How Osmolytes Counteract Pressure Denaturation on a Molecular Scale

Abstract

Life in the deep sea exposes enzymes to high hydrostatic pressure which decreases their stability. For survival, deep sea organisms tend to accumulate various osmolytes, most notably trimethylamine N-oxide (TMAO) used by fish, to counteract pressure denaturation. Yet, exactly how they work still remains unclear. Here, we use a rigorous statistical thermodynamics approach to clarify the mechanism of osmoprotection. We show that the weak, non-specific, and dynamic interactions of water and osmolytes with proteins can be characterized only statistically, and that the competition between protein-osmolyte and protein-water interactions is crucial in determining conformational stability. Osmoprotection is driven by a stronger exclusion of osmolytes from the denatured protein than from the native conformation, and the water distribution has no significant effect on these changes for low osmolyte concentrations.

Temperature, Hydrostatic Pressure, and Osmolyte Effects on Liquid–Liquid Phase Separation in Protein Condensates: Physical Chemistry and Biological Implications

“…[101] The unfavorability of protein-osmolyte interactions leads to preferentiale xclusion of osmolytes from protein surfaces and more favorable free energies for compact configurations. [102] Ap lausible physicalr ationalization of the combinedp ressure and osmolyte dependence of gD-crystallin LLPS, which needs to account also for the role of void volumes [98] in pressures ensitivity,w ill be discussed in Section4 below.…”

“…[183][184][185][186][187] Nonetheless, it is generally agreedt hat TMAO confers stability to more compact protein configurations because it interacts unfavorably with proteins, av iew that is supported by the observation that TMAO is depleted around protein surfaces, but forms strongh ydrogen bonds to water. [102,183,188] The degree of unfavorable interaction between TMAO and different amino acid residues differs as it depends on the residue involved. Nonetheless, experimental evidencep oints to ag eneralo verall exclusion of TMAO from both hydrophobic and polar surfaces of proteins.…”

“…The general stabilization effect of osmolytes on protein configurational states with less solvent exposures relative to states with high solvent exposures, such as stabilization of folded protein against unfolding, and stabilization of condensed LLPS states against dissolution, is likely related to the fact that interactions between osmolytes and with various components of polypeptide chains are generally unfavorable . The unfavorability of protein–osmolyte interactions leads to preferential exclusion of osmolytes from protein surfaces and more favorable free energies for compact configurations . A plausible physical rationalization of the combined pressure and osmolyte dependence of γD‐crystallin LLPS, which needs to account also for the role of void volumes in pressure sensitivity, will be discussed in Section 4 below.…”

“…There is no general consensus on the detailed structural and energetic mechanism by which TMAO affects solvent‐mediated biomolecular interactions because results from atomic simulations are highly sensitive to the forcefield employed . Nonetheless, it is generally agreed that TMAO confers stability to more compact protein configurations because it interacts unfavorably with proteins, a view that is supported by the observation that TMAO is depleted around protein surfaces, but forms strong hydrogen bonds to water . The degree of unfavorable interaction between TMAO and different amino acid residues differs as it depends on the residue involved.…”

Ammonium based stabilizers effectively counteract urea-induced denaturation in a small protein: insights from molecular dynamics simulations

“…14 Using neutron scattering experiments TMAO was earlier shown to interact directly with urea and offset its action on protein by Meersman et al 15 On the contrary, both experimental 16 as well as computational 17 studies have suggested that the enhancement of waterstructuring by TMAO is responsible for the preservation of the protein in its native state, not only from chemical denaturation but also from pressure induced structural collapse. 18 A very recent study by Dias and co-workers shows a competing mechanism of TMAO on Trp cage mini proteins, which stabilizes charge-charge interactions, induces collapse of the backbone and causes swelling of peptide with greater degree of hydrophobic residue. 19 Among other osmolytes, trehalose mediated counteraction of the effect of urea on a small peptide has been investigated thoroughly by Paul and Paul using extensive MD simulations.…”