Sugar-protein interactions control protein-complex stability in crowded Ficoll and dextran solutions

Traditional views of crowding emphasize excluded volume-stabilization of compact states-because they occupy less space, but the effects of high cosolute concentrations comprise several conjoined processes. We systematically explored the effects of dextrans, Ficolls, and their monomers, glucose and sucrose on the equilibrium thermodynamics of two protein complexes, both variants of the B1 domain of streptococcal protein G, a well-studied, small (6.2 kDa) globular protein. One is a simple side-by-side dimer of folded monomers. The other is a domain swap dimer, where, in buffer, the folded dimer dissociates to partially folded monomers. Dissociation was monitored as a function of cosolute molecular weight, cosolute concentration, and temperature using ¹⁹F NMR. Model fitting shows that, in contrast to sugar monomers whose interaction distance is dictated by their molecular volume, interaction distances of sugar polymers are dictated by their polymer structure and concentration. At high concentrations the effective length scale is the mesh size, emphasizing the decreasing effectiveness of large synthetic polymers. The model further dissects the measured parameters into fundamental processes: excluded volume, chemical interactions, and non-ideal mixing. For glucose and sucrose, dimer-stabilizing excluded volume contributions are nearly completely offset by destabilizing attractive chemical interactions with the monomers. As polymer molecular weight increases, the destabilizing effect of non-ideal mixing completely offsets the stabilizing effects of excluded volume. The results show that the crowding mechanism for polymers is distinct from those of their monomers and reflect their structural properties.