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Thermophilic Ferritin 24mer Assembly and Nanoparticle Encapsulation Modulated by Interdimer Electrostatic Repulsion
Katherine W Pulsipher1, Jose A Villegas1, Benjamin W Roose1
1Department of Chemistry, University of Pennsylvania , 231 South 34th Street, Philadelphia, Pennsylvania 19104, United States.
Biochemistry
|July 7, 2017
Summary
Researchers engineered protein cages by modifying Archaeoglobus fulgidus ferritin (AfFtn). Mutants showed altered assembly, with E65R forming stable cages across ionic strengths, demonstrating control over protein cage properties.
Area of Science:
- Biotechnology
- Structural Biology
- Protein Engineering
Background:
- Protein cages self-assemble for encapsulating diverse molecules, with Archaeoglobus fulgidus ferritin (AfFtn) assembly influenced by ionic strength and charged nanoparticles.
- Understanding the specific residues that control AfFtn assembly is crucial for designing tailored protein cages.
Purpose of the Study:
- To computationally design and characterize AfFtn mutants with altered electrostatic interactions at dimer-dimer interfaces.
- To investigate how specific charge mutations affect protein cage assembly, stability, and encapsulation capabilities.
Main Methods:
- Computational design of three AfFtn mutants (E65R, D138K, A127R) introducing positive charges at dimer-dimer interfaces.
- Analysis of mutant assembly kinetics, thermodynamics, and 24mer propensity across varying NaCl concentrations.
- X-ray crystallography to determine the structural basis of assembly changes.
- Assessment of gold nanoparticle (AuNP) encapsulation efficiency.
Main Results:
- Mutants displayed varied 24mer assembly propensity: A127R < wild type < D138K ≪ E65R.
- The E65R mutant assembled into a stable 24mer cage across 0-800 mM NaCl, with a high dissociation temperature (98 °C) and a compact structure.
- While A127R and D138K encapsulated AuNPs similarly to wild-type AfFtn, E65R showed reduced encapsulation yields.
Conclusions:
- Designed mutations can precisely control protein cage assembly and stability.
- The E65R mutant represents a robust protein cage platform with tunable properties for specific applications.
- This study provides insights into structure-function relationships for ferritin self-assembly and design.
