2PFP

DNA Polymerase lambda in complex with DNA and dCTP

Summary for 2PFP

• Entry DOI: 10.2210/pdb2pfp/pdb
• Related: 2PFN 2PFO 2PFQ
• Descriptor: Template, Primer, Downstream Primer, ... (8 entities in total)
• Functional Keywords: dna polymerase, dna repair, phosphoryl transfer reaction, manganese, transferase, lyase-dna complex, lyase/dna
• Biological source: Homo sapiens (human)
• Cellular location: Nucleus : Q9UGP5
• Total number of polymer chains: 4
• Total formula weight: 44429.35

Authors

Garcia-Diaz, M.,Bebenek, K.,Krahn, J.M.,Pedersen, L.C.,Kunkel, T.A. (deposition date: 2007-04-05, release date: 2007-05-15, Last modification date: 2023-08-30)

Primary citation

Garcia-Diaz, M.,Bebenek, K.,Krahn, J.M.,Pedersen, L.C.,Kunkel, T.A.
Role of the catalytic metal during polymerization by DNA polymerase lambda.
DNA Repair, 6:1333-1340, 2007

PubMed Abstract: The incorporation of dNMPs into DNA by polymerases involves a phosphoryl transfer reaction hypothesized to require two divalent metal ions. Here we investigate this hypothesis using as a model human DNA polymerase lambda (Pol lambda), an enzyme suggested to be activated in vivo by manganese. We report the crystal structures of four complexes of human Pol lambda. In a 1.9 A structure of Pol lambda containing a 3'-OH and the non-hydrolyzable analog dUpnpp, a non-catalytic Na+ ion occupies the site for metal A and the ribose of the primer-terminal nucleotide is found in a conformation that positions the acceptor 3'-OH out of line with the alpha-phosphate and the bridging oxygen of the pyrophosphate leaving group. Soaking this crystal in MnCl2 yielded a 2.0 A structure with Mn2+ occupying the site for metal A. In the presence of Mn2+, the conformation of the ribose is C3'-endo and the 3'-oxygen is in line with the leaving oxygen, at a distance from the phosphorus atom of the alpha-phosphate (3.69 A) consistent with and supporting a catalytic mechanism involving two divalent metal ions. Finally, soaking with MnCl2 converted a pre-catalytic Pol lambda/Na+ complex with unreacted dCTP in the active site into a product complex via catalysis in the crystal. These data provide pre- and post-transition state information and outline in a single crystal the pathway for the phosphoryl transfer reaction carried out by DNA polymerases.

PubMed: 17475573
DOI: 10.1016/j.dnarep.2007.03.005

Experimental method

X-RAY DIFFRACTION (2.1 Å)