Bulky Lesion Bypass Requires Dpo4 Binding in Distinct Conformations

Area of Science:

• Molecular Biology
• Biochemistry
• DNA Repair Mechanisms

Background:

• Translesion DNA synthesis (TLS) is crucial for resuming replication at stalled forks.
• Polycyclic aromatic hydrocarbons (PAHs) like Benzo[a]pyrene (B[a]P) form carcinogenic DNA adducts.
• The Y-family polymerase Dpo4 is involved in bypassing DNA lesions.

Purpose of the Study:

• To investigate the mechanism of Dpo4 bypassing a (+)-cis-B[a]P-N 2-dG adduct.
• To understand the role of environmental factors in adduct bypass.
• To elucidate the conditions for accurate versus error-prone replication past adducts.

Main Methods:

• Single-molecule fluorescence resonance energy transfer (smFRET) experiments.
• Classical molecular dynamics simulations.
• Nucleotide incorporation assays.

Main Results:

• The B[a]P moiety of the adduct blocks nucleotide incorporation when stacked within the DNA helix.
• Dimethyl sulfoxide (DMSO) promotes a solvent-exposed conformation of the adduct.
• DMSO enables error-prone replication past the adduct and accurate incorporation when primer-terminated.

Conclusions:

• The conformation of the B[a]P adduct is critical for Dpo4-mediated translesion synthesis.
• DMSO acts as an allosteric effector, facilitating adduct bypass.
• Modulating the cellular environment could influence the fidelity of DNA repair past carcinogen adducts.