Bidentate and tridentate metal-ion coordination states within ternary complexes of RB69 DNA polymerase
- Yale
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 {alpha}-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 {alpha},{beta},{gamma}-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 {beta},{gamma}-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 {angstrom} resolution. Our observations suggest that several distinct metal-ion 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.
- Research Organization:
- Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
- Sponsoring Organization:
- FOREIGNNIH
- OSTI ID:
- 1034949
- Journal Information:
- Protein Science, Vol. 21, Issue 3
- Country of Publication:
- United States
- Language:
- ENGLISH
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