4O5K: Structure Of Human Dna Polymerase Complexed With N7mg In The Template Base Paired With Incoming Non-hydrolyzable Ctp

Citation:
Abstract
N7-Methyl-2'-deoxyguanosine (m7dG) is the predominant lesion formed by methylating agents. A systematic investigation on the effect of m7dG on DNA replication has been difficult due to the chemical instability of m7dG. To gain insights into the m7dG effect, we employed a 2'-fluorine-mediated transition-state destabilzation strategy. Specifically, we determined kinetic parameters for dCTP insertion opposite a chemically stable m7dG analogue, 2'-fluoro-m7dG (Fm7dG), by human DNA polymerase beta (polbeta) and solved three X-ray structures of polbeta in complex with the templating Fm7dG paired with incoming dCTP or dTTP analogues. The kinetic studies reveal that the templating Fm7dG slows polbeta catalysis approximately 300-fold, suggesting that m7dG in genomic DNA may impede replication by some DNA polymerases. The structural analysis reveals that Fm7dG forms a canonical Watson-Crick base pair with dCTP, but metal ion coordination is suboptimal for catalysis in the polbeta-Fm7dG:dCTP complex, which partially explains the slow insertion of dCTP opposite Fm7dG by polbeta. In addition, the polbeta-Fm7dG:dTTP structure shows open protein conformations and staggered base pair conformations, indicating that N7-methylation of dG does not promote a promutagenic replication. Overall, the first systematic studies on the effect of m7dG on DNA replication reveal that polbeta catalysis across m7dG is slow, yet highly accurate.
PDB ID: 4O5KDownload
MMDB ID: 121153
PDB Deposition Date: 2013/12/19
Updated in MMDB: 2014/08
Experimental Method:
x-ray diffraction
Resolution: 2.06  Å
Source Organism:
synthetic construct
Similar Structures:
Biological Unit for 4O5K: tetrameric; determined by author and by software (PISA)
Molecular Components in 4O5K
Label Count Molecule
Protein (1 molecule)
1
DNA Polymerase Beta(Gene symbol: POLB)
Molecule annotation
Nucleotides(3 molecules)
1
DNA (5'-d(*cp*cp*gp*ap*cp*(fmg) P*tp*cp*gp*cp*ap*tp*cp*ap*gp*c)-3')
Molecule annotation
1
DNA (5'-d(*gp*cp*tp*gp*ap*tp*gp*cp*gp*a)-3')
Molecule annotation
1
DNA (5'-d(p*gp*tp*cp*gp*g)-3')
Molecule annotation
Chemicals (5 molecules)
1
2
2
2
3
1
* Click molecule labels to explore molecular sequence information.

Citing MMDB
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