DNA Polymerase

The importance of DNA polymerase fidelity both in continued survival and in scientific advancement is clear, though many of the details of polymerase behavior in the presence of a mismatch in the active site remain unknown. Using computational methods, the large fragment of Bacillus stearothermophilus DNA polymerase I is simulated in the presence of all sixteen possible template-trinucleotide pairs, including four Watson-Crick pairs and twelve mismatches. The pre-catalytic steps of nucleotide addition from nucleotide insertion to the moments immediately preceding chemical addition of a new nucleotide to the primer strand are explored using three starting structures representing different stages of nucleotide addition. All nucleotide pairs are simulated from these starting structures and details of interactions between the members of the nascent base pair and between the trinucleotide and the DNA polymerase itself are elucidated. Classification of nucleotide pairs based on strength and type of interactions, patterns of large-scale conformational shifts, description of stabilizing reactions between amino acids and bound trinucleotides, and binding affinity analysis are all completed on 50+ µs of simulation.