Mechanisms of ferritin assembly studied by time-resolved small-angle X-ray scattering

Area of Science:

• Biochemistry
• Structural Biology
• Biophysics

Background:

• Escherichia coli ferritin A (EcFtnA) self-assembles into a 24-subunit cage.
• Ferritin assembly is pH-dependent, with dissociation into dimers at acidic pH and reassembly at neutral pH.
• Understanding ferritin assembly mechanisms is crucial for its biological functions.

Purpose of the Study:

• To investigate the assembly kinetics of EcFtnA using time-resolved small-angle X-ray scattering (SAXS).
• To elucidate the role of ionic strength, pH, and electrostatic interactions in EcFtnA assembly.
• To examine the influence of net charge on ferritin subunit assembly rates.

Main Methods:

• Time-resolved small-angle X-ray scattering (SAXS) was employed to monitor EcFtnA assembly.
• Kinetic analysis of SAXS data was performed to determine assembly rates.
• Ferritin mutants with varying net charges were engineered and analyzed.

Main Results:

• Ferritin assembly intermediates (tetramers, hexamers, dodecamers) were modeled from SAXS data.
• Assembly rate increased with ionic strength and decreased with increasing pH (pH 6-8).
• Net charge repulsion between dimers significantly influenced assembly rates, particularly at low ionic strength.

Conclusions:

• Electrostatic repulsion between charged subunits is a key factor governing EcFtnA assembly kinetics.
• Ionic strength modulates the effect of electrostatic interactions on assembly.
• The study provides insights into the physical forces driving ferritin self-assembly.