PDBsum entry 1skr

Transferase/electron transport/DNA

PDB id: 1skr

Contents

Protein chains

669 a.a. *

105 a.a. *

DNA/RNA

Ligands

DAD

Metals

_MG ×3

Waters ×381

* Residue conservation analysis

PDB id:

1skr

Name:

Transferase/electron transport/DNA

Title:

T7 DNA polymerase complexed to DNA primer/template and ddatp

Structure:

5'-d( Cp Gp Ap Ap Ap Ap Cp Gp Ap C Gp Gp Cp Cp Ap Gp Tp Gp Cp Cp Ap (2Da))-3'. Chain: p. Engineered: yes. Other_details: DNA primer. 5'-d( Cp Cp Cp Tp Tp Tp Gp Gp Cp Ap Cp Tp Gp Gp Cp Cp Gp Tp Cp Gp Tp Tp Tp Tp Cp G)-3'. Chain: t. Engineered: yes.

Source:

Synthetic: yes. Enterobacteria phage t7. Organism_taxid: 10760. Gene: 5. Expressed in: escherichia coli. Expression_system_taxid: 562. Escherichia coli. Organism_taxid: 562. Gene: trxa, tsnc, fipa, b3781, c4701, z5291, ecs4714, stm3915,

Biol. unit:

Tetramer (from PQS)

Resolution:

2.40Å R-factor: 0.226 R-free: 0.270

Authors:

Y.Li,S.Dutta,S.Doublie,H.M.Bdour,J.S.Taylor,T.Ellenberger

Key ref:

Y.Li et al. (2004). Nucleotide insertion opposite a cis-syn thymine dimer by a replicative DNA polymerase from bacteriophage T7. Nat Struct Mol Biol, 11, 784-790. PubMed id: 15235589 DOI: 10.1038/nsmb792

Date:

05-Mar-04 Release date: 06-Jul-04

Protein chain

P00581  (DPOL_BPT7) -  DNA-directed DNA polymerase from Escherichia phage T7

Seq: 704 a.a.

Struc: 669 a.a.

Protein chain

P0AA25  (THIO_ECOLI) -  Thioredoxin 1 from Escherichia coli (strain K12)

Seq: 109 a.a.

Struc: 105 a.a.

DNA/RNA chains

G-C-C-A-G-T-G-C-C-A-2DA 11 bases

C-T-T-T-G-G-C-A-C-T-G-G-C 13 bases

Enzyme reactions

• Enzyme class 2: Chain A: 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: Chain A: E.C.3.1.11.- - ?????
• Enzyme class 4: Chain B: E.C.?

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

DOI no: 10.1038/nsmb792

Nat Struct Mol Biol 11:784-790 (2004)

PubMed id: 15235589

Nucleotide insertion opposite a cis-syn thymine dimer by a replicative DNA polymerase from bacteriophage T7.

Y.Li, S.Dutta, S.Doublié, H.M.Bdour, J.S.Taylor, T.Ellenberger.

ABSTRACT

Ultraviolet-induced DNA damage poses a lethal block to replication. To understand the structural basis for this, we determined crystal structures of a replicative DNA polymerase from bacteriophage T7 in complex with nucleotide substrates and a DNA template containing a cis-syn cyclobutane pyrimidine dimer (CPD). When the 3' thymine is the templating base, the CPD is rotated out of the polymerase active site and the fingers subdomain adopts an open orientation. When the 5' thymine is the templating base, the CPD lies within the polymerase active site where it base-pairs with the incoming nucleotide and the 3' base of the primer, while the fingers are in a closed conformation. These structures reveal the basis for the strong block of DNA replication that is caused by this photolesion.

Selected figure(s)

Figure 1.
Figure 1. Positioning of the cis-syn thymine dimer in the T7 DNA polymerase. (a) CPD-3'a complex, in which the CPD is rotated by 90° and lying outside of the polymerase active site. The fingers domain is in an open conformation. (b) CPD-5'b complex, in which the CPD is in the polymerase active site and it base-pairs with the 3' base of the primer strand and the incoming nucleotide. The fingers domain is in a closed conformation. (c) CPD-5'a complex, in which the DNA slides back by one base pair and the CPD curves back into the major grove of the duplex DNA. The fingers domain is in an open conformation. The protein is shown as surface representation. The DNA and nucleotide are ball-and-stick models in which the backbone of the primer strand is red and that of the template is light purple. The 5' base of the cis-syn thymine dimer is magenta and the 3' base is light magenta. The carbon atoms in the incoming nucleotide in b are green and the Mg2+ ions are gold spheres. For clarity, the thumb subdomain and the processivity factor E. coli thioredoxin were removed from Figures 1,2,3.

Figure 4.
Figure 4. The cis-syn thymine dimer base pairs with ddATP in the closed CPD-5'b complex. (a) View from the major groove side of the duplex DNA. (b) The view in a has been rotated by 90° along an axis (straight arrow). The 3' T of the CPD forms a normal Watson-Crick base pair with the 3' base of the primer. A twisted base pair is formed between the 5' T and the incoming ddATP. The view in b shows the distorted stacking of the 5' T base pair with that of the 3' T. The nucleic acids are in ball-and-stick representation with the same color scheme used in Figure 1.

The above figures are reprinted by permission from Macmillan Publishers Ltd: Nat Struct Mol Biol (2004, 11, 784-790) copyright 2004.

Figures were selected by an automated process.