Water-Mediated Protein-Protein Interactions at High Pressures are Controlled by a Deep-Sea Osmolyte

The influence of natural cosolvent mixtures on the pressure-dependent structure and protein-protein interaction potential of dense protein solutions is studied and analyzed using small-angle X-ray scattering in combination with a liquid-state theoretical approach. The deep-sea osmolyte trimethylamine-$N$-oxide is shown to play a crucial and singular role in its ability to not only guarantee sustainability of the native protein’s folded state under harsh environmental conditions, but it also controls water-mediated intermolecular interactions at high pressure, thereby preventing contact formation and hence aggregation of proteins.

Figure 1

Structure of $\alpha$ and $\beta$ amino acids, the denaturant urea, carbohydrates, and methylamines present as osmolytes in marine organisms. $\mathrm{GPC}=\mathrm{L}\text{−}\alpha$-glycerophosphorylcholine.

Figure 2

(a) Experimental SAXS data $I(q)$ for a 10% (w/v) lysozyme in 25 mM Bis-Tris solution at $25°\mathrm{C}$ and 1 bar for various trehalose concentrations $c$. Black lines display the refinement of the data. (b) Results of the refinement for the effective structure factor $S_{\text{eff}}(q)$ at various trehalose concentrations. (c) Strength of the attractive interaction $J(c,p)$ as a function of trehalose concentration $c$ and pressure $p$. (d) Total protein-protein interaction potential $V(r)$ (sum) displayed together with its contributing parts.

Figure 3

(a) Results of the refinement for the strength of the attractive part $J$ and the normalized second virial coefficient $b_2$ as a function of osmolyte concentration $c$ (in $\mathrm{M}=\mathrm{mol}{\mathrm{L}}^{-1}$), at $25°\mathrm{C}$ and ambient pressure, and as a function of pressure for midlevel (b) and highest (c) osmolyte concentrations. For comparison, the pure buffer system is depicted as well (black circle, solid line).

Figure 4

Pressure dependence of the strength of the attractive protein-protein interaction $J$ in 10% (w/v) lysozyme $+25\mathrm{mM}$ Bis-Tris ($p\mathrm{H}$ 7) at $25°\mathrm{C}$ with osmolyte mixtures added as found for shallow-living (top half) and deep-living (bottom half) crabs (a), shrimps (b), and skates (c) [11]. The results for the pure buffer (solid black line) and for the contributing single cosolvent solutions are shown as well.