Frey M.W. Lively C. Dahlberg M. Benkovic S.J. Kinetic characterization of the polymerase and exonuclease activities of the gene 43 protein of bacteriophage T4 Biochemistry 1992 31 10984 10994 1332748
  8. [10]Zhang H. Rhee C. Bebenek A. Drake J.W. Wang J. Konigsberg W. The L561A substitution in the nascent base-pair binding pocket of RB69 DNA polymerase reduces base discrimination Biochemistry 2006 45 2211 2220 16475809 PMC3373012
  9. [11]Zhang L. Brown J.A. Newmister S.A. Suo Z. Polymerization fidelity of a replicative DNA polymerase from the hyperthermophilic archaeon Sulfolobus solfataricus P2 Biochemistry 2009 48 7492 7501 19456141
  10. [12]Dieckman L.M. Johnson R.E. Prakash S. Washington M.T. Pre-steady state kinetic studies of the fidelity of nucleotide incorporation by yeast DNA polymerase delta Biochemistry 2010 49 7344 7350 20666462 PMC2941984
  11. [13]Lee H.R. Johnson K.A. Fidelity of the human mitochondrial DNA polymerase J. Biol. Chem. 2006 281 36236 36240 17005554
  12. [14]Wong I. Patel S.S. Johnson K.A. An induced-fit kinetic mechanism for DNA replication fidelity: direct measurement by single-turnover kinetics Biochemistry 1991 30 526 537 1846299
  13. [15]Loeb L.A. Kunkel T.A. Fidelity of DNA synthesis Annu. Rev. Biochem. 1982 51 429 457 6214209
  14. [16]Olson A.C. Patro J.N. Urban M. Kuchta R.D. The energetic difference between synthesis of correct and incorrect base pairs accounts for highly accurate DNA replication J. Am. Chem. Soc. 2013 135 1205 1208 23316816 PMC3561758
  15. [17]Kool E.T. Hydrogen bonding, base stacking, and steric effects in dna replication Annu. Rev. Biophys. Biomol. Struct. 2001 30 1 22 11340050
  16. [18]Petruska J. Sowers L.C. Goodman M.F. Comparison of nucleotide interactions in water, proteins, and vacuum: model for DNA polymerase fidelity Proc. Natl. Acad. Sci. U.S.A. 1986 83 1559 1562 3456600 PMC323122
  17. [19]Donlin M.J. Patel S.S. Johnson K.A. Kinetic partitioning between the exonuclease and polymerase sites in DNA error correction Biochemistry 1991 30 538 546 1988042
  18. [20]Johnson A.A. Johnson K.A. Exonuclease proofreading by human mitochondrial DNA polymerase J. Biol. Chem. 2001 276 38097 38107 11477094
  19. [21]Brieba L.G. Template dependent human DNA polymerases Curr. Top Med. Chem. 2008 8 1312 1326 18991720
  20. [22]Bermudez V.P. Farina A. Raghavan V. Tappin I. Hurwitz J. Studies on human DNA polymerase epsilon and GINS complex and their role in DNA replication J. Biol. Chem. 2011 286 28963 28977 21705323 PMC3190704
  21. [23]Hogg M. Osterman P. Bylund G.O. Ganai R.A. Lundstrom E.B. Sauer-Eriksson A.E. Johansson E. Structural basis for processive DNA synthesis by yeast DNA polymerase epsilon Nat. Struct. Mol. Biol. 2014 21 49 55 24292646
  22. [24]Jain R. Rajashankar K.R. Buku A. Johnson R.E. Prakash L. Prakash S. Aggarwal A.K. Crystal structure of yeast DNA polymerase epsilon catalytic domain PLoS One 2014 9 e94835 24733111 PMC3986358
  23. [25]McElhinny S.A.N. Gordenin D.A. Stith C.M. Burgers P.M.J. Kunkel T.A. Division of labor at the eukaryotic replication fork Mol. Cell 2008 30 137 144 18439893 PMC2654179
  24. [26]Pursell Z.F. Isoz I. Lundstrom E.B. Johansson E. Kunkel T.A. Yeast DNA polymerase epsilon participates in leading-strand DNA replication Science 2007 317 127 130 17615360 PMC2233713
  25. [27]Shinbrot E. Henninger E.E. Weinhold N. Covington K.R. Goksenin A.Y. Schultz N. Chao H. Doddapaneni H. Muzny D.M. Gibbs R.A. Exonuclease mutations In DNA Polymerase Epsilon reveal replication strand specific mutation patterns and human origins of replication Genome Res. 2014 1740 1750 PMC4216916 25228659
  26. [28]Miyabe I. Kunkel T.A. Carr A.M. The major roles of DNA polymerases epsilon and delta at the eukaryotic replication fork are evolutionarily conserved PLoS Genet. 2011 7 e1002407 22144917 PMC3228825
  27. [29]Zahurancik W.J. Klein S.J. Suo Z. Kinetic mechanism of DNA polymerization catalyzed by human DNA polymerase epsilon Biochemistry 2013 52 7041 7049 24020356 PMC3859840
  28. [30]Brown J.A. Suo Z. Elucidating the kinetic mechanism of DNA polymerization catalyzed by Sulfolobus solfataricus P2 DNA polymerase B1 Biochemistry 2009 48 7502 7511 19456143
  29. [31]Dahlberg M.E. Benkovic S.J. Kinetic mechanism of DNA polymerase I (Klenow fragment): identification of a second conformational change and evaluation of the internal equilibrium constant Biochemistry 1991 30 4835 4843 1645180
  30. [32]Fiala K.A. Suo Z. Mechanism of DNA polymerization catalyzed by Sulfolobus solfataricus P2 DNA polymerase IV Biochemistry 2004 43 2116 2125 14967051
  31. [33]Hsieh J.C. Zinnen S. Modrich P. Kinetic mechanism of the DNA-dependent DNA polymerase activity of human immunodeficiency virus reverse transcriptase J. Biol. Chem. 1993 268 24607 24613 7693703
  32. [34]Kuchta R.D. Mizrahi V. Benkovic P.A. Johnson K.A. Benkovic S.J. Kinetic mechanism of DNA polymerase I (Klenow) Biochemistry 1987 26 8410 8417 3327522
  33. [35]Patel S.S. Wong I. Johnson K.A. Pre-steady-state kinetic analysis of processive DNA replication including complete characterization of an exonuclease-deficient mutant Biochemistry 1991 30 511 525 1846298
  34. [36]Washington M.T. Prakash L. Prakash S. Yeast DNA polymerase eta utilizes an induced-fit mechanism of nucleotide incorporation Cell 2001 107 917 927 11779467
  35. [37]Korona D.A. Lecompte K.G. Pursell Z.F. The high fidelity and unique error signature of human DNA polymerase epsilon Nucleic Acids Res. 2010 39 1763 1773 21036870 PMC3061053
  36. [38]Fiala K.A. Suo Z. Pre-steady-state kinetic studies of the fidelity of Sulfolobus solfataricus P2 DNA polymerase IV Biochemistry 2004 43 2106 2115 14967050
  37. [39]Xia S. Konigsberg W.H. RB69 DNA polymerase structure, kinetics, and fidelity Biochemistry 2014 53 2752 2767 24720884 PMC4018061
  38. [40]Kuchta R.D. Benkovic P. Benkovic S.J. Kinetic mechanism whereby DNA polymerase I (Klenow) replicates DNA with high fidelity Biochemistry 1988 27 6716 6725 3058205
  39. [41]Loeb L.A. Mutator phenotype may be required for multistage carcinogenesis Cancer Res. 1991 51 3075 3079 2039987
  40. [42]McCulloch S.D. Kunkel T.A. The fidelity of DNA synthesis by eukaryotic replicative and translesion synthesis polymerases Cell Res. 2008 18 148 161 18166979 PMC3639319
  41. [44]Aksenova A. Volkov K. Maceluch J. Pursell Z.F. Rogozin I.B. Kunkel T.A. Pavlov Y.I. Johansson E. Mismatch repair-independent increase in spontaneous mutagenesis in yeast lacking non-essential subunits of DNA polymerase epsilon PLoS Genet. 2010 6 e1001209 21124948 PMC2987839
  42. [45]Lujan S.A. Williams J.S. Pursell Z.F. Abdulovic-Cui A.A. Clark A.B. Nick McElhinny S.A. Kunkel T.A. Mismatch repair balances leading and lagging strand DNA replication fidelity PLoS Genet. 2012 8 e1003016 23071460 PMC3469411
  43. [47]Umar A. Kunkel T.A. DNA-replication fidelity, mismatch repair and genome instability in cancer cells Eur. J. Biochem. 1996 238 297 307 8681938

