Polymerase Simulations Reveal That Arg258 Rotation is a Slow Step Rather Than Large Subdomain Motions Per Se

The large-scale opening motion of mammalian polymerase is followed at atomic resolution by dynamic simulations that link crystal `closed' and `open' conformations. This conformational change is thought to be key to the ability of polymerases to choose a correct nucleotide and hence maintain DNA synthesis fidelity. Corroborating available structural and kinetic measurements, our studies delineate a sequence of steps after the chemical DNA extension step: Phe272 ring flip, large thumb movement, and Arg258 rotation with release of catalytic Mg2+. Significantly, we identify the last step as slow rather than large subdomain motion per se. In the closing, these slow rearrangements could be further hampered if pol binds to an incorrect nucleotide. Our results suggest experiments that could provide further insights into mechanisms of error discrimination and, more broadly, aid in our understanding of recently discovered polymerases that exhibit extraordinarily low fidelity .

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