skip to main content
OSTI.GOV title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Mechanism of Error-Free DNA Replication Past Lucidin-Derived DNA Damage by Human DNA Polymerase κ

Authors:
;  [1];  [2]; ; ;  [1]; ORCiD logo [2]; ORCiD logo
  1. Department of Biochemistry, Schulich School of Medicine &, Dentistry, University of Western Ontario, London, Ontario N6A 5C1, Canada
  2. Department of Chemistry, Indian Institute of Technology Bombay, Mumbai 400076, India
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
UNIVERSITYNIHFOREIGN
OSTI Identifier:
1411784
Resource Type:
Journal Article
Resource Relation:
Journal Name: Chemical Research in Toxicology; Journal Volume: 30; Journal Issue: 11
Country of Publication:
United States
Language:
ENGLISH

Citation Formats

Yockey, Oliver P., Jha, Vikash, Ghodke, Pratibha P., Xu, Tianzuo, Xu, Wenyan, Ling, Hong, Pradeepkumar, P. I., and Zhao, Linlin. Mechanism of Error-Free DNA Replication Past Lucidin-Derived DNA Damage by Human DNA Polymerase κ. United States: N. p., 2017. Web. doi:10.1021/acs.chemrestox.7b00227.
Yockey, Oliver P., Jha, Vikash, Ghodke, Pratibha P., Xu, Tianzuo, Xu, Wenyan, Ling, Hong, Pradeepkumar, P. I., & Zhao, Linlin. Mechanism of Error-Free DNA Replication Past Lucidin-Derived DNA Damage by Human DNA Polymerase κ. United States. doi:10.1021/acs.chemrestox.7b00227.
Yockey, Oliver P., Jha, Vikash, Ghodke, Pratibha P., Xu, Tianzuo, Xu, Wenyan, Ling, Hong, Pradeepkumar, P. I., and Zhao, Linlin. 2017. "Mechanism of Error-Free DNA Replication Past Lucidin-Derived DNA Damage by Human DNA Polymerase κ". United States. doi:10.1021/acs.chemrestox.7b00227.
@article{osti_1411784,
title = {Mechanism of Error-Free DNA Replication Past Lucidin-Derived DNA Damage by Human DNA Polymerase κ},
author = {Yockey, Oliver P. and Jha, Vikash and Ghodke, Pratibha P. and Xu, Tianzuo and Xu, Wenyan and Ling, Hong and Pradeepkumar, P. I. and Zhao, Linlin},
abstractNote = {},
doi = {10.1021/acs.chemrestox.7b00227},
journal = {Chemical Research in Toxicology},
number = 11,
volume = 30,
place = {United States},
year = 2017,
month =
}
  • N1-methyl-deoxyadenosine (1-MeA) is formed by methylation of deoxyadenosine at the N1 atom. 1-MeA presents a block to replicative DNA polymerases due to its inability to participate in Watson-Crick (W-C) base pairing. Here we determine how human DNA polymerase-ι (Polι) promotes error-free replication across 1-MeA. Steady state kinetic analyses indicate that Polι is ~100 fold more efficient in incorporating the correct nucleotide T versus the incorrect nucleotide C opposite 1-MeA. To understand the basis of this selectivity, we determined ternary structures of Polι bound to template 1-MeA and incoming dTTP or dCTP. In both structures, template 1-MeA rotates to the synmore » conformation but pairs differently with dTTP versus dCTP. Thus, whereas dTTP partakes in stable Hoogsteen base pairing with 1-MeA, dCTP fails to gain a “foothold” and is largely disordered. Together, our kinetic and structural studies show how Polι maintains discrimination between correct and incorrect incoming nucleotide opposite 1-MeA in preserving genome integrity.« less
  • 7,8-dihydro-8-oxoguanine (8-oxoG) adducts are formed frequently by the attack of oxygen-free radicals on DNA. They are among the most mutagenic lesions in cells because of their dual coding potential, where, in addition to normal base-pairing of 8-oxoG(anti) with dCTP, 8-oxoG in the syn conformation can base pair with dATP, causing G to T transversions. We provide here for the first time a structural basis for the error-free replication of 8-oxoG lesions by yeast DNA polymerase {eta} (Pol{eta}). We show that the open active site cleft of Pol{eta} can accommodate an 8-oxoG lesion in the anti conformation with only minimal changesmore » to the polymerase and the bound DNA: at both the insertion and post-insertion steps of lesion bypass. Importantly, the active site geometry remains the same as in the undamaged complex and provides a basis for the ability of Pol to prevent the mutagenic replication of 8-oxoG lesions in cells.« less
