We will use computational techniques that were previously developed and tested by our research group in studies of DNA replication and related biochemical processes. These computer simulations will systematically examine changes in the activation barrier of the chemical step due to different factors including the effects of point mutations of distant pol B residues, to substitutions of the p-y bridging oxygen by the - C X Y - groups where X. Y = H, CH3, F, CI, Br or N3, and substitutions in the deoxyribose residues of the primer or dNTP nucleotides. Additionally, calculations of the substrate binding in its ground- and transition-state configurations will be investigated by proven computational strategies with extended simulation times. The results of the simulations (including TS structures and charge distributions as well as simulation protocols) will form a database that will include numerous experimentally-testable predictions along with the list of the assumptions and theoretical models that were used to generate these predictions. This database, which will be made accessible online, will be continually updated with new experimental and computational results, helping in selecting the best models and simulation strategies, and eventually to understanding of the functionally important properties of DNA polymerases at the atomic level. Additionally, the comparative studies will help in selecting the most effective approach for computational inhibitor and mutational screening.
The integration of structural, biochemical and computational studies of DNA replication fidelity requires a collaborative approach, where the theoretical analysis should be treated as an integral part of the experimental effort. Thus, the proposed core will focus on supporting Projects 1-3 with routine calculations of the properties of DNA polymerase B(pol B).
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|Alnajjar, Khadijeh S; Garcia-Barboza, Beatriz; Negahbani, Amirsoheil et al. (2017) A Change in the Rate-Determining Step of Polymerization by the K289M DNA Polymerase ? Cancer-Associated Variant. Biochemistry 56:2096-2105|
|Alnajjar, Khadijeh S; Negahbani, Amirsoheil; Nakhjiri, Maryam et al. (2017) DNA Polymerase ? Cancer-Associated Variant I260M Exhibits Nonspecific Selectivity toward the ?-? Bridging Group of the Incoming dNTP. Biochemistry 56:5449-5456|
|Ni, Feng; Kung, Alvin; Duan, Yankun et al. (2017) Remarkably Stereospecific Utilization of ATP ?,?-Halomethylene Analogues by Protein Kinases. J Am Chem Soc 139:7701-7704|
|Petruska, John; Goodman, Myron F (2017) Relating DNA base-pairing in aqueous media to DNA polymerase fidelity. Nat Rev Chem 1:|
|Yoon, Hanwool; Warshel, Arieh (2017) Simulating the fidelity and the three Mg mechanism of pol ? and clarifying the validity of transition state theory in enzyme catalysis. Proteins 85:1446-1453|
|Maximoff, Sergey N; Kamerlin, Shina Caroline Lynn; Florián, Jan (2017) DNA Polymerase ? Active Site Favors a Mutagenic Mispair between the Enol Form of Deoxyguanosine Triphosphate Substrate and the Keto Form of Thymidine Template: A Free Energy Perturbation Study. J Phys Chem B 121:7813-7822|
|Hwang, Candy S; Xu, Liang; Wang, Wei et al. (2016) Functional interplay between NTP leaving group and base pair recognition during RNA polymerase II nucleotide incorporation revealed by methylene substitution. Nucleic Acids Res 44:3820-8|
|Klva?a, Martin; Bren, Urban; Florián, Jan (2016) Uniform Free-Energy Profiles of the P-O Bond Formation and Cleavage Reactions Catalyzed by DNA Polymerases ? and ?. J Phys Chem B 120:13017-13030|
|Matute, Ricardo A; Yoon, Hanwool; Warshel, Arieh (2016) Exploring the mechanism of DNA polymerases by analyzing the effect of mutations of active site acidic groups in Polymerase ?. Proteins 84:1644-1657|
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