Mutagenesis studies to determine the key interfacial amino acid residues that govern the self-assembly of the ferritin protein cages.
Date of Issue2011
School of Physical and Mathematical Sciences
Investigations into protein quaternary structure can lead to deeper insight into the fundamentals governing both protein folding and protein-protein interactions. In addition they can provide a foundation for the eventual rational design of novel complex protein architectures. A maxi-ferritin, bacterioferritin from E. coli (BFR), and a mini-ferritin, DNA-binding protein from starved cells (DPS), despite their similar four-helix bundle tertiary structure, assemble into quaternary structure with different symmetries, octahedral and tetrahedral and oligomerization states, 24-mer and 12-mer, respectively. To understand how these two structurally analogous proteins assemble into nano-structures with different sizes and shapes, both proteins were chosen as the basis for a mutagenesis study to investigate the importance of key amino acid residues, located at symmetry-related protein-protein interfaces, in controlling protein stability and self-assembly. Several mutants were designed for each protein through simple inspection and computational analysis, synthesized and subjected to different chemical and biophysical methods to determine their thermal stability, self-assemble ability and structure. The data indicate that many of these residues may be hot spot residues. Several mutants were observed to completely shut down detectable solution formation of 24-mer, favoring a cooperatively folded dimer, suggesting that they may be oligomerization “switch residues”. This investigation into the structure and energetics of these self-assembling nano-cage proteins not only can act as a jumping off point for the eventual design of novel protein nano-structures and their applications, but it can also help to understand the role that structure plays in the function of these important classes of proteins.