Preferred WMSA catalytic mechanism of the nucleotidyl transfer reaction in human DNA polymerase κ elucidates error-free bypass of a bulky DNA lesion

Lee Lior-Hoffmann, Lihua Wang, Shenglong Wang, Nicholas E. Geacintov, Suse Broyde, Yingkai Zhang

Research output: Contribution to journalArticle

Abstract

Human DNA Pol i is a polymerase enzyme, specialized for near error-free bypass of certain bulky chemical lesions to DNA that are derived from environmental carcinogens present in tobacco smoke, automobile exhaust and cooked food. By employing ab initio QM/MM-MD (Quantum Mechanics/ Molecular Mechanics-Molecular Dynamics) simulations with umbrella sampling, we have determined the entire free energy profile of the nucleotidyl transfer reaction catalyzed by Pol κ and provided detailed mechanistic insights. Our results show that a variant of the Water Mediated and Substrate Assisted (WMSA) mechanism that we previously deduced for Dpo4 and T7 DNA polymerases is preferred for Pol κ as well, suggesting its broad applicability. The hydrogen on the 3'-OHprimer terminus is transferred through crystal and solvent waters to the γ-phosphate of the dNTP, followed by the associative nucleotidyl transfer reaction; this is facilitated by a proton transfer from the γ-phosphate to the α,β-bridging oxygen as pyrophosphate leaves, to neutralize the evolving negative charge. MD simulations show that the near error-free incorporation of dCTP opposite the major benzo[a]pyrene-derived dG lesion is compatible with the WMSA mechanism, allowing for an essentially undisturbed pentacovalent phosphorane transition state, and explaining the bypass of this lesion with little mutation by Pol κ.

Original languageEnglish (US)
Pages (from-to)9193-9205
Number of pages13
JournalNucleic Acids Research
Volume40
Issue number18
DOIs
StatePublished - Oct 2012

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DNA-Directed DNA Polymerase
Mechanics
Water
DNA
Phosphoranes
Phosphates
Environmental Carcinogens
Vehicle Emissions
Benzo(a)pyrene
Molecular Dynamics Simulation
Smoke
Tobacco
Protons
Hydrogen
Oxygen
Food
Mutation
Enzymes

ASJC Scopus subject areas

  • Genetics

Cite this

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title = "Preferred WMSA catalytic mechanism of the nucleotidyl transfer reaction in human DNA polymerase κ elucidates error-free bypass of a bulky DNA lesion",
abstract = "Human DNA Pol i is a polymerase enzyme, specialized for near error-free bypass of certain bulky chemical lesions to DNA that are derived from environmental carcinogens present in tobacco smoke, automobile exhaust and cooked food. By employing ab initio QM/MM-MD (Quantum Mechanics/ Molecular Mechanics-Molecular Dynamics) simulations with umbrella sampling, we have determined the entire free energy profile of the nucleotidyl transfer reaction catalyzed by Pol κ and provided detailed mechanistic insights. Our results show that a variant of the Water Mediated and Substrate Assisted (WMSA) mechanism that we previously deduced for Dpo4 and T7 DNA polymerases is preferred for Pol κ as well, suggesting its broad applicability. The hydrogen on the 3'-OHprimer terminus is transferred through crystal and solvent waters to the γ-phosphate of the dNTP, followed by the associative nucleotidyl transfer reaction; this is facilitated by a proton transfer from the γ-phosphate to the α,β-bridging oxygen as pyrophosphate leaves, to neutralize the evolving negative charge. MD simulations show that the near error-free incorporation of dCTP opposite the major benzo[a]pyrene-derived dG lesion is compatible with the WMSA mechanism, allowing for an essentially undisturbed pentacovalent phosphorane transition state, and explaining the bypass of this lesion with little mutation by Pol κ.",
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T1 - Preferred WMSA catalytic mechanism of the nucleotidyl transfer reaction in human DNA polymerase κ elucidates error-free bypass of a bulky DNA lesion

AU - Lior-Hoffmann, Lee

AU - Wang, Lihua

AU - Wang, Shenglong

AU - Geacintov, Nicholas E.

AU - Broyde, Suse

AU - Zhang, Yingkai

PY - 2012/10

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