4P2H

Structure of human DNA polymerase complexed with N7MG in the template opposite to incoming non-hydrolyzable TTP with manganese in the active site

Summary for 4P2H

• Entry DOI: 10.2210/pdb4p2h/pdb
• Related: 1bpx
• Descriptor: DNA polymerase beta, DNA (5'-D(*CP*CP*GP*AP*CP*(FMG)P*TP*CP*GP*CP*AP*TP*CP*AP*GP*C)-3'), DNA (5'-D(*GP*CP*TP*GP*AP*TP*GP*CP*GP*A)-3'), ... (8 entities in total)
• Functional Keywords: nucleotidyl transfer, dna binding, dna polymerase fold, transferase-dna complex, transferase/dna
• Biological source: Homo sapiens (Human); More
• Total number of polymer chains: 4
• Total formula weight: 47317.68

Authors

Lee, S.,Koag, M.C. (deposition date: 2014-03-04, release date: 2014-06-25, Last modification date: 2023-09-27)

Primary citation

Koag, M.C.,Kou, Y.,Ouzon-Shubeita, H.,Lee, S.
Transition-state destabilization reveals how human DNA polymerase beta proceeds across the chemically unstable lesion N7-methylguanine.
Nucleic Acids Res., 42:8755-8766, 2014

PubMed 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 β (polβ) and solved three X-ray structures of polβ in complex with the templating Fm7dG paired with incoming dCTP or dTTP analogues. The kinetic studies reveal that the templating Fm7dG slows polβ catalysis ∼ 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 polβ-Fm7dG:dCTP complex, which partially explains the slow insertion of dCTP opposite Fm7dG by polβ. In addition, the polβ-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 polβ catalysis across m7dG is slow, yet highly accurate.

PubMed: 24966350
DOI: 10.1093/nar/gku554

Experimental method

X-RAY DIFFRACTION (1.987 Å)