Cited by

Sorted by citations

Authors
Zahurancik, W. J., Klein, S. J., & Suo, Z.
Affiliations
The Ohio State University (US)
Corresponding author
Suo, Zucai
ORCID
Venue
Nucleic Acids Research
Publisher
Oxford University Press (OUP)
Published
2014-11-20
Volume
42
Issue
22
Pages
13853-13860
ISSN
  • 0305-1048 (ISSN-L)
  • 1362-4962
DOI
10.1093/nar/gku1184
PMC
PMC4267634
OpenAlex
W2162935880
Cited by
22
References
4
Citations by year
2025: 1 · 2024: 2 · 2023: 1 · 2022: 4 · 2021: 1 · 2020: 4 · 2019: 1 · 2018: 3 · 2017: 1 · 2016: 1 · 2015: 2 · 2014: 1
FWCI
0.68
Type
journal article
Language
en
Open access
Licence
Locations
  • Nucleic Acids Research (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
Funders
  • National Institute of General Medical Sciences (T32GM008512)
  • National Institute of Environmental Health Sciences (ES009127)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA Repair Mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › DNA and Nucleic Acid Chemistry
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Biochemical and Molecular Research
Keywords
DNA polymerase epsilon, 3'-5' exonuclease activity, polymerase fidelity, nucleotide incorporation kinetics, misincorporation, leading strand synthesis, DNA replication, mutator phenotype, somatic mutations
MeSH terms
Poly-ADP-Ribose Binding Proteins, DNA, DNA Polymerase II, Exodeoxyribonucleases, Humans, Nucleotides, Substrate Specificity, Base Pair Mismatch, Biocatalysis

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review