All cells rely on high fidelity polymerases for faithful DNA replication.  DNA polymerase III (PolIII), the main enzyme responsible for replication in E. coli, synthesizes over 100,000 basepairs (bp) per binding event, with an error rate of ~1 per million.  On the rare occasion that a misincorporation does occur, PolIII utilizes a 3’-5’ exonuclease to remove the misincorporated nucleotide so that synthesis may continue.  Combined with its exonuclease activity, PolIII’s error rate reaches ~1 per billion.  New cryo-EM data has provided atomic coordinates of PolIII in conformations involving both the synthesis of DNA (polymerase mode) and the correction of a misincorporated nucleotide (editing mode).  Using this information, we have implemented targeted molecular dynamics (tMD) to generate a transition pathway between the polymerase and editing modes.  Evenly spaced snapshots, taken from the tMD trajectory, were further simulated using molecular dynamics (MD).  In order to gain insight into the global motions that govern the transition from polymerase mode to editing mode, we preformed principal component analysis on the resultant MD trajectories.  Our initial results show that PolIII must undergo a number of large-scale motions before it can begin to repair the mismatched segment of DNA.  Furthermore, our results indicate that several short-lived intermediate states exist along this pathway, most of which have remained elusive to current experimental techniques.

Molecular dynamics simulations reveal structural details on the proofreading mechanism of DNA polymerase III

256th National Meeting (2018)

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