β-Hairpin Crowding Agents Affect α-Helix Stability in Crowded Environments

Abstract

The dense, heterogeneous cellular environment is known to affect protein stability. It is now recognized that attractive "quinary" interactions with other biomacromolecules in the cell, referred to as the crowding agents, play a significant role in determining the stability of the protein of interest or test protein. These attractive interactions can reduce or overcome the stabilizing effect of the excluded volume of the crowding agents. However, the roles of specific interactions, such as hydrogen bonding and side chain-side chain hydrophobic interactions, are still unclear. Here, we use molecular simulation to investigate the roles played by hydrophobic interactions and hydrogen bonding between a small helical test protein and equally sized crowding agent proteins in a fixed β-hairpin configuration. The test protein and crowding agents are represented by a coarse-grained protein model, and we use multicanonical molecular dynamics to study the folding thermodynamics of the test protein. Our results confirm that the stability of the test protein depends on the hydrophobicity of the crowding agents and that the stability of the test protein is reduced through favorable side chain-side chain interactions that preferentially stabilize the unfolded states. In addition, we show that when the intermolecular hydrophobic interactions are more favorable than the intramolecular hydrophobic interactions, the β-rich crowding agents can completely destabilize the test protein, causing it to adopt configurations with increased β-content and preventing it from forming its native helical state. Similarities between our results and those seen in the formation of amyloid fibrils are also discussed.

Peptide folding in the presence of interacting protein crowders

“…Pioneering simulations of this kind, based on molecular dynamics methods, have recently been reported. 8,[35][36][37][38] The present study focuses on crowder-induced changes in the equilibrium properties of our two peptides. The problem of determining folding/unfolding equilibria in the presence of the crowders is tackled by direct Monte Carlo-based simulation.…”

Stabilizing or Destabilizing: Simulations of Chymotrypsin Inhibitor 2 under Crowding Reveal Existence of a Crossover Temperature

“…The stability of the native states of proteins immersed in a crowded macromolecular environment, such as that present in a living cell, can substantially differ from that in dilute conditions. Previous experiments − and simulations − showed that macromolecular crowding can lead to both stabilization and destabilization of the native fold. This variable outcome arises from an interplay of two phenomena, the stabilizing excluded volume effect and the potentially destabilizing effect of weak transient (quinary) interactions, and thus, it depends on the composition of the environment and on the amino acid sequence. ,,,− …”

Crowding in Cellular Environments at an Atomistic Level from Computer Simulations

“…97 A second theme is the modulation of secondary structure propensities as a result of crowder interactions. While this has been systematically analyzed using coarse-grained models, 95,96 atomistic simulations have provided evidence for an apparent stabilization of secondary structure elements, especially helices, as a result of crowding. 27,97,98,174 One way to understand that is via crowder amino acid side chain interactions that stabilize helices 95 or β-hairpin turns.…”