  • Human DNA polymerase iota (pol iota) is a unique member of Y-family polymerases, which preferentially misincorporates nucleotides opposite thymines (T) and halts replication at T bases. The structural basis of the high error rates remains elusive. We present three crystal structures of pol complexed with DNA containing a thymine base, paired with correct or incorrect incoming nucleotides. A narrowed active site supports a pyrimidine to pyrimidine mismatch and excludes Watson-Crick base pairing by pol. The template thymine remains in an anti conformation irrespective of incoming nucleotides. Incoming ddATP adopts a syn conformation with reduced base stacking, whereas incorrect dGTP andmore » dTTP maintain anti conformations with normal base stacking. Further stabilization of dGTP by H-bonding with Gln59 of the finger domain explains the preferential T to G mismatch. A template 'U-turn' is stabilized by pol and the methyl group of the thymine template, revealing the structural basis of T stalling. Our structural and domain-swapping experiments indicate that the finger domain is responsible for pol's high error rates on pyrimidines and determines the incorporation specificity.« less
  • The aromatic amine carcinogen 2-aminofluorene (AF) forms covalent adducts with DNA, predominantly with guanine at the C8 position. Such lesions are bypassed by Y-family polymerases such as Dpo4 via error-free and error-prone mechanisms. We show that Dpo4 catalyzes elongation from a correct 3{prime}-terminal cytosine opposite [AF]G in a nonrepetitive template sequence with low efficiency. This extension leads to cognate full-length product, as well as mis-elongated products containing base mutations and deletions. Crystal structures of the Dpo4 ternary complex, with the 3{prime}-terminal primer cytosine base opposite [AF]G in the anti conformation and with the AF moiety positioned in the major groove,more » reveal both accurate and misalignment-mediated mutagenic extension pathways. The mutagenic template-primer-dNTP arrangement is promoted by interactions between the polymerase and the bulky lesion rather than by a base pair-stabilized misaligment. Further extension leads to semitargeted mutations via this proposed polymerase-guided mechanism.« less
  • Human polymerase kappa (hPol {kappa}) is one of four eukaryotic Y-class DNA polymerases and may be an important element in the cellular response to polycyclic aromatic hydrocarbons such as benzo[a]pyrene, which can lead to reactive oxygenated metabolite-mediated oxidative stress. Here, we present a detailed analysis of the activity and specificity of hPol {kappa} bypass opposite the major oxidative adduct 7,8-dihydro-8-oxo-2{prime}-deoxyguanosine (8-oxoG). Unlike its archaeal homolog Dpo4, hPol {kappa} bypasses this lesion in an error-prone fashion by inserting mainly dATP. Analysis of transient-state kinetics shows diminished 'bursts' for dATP:8-oxoG and dCTP:8-oxoG incorporation, indicative of non-productive complex formation, but dATP:8-oxoG insertion eventsmore » that do occur are 2-fold more efficient than dCTP:G insertion events. Crystal structures of ternary hPol {kappa} complexes with adducted template-primer DNA reveal non-productive (dGTP and dATP) alignments of incoming nucleotide and 8-oxoG. Structural limitations placed upon the hPol {kappa} by interactions between the N-clasp and finger domains combined with stabilization of the syn-oriented template 8-oxoG through the side chain of Met-135 both appear to contribute to error-prone bypass. Mutating Leu-508 in the little finger domain of hPol {kappa} to lysine modulates the insertion opposite 8-oxoG toward more accurate bypass, similar to previous findings with Dpo4. Our structural and activity data provide insight into important mechanistic aspects of error-prone bypass of 8-oxoG by hPol {kappa} compared with accurate and efficient bypass of the lesion by Dpo4 and polymerase {eta}.« less