Threonine 79 Is a Hinge Residue That Governs the Fidelity of DNA Polymerase β by Helping to Position the DNA within the Active Site*

DNA polymerase β (pol β) is an ideal system for studying the role of its different amino acid residues in the fidelity of DNA synthesis. In this study, the T79S variant of pol β was identified using an in vivo genetic screen. T79S is located in the N-terminal 8-kDa domain of pol β and has no contact with either the DNA template or the incoming dNTP substrate. The T79S protein produced 8-fold more multiple mutations in the herpes simplex virus type 1-thymidine kinase assay than wild-type pol β. Surprisingly, T79S is a misincorporation mutator only when using a 3′-recessed primer-template. In the presence of a single nucleotide-gapped DNA substrate, T79S displays an antimutator phenotype when catalyzing DNA synthesis opposite template C and has similar fidelity as wild type opposite templates A, G, or T. Threonine 79 is located directly between two helix-hairpin-helix motifs located within the 8-kDa and thumb domains of pol β. As the pol β enzyme closes into its active form, the helix-hairpin-helix motifs appear to assist in the production and stabilization of a 90o bend of the DNA. The function of the bent DNA is to present the templating base to the incoming nucleotide substrate. We propose that Thr-79 is part of a hydrogen bonding network within the helix-hairpin-helix motifs that is important for positioning the DNA within the active site. We suggest that alteration of Thr-79 to Ser disrupts this hydrogen bonding network and results in an enzyme that is unable to bend the DNA into the proper geometry for accurate DNA synthesis.

• Publication year

  2002

• Venue

  Journal of Biological Chemistry

• Publication date

  2002-09-20

• Fields of study

  Biology, Medicine, Chemistry

• Identifiers
  DOI 10.1074/jbc.M204953200PMID 12121998
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• No claims are published for this paper.

• No concepts are published for this paper.

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