title: Pre-Steady-State Kinetic Analysis of Truncated and Full-Length Saccharomyces cerevisiae DNA Polymerase Eta.
abstract: Understanding polymerase fidelity is an important objective towards ascertaining the overall stability of an organism's genome. Saccharomyces cerevisiae DNA polymerase eta (yPoleta), a Y-family DNA polymerase, is known to efficiently bypass DNA lesions (e.g., pyrimidine dimers) in vivo. Using pre-steady-state kinetic methods, we examined both full-length and a truncated version of yPoleta which contains only the polymerase domain. In the absence of yPoleta's C-terminal residues 514-632, the DNA binding affinity was weakened by 2-fold and the base substitution fidelity dropped by 3-fold. Thus, the C-terminus of yPoleta may interact with DNA and slightly alter the conformation of the polymerase domain during catalysis. In general, yPoleta discriminated between a correct and incorrect nucleotide more during the incorporation step (50-fold on average) than the ground-state binding step (18-fold on average). Blunt-end additions of dATP or pyrene nucleotide 5'-triphosphate revealed the importance of base stacking during the binding of incorrect incoming nucleotides.
authors.affiliation: Department of Biochemistry, The Ohio State University, Columbus, OH 43210, USA.
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authors.lastname: Burgers
authors.forename: Zucai
authors.initials: Z
authors.lastname: Suo
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