Bidentate and tridentate metal-ion coordination states within ternary complexes of RB69 DNA polymerase

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Abstract

Two divalent metal ions are required for primer-extension catalyzed by DNA polymerases. One metal ion brings the 3′-hydroxyl of the primer terminus and the α-phosphorus atom of incoming dNTP together for bond formation so that the catalytically relevant conformation of the triphosphate tail of the dNTP is in an α,β,γ-tridentate coordination complex with the second metal ion required for proper substrate alignment. A probable base selectivity mechanism derived from structural studies on Dpo4 suggests that the inability of mispaired dNTPs to form a substrate-aligned, tridentate coordination complex could effectively cause the mispaired dNTPs to be rejected before catalysis. Nevertheless, we found that mispaired dNTPs can actually form a properly aligned tridentate coordination complex. However, complementary dNTPs occasionally form misaligned complexes with mutant RB69 DNA polymerases (RB69pols) that are not in a tridentate coordination state. Here, we report finding a β,γ-bidentate coordination complex that contained the complementary dUpNpp opposite dA in the structure of a ternary complex formed by the wild type RB69pol at 1.88 Å resolution. Our observations suggest that several distinct metalion coordination states can exist at the ground state in the polymerase active site and that base selectivity is unlikely to be based on metal-ion coordination alone. © 2012 The Protein Society.

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Xia, S., Eom, S. H., Konigsberg, W. H., & Wang, J. (2012). Bidentate and tridentate metal-ion coordination states within ternary complexes of RB69 DNA polymerase. Protein Science, 21(3), 447–451. https://doi.org/10.1002/pro.2026

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