Escherichia coli processivity clamp β from DNA polymerase III is dynamic in solution

Area of Science:

• Molecular Biology
• Biochemistry
• Structural Biology

Background:

• Escherichia coli DNA polymerase III requires the beta (β) clamp protein for high processivity during DNA replication.
• The β clamp is a ring-shaped homodimer, crucial for tethering DNA polymerases to DNA.
• Previous research has explored β clamp functions, but its solution dynamics remain poorly understood.

Purpose of the Study:

• To investigate the conformational dynamics of the intact β clamp dimer and a dimerization-impaired variant in solution.
• To characterize the flexibility and unfolding behavior of different β clamp domains.

Main Methods:

• Hydrogen-deuterium exchange mass spectrometry (HDX-MS) was employed.
• Conformational dynamics of the wild-type β clamp and an I272A/L273A variant were analyzed in solution.

Main Results:

• The β clamp exists as a dynamic, not a static, closed ring in solution.
• Domain I, involved in clamp opening, showed significant flexibility and partial unfolding (EX1 kinetics, ~4h half-life).
• The β clamp monomer variant exhibited greater dynamics than the wild-type dimer, with faster unfolding in Domain III (~1h half-life).

Conclusions:

• The β clamp's conformational flexibility is essential for its function.
• The δ subunit of the clamp loader may stabilize transient clamp opening rather than actively opening the ring.