of Work: We are examining the relationship between the structures of DNA polymerases and their functions, including fidelity. Accomplishments this year include the following. We determined that the fidelity of a archeal Y family DNA polymerase (Sso Dpo4) is very low. We also provided evidence that nucleotide misalignment in the active sites of this and likely other TLS polymerases generates frameshift mutations. We discovered that Pol lambda (family X) has 5'-dRPase activity and other properties suggesting its participation in base excision repair of DNA damage resulting from environmental stress. We collaborated with the NMR group to determine the NMR structure of the 8 kDa dRP lyase domain of DNA polymerase lambda. Based on structural information and biochemical properties, we proposed and tested a model for altered interactions of the polymerases active site with the DNA minor groove that explains the unusual base substitution error specificity of the (Family A) Klenow fragment of E. coli DNA polymerase I containing an amino acid replacement in the binding pocket for the nascent base pair. We presented evidence that DNA polymerase eta also likely participates in somatic hypermutation of immunoglobulin genes, a process responsible to development of high affinity antibodies. We presented evidence that a specific tyrosine residue in the site of DNA polymerase eta participates in determining the limited base selectivity of the highly inaccurate enzyme, as well as in the efficiency with which it bypasses a helix-distorting UV photoproduct. This study has implications for the ability of pol eta to strongly suppress human susceptibility to skin cancer. In collaboration with the Wilson group, we further characterized the dRP lyase activity of pol iota and localized this activity by controlled proteolysis of the recombinant protein expressed in insect cells. In collaboration with the Copeland group we discovered that, in the gene for the mitochondrial replicative DNA polymerase gamma, an active site point mutation that is associated with the human genetic disease Progressive External Opthalmoplegia encodes a polymerase with reduced catalytic efficiency and reduced DNA replication fidelity.
| Kaminski, Andrea M; Tumbale, Percy P; Schellenberg, Matthew J et al. (2018) Structures of DNA-bound human ligase IV catalytic core reveal insights into substrate binding and catalysis. Nat Commun 9:2642 |
| Orebaugh, Clinton D; Lujan, Scott A; Burkholder, Adam B et al. (2018) Mapping Ribonucleotides Incorporated into DNA by Hydrolytic End-Sequencing. Methods Mol Biol 1672:329-345 |
| Burkholder, Adam B; Lujan, Scott A; Lavender, Christopher A et al. (2018) Muver, a computational framework for accurately calling accumulated mutations. BMC Genomics 19:345 |
| Zhou, Zhi-Xiong; Williams, Jessica S; Kunkel, Thomas A (2018) Studying Ribonucleotide Incorporation: Strand-specific Detection of Ribonucleotides in the Yeast Genome and Measuring Ribonucleotide-induced Mutagenesis. J Vis Exp : |
| Williams, Jessica S; Kunkel, Thomas A (2018) Studying Topoisomerase 1-Mediated Damage at Genomic Ribonucleotides. Methods Mol Biol 1703:241-257 |
| Huang, Shar-Yin N; Williams, Jessica S; Arana, Mercedes E et al. (2017) Topoisomerase I-mediated cleavage at unrepaired ribonucleotides generates DNA double-strand breaks. EMBO J 36:361-373 |
| Jamsen, Joonas A; Beard, William A; Pedersen, Lars C et al. (2017) Time-lapse crystallography snapshots of a double-strand break repair polymerase in action. Nat Commun 8:253 |
| Burgers, Peter M J; Kunkel, Thomas A (2017) Eukaryotic DNA Replication Fork. Annu Rev Biochem 86:417-438 |
| Lujan, Scott A; Williams, Jessica S; Kunkel, Thomas A (2016) DNA Polymerases Divide the Labor of Genome Replication. Trends Cell Biol 26:640-654 |
| Watt, Danielle L; Buckland, Robert J; Lujan, Scott A et al. (2016) Genome-wide analysis of the specificity and mechanisms of replication infidelity driven by imbalanced dNTP pools. Nucleic Acids Res 44:1669-80 |
Showing the most recent 10 out of 120 publications
