PDBsum entry 2br0

Transferase

PDB id: 2br0

Contents

Protein chain

342 a.a. *

DNA/RNA

Ligands

_DG

Metals

_CA ×3

Waters ×118

* Residue conservation analysis

PDB id:

2br0

Name:

Transferase

Title:

DNA adduct bypass polymerization by sulfolobus solfataricus dpo4. Analysis and crystal structures of multiple base-pair substitution and frameshift products with the adduct 1,n2-ethenoguanine

Structure:

DNA polymerase iv. Chain: a. Synonym: pol iv. Engineered: yes. 5'-d( Gp Gp Gp Gp Gp Ap Ap Gp Gp Ap Tp Tp C)-3'. Chain: p. Engineered: yes. 5'-d( Tp Cp Ap Cp Gnep Gp Ap Ap Tp Cp Cp Tp Tp Cp Cp Cp Cp C)-3'.

Source:

Sulfolobus solfataricus. Organism_taxid: 273057. Strain: p2. Gene: dpo4. Expressed in: escherichia coli. Expression_system_taxid: 469008. Synthetic: yes. Synthetic: yes

Biol. unit:

Trimer (from PDB file)

Resolution:

2.17Å R-factor: 0.239 R-free: 0.266

Authors:

A.Irimia,L.V.Loukachevitch,M.Egli

Key ref:

H.Zang et al. (2005). DNA adduct bypass polymerization by Sulfolobus solfataricus DNA polymerase Dpo4: analysis and crystal structures of multiple base pair substitution and frameshift products with the adduct 1,N2-ethenoguanine. J Biol Chem, 280, 29750-29764. PubMed id: 15965231 DOI: 10.1074/jbc.M504756200

Date:

28-Apr-05 Release date: 23-Jun-05

Protein chain

Q97W02  (DPO4_SULSO) -  DNA polymerase IV from Saccharolobus solfataricus (strain ATCC 35092 / DSM 1617 / JCM 11322 / P2)

Seq: 352 a.a.

Struc: 343 a.a.

DNA/RNA chains

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

C-A-C-GNE-G-A-A-T-C-C-T-T-C-C-C-C-C 17 bases

Enzyme reactions

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

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

Added reference

DOI no: 10.1074/jbc.M504756200

J Biol Chem 280:29750-29764 (2005)

PubMed id: 15965231

DNA adduct bypass polymerization by Sulfolobus solfataricus DNA polymerase Dpo4: analysis and crystal structures of multiple base pair substitution and frameshift products with the adduct 1,N2-ethenoguanine.

H.Zang, A.K.Goodenough, J.Y.Choi, A.Irimia, L.V.Loukachevitch, I.D.Kozekov, K.C.Angel, C.J.Rizzo, M.Egli, F.P.Guengerich.

ABSTRACT

1,N(2)-Etheno(epsilon)guanine is a mutagenic DNA lesion derived from lipid oxidation products and also from some chemical carcinogens. Gel electrophoretic analysis of the products of primer extension by Sulfolobus solfataricus P2 DNA polymerase IV (Dpo4) indicated preferential incorporation of A opposite 3'-(1,N(2)-epsilon-G)TACT-5', among the four dNTPs tested individually. With the template 3'-(1,N(2)-epsilon-G)CACT-5', both G and A were incorporated. When primer extension was done in the presence of a mixture of all four dNTPs, high pressure liquid chromatography-mass spectrometry analysis of the products indicated that (opposite 3'-(1,N(2)-epsilon-G)CACT-5') the major product was 5'-GTGA-3' and the minor product was 5'-AGTGA-3'. With the template 3'-(1,N(2)-epsilon-G)TACT-5', the following four products were identified by high pressure liquid chromatography-mass spectrometry: 5'-AATGA-3', 5'-ATTGA-3', 5'-ATGA-3', and 5'-TGA-3'. An x-ray crystal structure of Dpo4 was solved (2.1 A) with a primer-template and A placed in the primer to be opposite the 1,N(2)-epsilon-G in the template 3'-(1,N(2)-epsilon-G)TACT 5'. The added A in the primer was paired across the template T with classic Watson-Crick geometry. Similar structures were observed in a ternary Dpo4-DNA-dATP complex and a ternary Dpo4-DNA-ddATP complex, with d(d)ATP opposite the template T. A similar structure was observed with a ddGTP adjacent to the primer and opposite the C next to 1,N(2)-epsilon-G in 3'-(1,N(2)-epsilon-G)CACT-5'. We concluded that Dpo4 uses several mechanisms, including A incorporation opposite 1,N(2)-epsilon-G and also a variation of dNTP-stabilized misalignment, to generate both base pair and frameshift mutations.

Selected figure(s)

Figure 2.
FIG. 2. Extension of a 32P-labeled primer opposite template adduct by pol T7^- and Dpo4 in the presence of single dNTPs. The concentrations of each polymerase used are indicated. A, the percentage of extension with 20 nM pol T7^- was 59, 4, 5, and 9% for A, G, C, and T, respectively. B, the percentage of extension with 100 nM Dpo4 was 23, 26, 8, and 11% for A, G., C, and T, respectively.

Figure 3.
FIG. 3. Extension of a 32P-labeled primer opposite the template adduct by pol T7^- or Dpo4 in the presence of single dNTPs. The concentrations of each polymerase used are indicated. A, pol T7^-. B, Dpo4.

The above figures are reprinted by permission from the ASBMB: J Biol Chem (2005, 280, 29750-29764) copyright 2005.

Figures were selected by an automated process.