Hydrophobic nanoparticles as bio-mimetic chaperones

Selective unfolding of native proteins may occur on the hydrophobic surface of nanoparticles, due to hydrophobic interactions among patches of hydrophobic residues in the protein core and the non polar surface of the nanoparticles. We reasoned that these same interactions may be helpful in dissociating hydrophobic bonds in the insoluble protein aggregates often formed as inclusion bodies when overexpressing proteins in heterologous systems. These aggregation events are hard to control, and are limiting the yield in native and functional proteins, in particular when proper folding requires uptake of cofactors. We report here on results obtained by stabilizing the unfolded form of the protein on the surface of geometrically suitable polystyrene nanoparticles, and by promoting subsequent refolding by supplying appropriate "structure-promoting" compounds. In this frame, we tested organic and inorganic cofactors, as appropriate to the specific protein under investigation, as well as substrates and their analogues in the case of enzymes. Under conditions where formation of a native structure is favored (e.g., in the presence of stabilizing agents), refolding of the adsorbed protein around the chemicals used as "structural primers" to promote refolding was proven to lead to formation of a proper hydrophobic core at the protein interior. As anticipated, burial of the exposed hydrophobic sites uopn refolding led to the release of the folded protein from the nanoparticle surface. Results will be discussed in terms of their possible practical relevance, and in terms of the specific conditions required for refolding of individual protein classes.