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In crystallo observation of three metal ion promoted DNA polymerase misincorporation

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  • Caleb Chang

    (Rice University)

  • Christie Lee Luo

    (Rice University)

  • Yang Gao

    (Rice University)

Abstract

Error-free replication of DNA is essential for life. Despite the proofreading capability of several polymerases, intrinsic polymerase fidelity is in general much higher than what base-pairing energies can provide. Although researchers have investigated this long-standing question with kinetics, structural determination, and computational simulations, the structural factors that dictate polymerase fidelity are not fully resolved. Time-resolved crystallography has elucidated correct nucleotide incorporation and established a three-metal-ion-dependent catalytic mechanism for polymerases. Using X-ray time-resolved crystallography, we visualize the complete DNA misincorporation process catalyzed by DNA polymerase η. The resulting molecular snapshots suggest primer 3´-OH alignment mediated by A-site metal ion binding is the key step in substrate discrimination. Moreover, we observe that C-site metal ion binding preceded the nucleotidyl transfer reaction and demonstrate that the C-site metal ion is strictly required for misincorporation. Our results highlight the essential but separate roles of the three metal ions in DNA synthesis.

Suggested Citation

  • Caleb Chang & Christie Lee Luo & Yang Gao, 2022. "In crystallo observation of three metal ion promoted DNA polymerase misincorporation," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
    • Handle: RePEc:nat:natcom:v:13:y:2022:i:1:d:10.1038_s41467-022-30005-3
    DOI: 10.1038/s41467-022-30005-3
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    References listed on IDEAS

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    2. Nicholas Chim & Roman A. Meza & Anh M. Trinh & Kefan Yang & John C. Chaput, 2021. "Following replicative DNA synthesis by time-resolved X-ray crystallography," Nature Communications, Nature, vol. 12(1), pages 1-8, December.
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