3uiq

Publication Abstract from PubMed

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 A 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.

Bidentate and tridentate metal-ion coordination states within ternary complexes of RB69 DNA polymerase.,Xia S, Eom SH, Konigsberg WH, Wang J Protein Sci. 2012 Mar;21(3):447-51. doi: 10.1002/pro.2026. Epub 2012 Jan 31. PMID:22238207^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.