3SPY

RB69 DNA Polymerase(L415A/L561A/S565G/Y567A) Ternary Complex with dUpCpp Opposite dA

Summary for 3SPY

• Entry DOI: 10.2210/pdb3spy/pdb
• Related: 3SPZ 3SQ0 3SQ1 3SQ2 3SQ4
• Descriptor: DNA polymerase, 5'-D(*TP*CP*GP*AP*GP*TP*AP*AP*GP*CP*AP*GP*TP*CP*CP*GP*CP*G)-3', 5'-D(*GP*CP*GP*GP*AP*CP*TP*GP*CP*TP*TP*AP*C)-3', ... (6 entities in total)
• Functional Keywords: dupcpp, quadruple mutant, transferase-dna complex, transferase/dna
• Biological source: Enterobacteria phage RB69
• Total number of polymer chains: 3
• Total formula weight: 114283.48

Authors

Xia, S.,Konigsberg, W.H.,Wang, J. (deposition date: 2011-07-04, release date: 2011-10-12, Last modification date: 2024-02-28)

Primary citation

Xia, S.,Wang, M.,Blaha, G.,Konigsberg, W.H.,Wang, J.
Structural Insights into Complete Metal Ion Coordination from Ternary Complexes of B Family RB69 DNA Polymerase.
Biochemistry, 50:9114-9124, 2011

PubMed Abstract: We have captured a preinsertion ternary complex of RB69 DNA polymerase (RB69pol) containing the 3' hydroxyl group at the terminus of an extendable primer (ptO3') and a nonhydrolyzable 2'-deoxyuridine 5'-α,β-substituted triphosphate, dUpXpp, where X is either NH or CH(2), opposite a complementary templating dA nucleotide residue. Here we report four structures of these complexes formed by three different RB69pol variants with catalytically inert Ca(2+) and four other structures with catalytically competent Mn(2+) or Mg(2+). These structures provide new insights into why the complete divalent metal-ion coordination complexes at the A and B sites are required for nucleotidyl transfer. They show that the metal ion in the A site brings ptO3' close to the α-phosphorus atom (Pα) of the incoming dNTP to enable phosphodiester bond formation through simultaneous coordination of both ptO3' and the nonbridging Sp oxygen of the dNTP's α-phosphate. The coordination bond length of metal ion A as well as its ionic radius determines how close ptO3' can approach Pα. These variables are expected to affect the rate of bond formation. The metal ion in the B site brings the pyrophosphate product close enough to Pα to enable pyrophosphorolysis and assist in the departure of the pyrophosphate. In these dUpXpp-containing complexes, ptO3' occupies the vertex of a distorted metal ion A coordination octahedron. When ptO3' is placed at the vertex of an undistorted, idealized metal ion A octahedron, it is within bond formation distance to Pα. This geometric relationship appears to be conserved among DNA polymerases of known structure.

PubMed: 21923197
DOI: 10.1021/bi201260h

Experimental method

X-RAY DIFFRACTION (2.14 Å)