Mixture of Macromolecular Crowding Agents Has a Non-additive Effect on the Stability of Proteins.

The folding and unfolding of proteins inside a cell take place in the presence of macromolecules of various shapes and sizes. Such crowded conditions can significantly affect folding, stability, and biophysical properties of proteins. Thus, to logically mimic the intracellular environment, the thermodynamic stability of two different proteins (lysozyme and α-lactalbumin) was investigated in the presence of mixtures of three crowding agents (ficoll 70, dextran 70, and dextran 40) at different pH values. These crowders possess different shapes and sizes. It was observed that the stabilizing effect of mixtures of crowders is more than the sum effects of the individual crowder, i.e., the stabilizing effect is non-additive in nature. Moreover, dextran 40 (in the mixture) has been found to exhibit the greatest stabilization when compared with other crowders in the mixture. In other words, the small size of the crowder has been observed to be a dominant factor in stabilization of the proteins. Graphical Abstract.