Dynamic look at DNA unwinding by a replicative helicase.

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• DNA replication requires unwinding of duplex DNA by replicative hexameric helicases.
• Current mechanistic models for helicase function present contrasting views on DNA strand encirclement and ring orientation at the replication fork.

Purpose of the Study:

• To elucidate the DNA unwinding mechanism of the papillomavirus E1 helicase.
• To determine the number of DNA strands encircled by the E1 helicase ring and its orientation at the replication fork.

Main Methods:

• Single-molecule assays
• Ensemble assays
• Biochemical characterization of papillomavirus E1 helicase activity

Main Results:

• The E1 helicase unwinds DNA via a strand-exclusion mechanism.
• The N-terminal side of the E1 helicase ring is oriented towards the replication fork.
• E1 exhibits heterogeneous unwinding patterns due to repetitive movements modulated by its DNA-binding domain.

Conclusions:

• The papillomavirus E1 helicase employs a strand-exclusion mechanism with a specific orientation at the replication fork.
• Unrecognized dynamic aspects of replicative helicase unwinding are revealed, highlighting the role of repetitive movements and DNA binding.