Temperature-independent solubility and interactions between apoferritin monomers and dimers in solution

We used chromatographic, static and dynamic light scattering techniques, an d atomic force microscopy (AFM) to study the structure of the protein speci es and the protein-protein interactions in solutions containing two apoferr itin molecular forms, monomers and dimers, in the presence of NaAc buffer a nd CdSO4. The sizes and shapes of the monomers and dimers, separated by siz e-exclusion chromatography, were determined by dynamic light scattering and AFM. While the monomer is an apparent sphere with a diameter corresponding to previous X-ray crystallography determinations, the dimer shape correspo nds to two, bound monomer spheres. Static light scattering was used to char acterize the interactions between solute molecules of monomers and dimers i n terms of the second osmotic virial coefficients. The addition of even sma ll amounts of Cd2+ causes attraction between the monomer molecules. Further more, we found that the second virial coefficient and the protein solubilit y do not noticeably depend on temperature in the range from 0 degreesC to 4 0 degreesC. This suggests that the enthalpy for crystallization of apoferri tin is close to zero, and the gain of entropy is essentially constant in th is temperature range. We also found that in solutions of the apoferritin di mer, the molecules attract even in the presence of acetate buffer only, ind icating a change in the surface of the apoferritin molecule. In view of the repulsion between the monomers at the same conditions, this suggests that the dimers and higher oligomers form only after partial unfolding of some o f the apoferritin subunits. These observations suggest that aggregation and self-assembly of protein molecules or molecular subunits may be driven by forces other than those responsible for crystallization in the protein solu tion. (C) 2001 Elsevier Science B.V. All rights reserved.