PDBsum entry 2hhs

Transferase/DNA

PDB id: 2hhs

Contents

Protein chain

580 a.a. *

DNA/RNA

Ligands

GLC-FRU ×2

SO4 ×3

Metals

_MG

Waters ×318

* Residue conservation analysis

PDB id:

2hhs

Name:

Transferase/DNA

Title:

O6-methyl:c pair in the polymerase-10 basepair position

Structure:

5'-d( Cp Ap Tp (6Og)p Cp Gp Ap Gp Tp Cp Ap Gp G)-3'. Chain: b. Engineered: yes. Other_details: DNA primer strand. 5'-d( Cp Cp Tp Gp Ap Cp Tp Cp Gp Cp Ap Tp Gp A)-3'. Chain: c. Engineered: yes. Other_details: DNA template strand. DNA polymerase i.

Source:

Synthetic: yes. Other_details: chemically synthesized. Geobacillus stearothermophilus. Organism_taxid: 1422. Gene: pola. Expressed in: escherichia coli bl21. Expression_system_taxid: 511693.

Biol. unit:

Trimer (from PQS)

Resolution:

1.80Å R-factor: 0.218 R-free: 0.240

Authors:

J.J.Warren,L.J.Forsberg,L.S.Beese

Key ref:

J.J.Warren et al. (2006). The structural basis for the mutagenicity of O(6)-methyl-guanine lesions. Proc Natl Acad Sci U S A, 103, 19701-19706. PubMed id: 17179038 DOI: 10.1073/pnas.0609580103

Date:

28-Jun-06 Release date: 12-Dec-06

Protein chain

Q45458  (Q45458_GEOSE) -  DNA polymerase I from Geobacillus stearothermophilus

Seq: 876 a.a.

Struc: 580 a.a.*

* PDB and UniProt seqs differ at 67 residue positions (black crosses)

DNA/RNA chains

C-A-T-6OG-C-G-A-G-T-C-A-G-G 13 bases

C-C-T-G-A-C-T-C-G-C-A-T-G-A 14 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: 10.1073/pnas.0609580103

Proc Natl Acad Sci U S A 103:19701-19706 (2006)

PubMed id: 17179038

The structural basis for the mutagenicity of O(6)-methyl-guanine lesions.

J.J.Warren, L.J.Forsberg, L.S.Beese.

ABSTRACT

Methylating agents are widespread environmental carcinogens that generate a broad spectrum of DNA damage. Methylation at the guanine O(6) position confers the greatest mutagenic and carcinogenic potential. DNA polymerases insert cytosine and thymine with similar efficiency opposite O(6)-methyl-guanine (O6MeG). We combined pre-steady-state kinetic analysis and a series of nine x-ray crystal structures to contrast the reaction pathways of accurate and mutagenic replication of O6MeG in a high-fidelity DNA polymerase from Bacillus stearothermophilus. Polymerases achieve substrate specificity by selecting for nucleotides with shape and hydrogen-bonding patterns that complement a canonical DNA template. Our structures reveal that both thymine and cytosine O6MeG base pairs evade proofreading by mimicking the essential molecular features of canonical substrates. The steric mimicry depends on stabilization of a rare cytosine tautomer in C.O6MeG-polymerase complexes. An unusual electrostatic interaction between O-methyl protons and a thymine carbonyl oxygen helps stabilize T.O6MeG pairs bound to DNA polymerase. Because DNA methylators constitute an important class of chemotherapeutic agents, the molecular mechanisms of replication of these DNA lesions are important for our understanding of both the genesis and treatment of cancer.

Selected figure(s)

Figure 1.
Fig. 1. Structures of O6MeG·C (a) and O6MeG·T (b) pairs in DNA duplexes, unbound by protein. Hydrogen bonds are shown as dashed lines. (a Left) O6MeG·C wobble pair from ref. 27. (a Right) Model of Watson–Crick O6MeG·protonated C pair from ref. 52. (b) O6MeG·T pairs from refs. 29 and 30. The presence or absence of the H bond indicated by the "?" has been the subject of some controversy (29, 30).

Figure 3.
Fig. 3. T·O6MeG (a) and C·O6MeG (b) conformations in the –2 base pair position. Mesh shows 1 2 F[o] – F[c] electron density for the base pairs.

Figures were selected by an automated process.