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.