Dynamic basis for dG•dT misincorporation via tautomerization and ionization
Nature
doi:10.1038/nature25487
Received:10 June 2016 Accepted:21 December 2017 Published online:31 January 2018
Abstract
Tautomeric and anionic Watson–Crick-like mismatches have important roles in replication and translation errors through mechanisms that are not fully understood. Here, using NMR relaxation dispersion, we resolve a sequence-dependent kinetic network connecting G•T/U wobbles with three distinct Watson–Crick mismatches: two rapidly exchanging
tautomeric species (Genol•T/U
G•Tenol/Uenol; population less than 0.4%) and one anionic species (G•T–/U–; population around 0.001% at neutral pH). The sequence-dependent
tautomerization or ionization step was inserted into a minimal kinetic mechanism for correct incorporation during replication after the initial binding of the nucleotide, leading to accurate predictions of the probability of dG•dT
misincorporation across different
polymerases and pH conditions and for a chemically modified nucleotide, and providing mechanisms for sequence-dependent
misincorporation. Our results indicate that the energetic penalty for
tautomerization and/or ionization accounts for an approximately 10−2 to 10−3-fold discrimination against
misincorporation, which proceeds primarily via
tautomeric dGenol•dT and dG•dTenol, with contributions from anionic dG•dT– dominant at pH 8.4 and above or for some
mutagenic nucleotides.
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