Controlling protein nanocage assembly with hydrostatic pressure

Controlling the assembly and disassembly of nanoscale protein cages is fundamental to the internalisation of protein and non-proteinaceous components for diverse bionanotechnological applications. To this end, here we study the reversible, pressure-induced dissociation of a natural protein nanocage, E. coli bacterioferritin (Bfr), principally using synchrotron radiation small angle X-ray scattering and circular dichroism. We demonstrate that hydrostatic pressures of 450 MPa are sufficient to completely dissociate the Bfr icositetramer into protein dimers, and the reversibility and kinetics of the reassembly process can be controlled by selecting appropriate buffer conditions. We also demonstrate that the heme B prosthetic group present at the subunit dimer interface influences the stability and pressure lability of the cage, despite its location being discrete from the inter-dimer interface that is key to cage assembly. This indicates a major cage-stabilising role for heme within this family of ferritins.