Plasticity and Co-Factor-Dependent Structural Changes in the RecA Nucleoprotein Filament Studied by Small-Angle X-Ray...

Area of Science:

• Structural biology
• Biophysics
• Molecular mechanisms

Background:

• Protein filament structural analysis faces challenges like crystallization difficulties and dynamic behavior.
• Cryo-electron microscopy provides static snapshots, but in-solution observations remain limited, especially for plastic filaments like RecA.
• RecA protein filaments are crucial for homologous recombination but their dynamic nature complicates structural studies.

Purpose of the Study:

• To determine the solution structures of two RecA nucleoprotein filament forms (ATP-bound and ADP-bound).
• To investigate the transition mechanism between these RecA filament forms in solution.
• To compare solution structures with existing crystal and cryo-EM data.

Main Methods:

• Small-angle X-ray scattering (SAXS) measurements.
• Integrative modeling using SAXS data and existing structural information.
• Monitoring structural transitions induced by varying magnesium ion concentrations.

Main Results:

• SAXS data validated the ATP-RecA filament crystal structure.
• SAXS profiles indicated differences in pitch and monomer-per-turn for the ADP-RecA filament compared to its crystal structure.
• A highly cooperative transition between ATP and ADP forms was observed upon addition of 0.3 mM Mg2+.

Conclusions:

• Integrative modeling and SAXS provide insights into RecA filament solution structures.
• The RecA filament undergoes a cooperative structural transition between ATP and ADP states.
• Understanding these transitions is key to elucidating RecA's role in homologous recombination.