Most theories about macromolecular crowding focus on two ideas: the macromolecular nature of the crowder and entropy. For proteins, the volume excluded by the crowder favors compact native states over expanded denatured states, enhancing protein stability by decreasing the entropy of unfolding.
Researchers in the Pielak Group tested these ideas with the widely used crowding agent Ficoll-70 and its monomer, sucrose. Contrary to expectations, Ficoll and sucrose have approximately the same stabilizing effect on chymotrypsin inhibitor 2. Furthermore, the stabilization is driven by enthalpy, not entropy. These results point to the need for carefully controlled studies and more sophisticated theories for understanding crowding effects.
An understanding of cellular chemistry requires knowledge of how crowded environments affect proteins. The influence of crowding on protein stability arises from two phenomena, hard-core repulsions and soft, that is chemical, interactions. Most efforts to understand crowding effects on protein stability, however, focus on hard-core repulsions, which are inherently entropic and stabilizing.
Published in JACS, researchers in the Pielak Group assess these phenomena by measuring the temperature dependence of NMR-detected amide proton exchange and used these data to extract the entropic and enthalpic contributions of crowding to the stability of ubiquitin. Contrary to expectations, the contribution of chemical interactions is large and in many cases dominates the contribution from hardcore repulsions. The results show that both chemical interactions and hard-core repulsions must be considered when assessing the effects of crowding and help explain previous observations about protein stability and dynamics in cells.