A quantum mechanical investigation of possible mechanisms for the nucleotidyl transfer reaction catalyzed by DNA polymerase β

Mihaela D. Bojin, Tamar Schlick

Research output: Contribution to journalArticlepeer-review


Several quantum mechanical (QM) and hybrid quantum/molecular mechanical (QM/MM) studies have been employed recently to analyze the nucleotidyl transfer reaction in DNA polymerase β(pol β). Our examination reveals strong dependence of the reported mechanism on the initial molecular model. Thus, we explore here several model systems by QM methods to investigate pol β's possible pathway variations. Although our most favorable pathway involves a direct proton transfer from O3′(primer) to O2α(P α)> we also discuss other initial proton-transfer steps - to an adjacent water, to triphosphate, or to aspartic units - and the stabilizing effect of crystallographic water molecules in the active site. Our favored reaction route has an energetically undemanding initial step of less than 1.0 kcal/mol (at the B3LYP/6-31G(d,p) level), and involves a slight rearrangement in the geometry of the active site. This is followed by two major steps: (1) direct proton transfer from 03′(primer) to O2α(P α) leading to the formation of a pentavalent, trigonal bipyramidal Pα center, via an associative mechanism, at a cost of about 28 kcal/mol, and (2) breakage of the triphosphate unit (exothermic process, ∼22 kcal/mol) that results in the full transfer of the nucleotide to the DNA and the formation of pyrophosphate. These energy values are expected to be lower in the physical system when full protein effects are incorporated. We also discuss variations from this dominant pathway, and their impact on the overall repair process. Our calculated barrier for the chemical reaction clearly indicates that chemistry is rate-limiting overall for correct nucleotide insertion in pol β, in accord with other studies. Protonation studies on relevant intermediates suggest that, although protonation at a single aspartic residue may occur, the addition of a second proton to the system significantly disturbs the active site. We conclude that the active site rearrangement step necessary to attain a reaction-competent geometry is essential and closely related to the "pre-chemistry" avenue described recently as a key step in the overall kinetic cycle of DNA polymerases. Thus, our work emphasizes the many possible ways for DNA polymerase β's chemical reaction to occur, determined by the active site environment and initial models.

Original languageEnglish (US)
Pages (from-to)11244-11252
Number of pages9
JournalJournal of Physical Chemistry B
Issue number38
StatePublished - Sep 27 2007

ASJC Scopus subject areas

  • Physical and Theoretical Chemistry
  • Surfaces, Coatings and Films
  • Materials Chemistry


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