A triad interaction in the fingers subdomain of DNA polymerase beta controls polymerase activity

Drew L. Murphy, Joachim Jaeger, Joann B. Sweasy

Research output: Contribution to journalArticlepeer-review

18 Scopus citations

Abstract

DNA polymerase beta (pol beta) is the main polymerase involved in the base excision repair pathway responsible for repairing damaged bases in the DNA. Previous studies on the H285D mutant of pol beta suggested that the C-terminal region of the polymerase is important for polymerase function. In this study, the C-terminal region of pol beta was mutated to assess its role in polymerization. Kinetic experiments showed that the C-terminal region is required for wild-type polymerase activity. Additionally, an interaction between the fingers and palm subdomain revealed itself to be required for polymerase activity. The E316R mutant of pol beta was shown to have a 29 000-fold reduction in polymerization rate with no reduction in nucleotide binding, suggesting that there exists a noncovalent mechanistic step between nucleotide binding and nucleophilic attack of the primer 3′-hydroxyl group on the α-PO 4 of the nucleotide. Molecular modeling studies of the E316R mutant demonstrate that disrupting the interaction between Arg182 and Glu316 disrupts the packing of side chains in the hydrophobic hinge region and may be hampering the conformational change during polymerization. Taken together, these data demonstrate that the triad interaction of Arg182, Glu316, and Arg333 is crucial for polymerase function.

Original languageEnglish (US)
Pages (from-to)6279-6287
Number of pages9
JournalJournal of the American Chemical Society
Volume133
Issue number16
DOIs
StatePublished - Apr 27 2011
Externally publishedYes

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

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