PDBsum entry 2pfo

Transferase, lyase/DNA

PDB id: 2pfo

Contents

Protein chain

323 a.a. *

DNA/RNA

Ligands

DUP

EDO

Metals

_MG

_MN

_NA ×4

Waters ×329

* Residue conservation analysis

PDB id:

2pfo

Name:

Transferase, lyase/DNA

Title:

DNA polymerase lambda in complex with DNA and dupnpp

Structure:

Template. Chain: t. Engineered: yes. Primer. Chain: p. Engineered: yes. Downstream primer. Chain: d. Engineered: yes.

Source:

Synthetic: yes. Homo sapiens. Human. Organism_taxid: 9606. Gene: poll. Expressed in: escherichia coli. Expression_system_taxid: 562

Resolution:

2.00Å R-factor: 0.217 R-free: 0.242

Authors:

M.Garcia-Diaz,K.Bebenek,J.M.Krahn,L.C.Pedersen,T.A.Kunkel

Key ref:

M.Garcia-Diaz et al. (2007). Role of the catalytic metal during polymerization by DNA polymerase lambda. Dna Repair (amst), 6, 1333-1340. PubMed id: 17475573

Date:

05-Apr-07 Release date: 15-May-07

Protein chain

Q9UGP5  (DPOLL_HUMAN) -  DNA polymerase lambda from Homo sapiens

Seq: 575 a.a.

Struc: 323 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

DNA/RNA chains

C-G-G-C-A-G-T-A-C-T-G 11 bases

C-A-G-T-A-C 6 bases

G-C-C-G 4 bases

Enzyme reactions

• Enzyme class 2: E.C.2.7.7.7 - DNA-directed Dna polymerase.
• Reaction: DNA(n) + a 2'-deoxyribonucleoside 5'-triphosphate = DNA(n+1) + diphosphate
• Enzyme class 3: E.C.4.2.99.- - ?????

Note, where more than one E.C. class is given (as above), each may correspond to a different protein domain or, in the case of polyprotein precursors, to a different mature protein.

Molecule diagrams generated from .mol files obtained from the KEGG ftp site

Added reference

Dna Repair (amst) 6:1333-1340 (2007)

PubMed id: 17475573

Role of the catalytic metal during polymerization by DNA polymerase lambda.

M.Garcia-Diaz, K.Bebenek, J.M.Krahn, L.C.Pedersen, T.A.Kunkel.

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.