Protein-Protein Interactions Affect Native State Stability in Crowded Environments

Abstract

while the entropic change can even be unfavorable. To elucidate the molecular basis of these depletion interactions, we use simulations and analytic theory. Monte-Carlo simulations follow the association of two rod ''macromolecules'' in binary Lennard-Jones solutions. By dissecting the association free energy change into the respective thermodynamic components, we find different cosolutes induce stabilization through different thermodynamic driving mechanisms. Even for these simple liquids, considering intermolecular interactions beyond hard-cores can result in depletion forces that are completely enthalpic. We discuss how this newly resolved mechanism originates from intermolecular interactions and solvent restructuring. Finally, a mean-field theoretical model based on regular solution theories complements the simulation analysis. The dense, heterogeneous cellular environment is known to affect protein stability through interactions with other biomacromolecules. The effect of excluded volume due to these biomolecules, also known as crowding agents, on a protein of interest, or test protein, has long been known to increase the stability of a test protein. Recently, it has been recognized that attractive proteincrowder interactions play an important role. These interactions affect protein stability and can destabilize the test protein. However, most computational work investigating the role of attractive interactions has used spherical crowding agents and has neglected the specific roles of crowding agent hydrophobicity and hydrogen bonding. Here, we use multicanonical molecular dynamics and a coarse-grained protein model to study the folding thermodynamics of a small helical test protein in the presence of crowding agents that are themselves proteins. Our results show that the stability of the test protein depends on the hydrophobicity of the crowding agents. For low values of crowding agent hydrophobicity, the excluded volume effect is dominant and the test protein is stabilized relative to the dilute solution. For intermediate values of the crowding agent hydrophobicity, the test protein is destabilized by favorable side chainside chain interactions stabilizing the unfolded states. For high values of the crowding agent hydrophobicity, the native state is stabilized by the strong intermolecular attractions causing the formation of a packed structure that increases the stability of the test protein through favorable side chain-side chain interactions. In addition, increasing crowding agent hydrophobicity increases the ''foldability'' of the test protein and alters the potential energy landscape by simultaneously deepening the basins corresponding to the folded and unfolded states and increasing the energy barrier between them. Molecular chaperones are commonly identified by their ability to suppress heat induced protein aggregation. The muscle specific molecular chaperone UNC-45B is known to be critical in folding myosin and is trafficked to the sarcomeres A-band during thermal stress. Here, we identify thermally...