Enthalpic stabilization of an SH3 domain by D₂O

Abstract

The stability of a protein is vital for its biological function, and proper folding is partially driven by intermolecular interactions between protein and water. In many studies, H O is replaced by D O because H O interferes with the protein signal. Even this small perturbation, however, affects protein stability. Studies in isotopic waters also might provide insight into the role of solvation and hydrogen bonding in protein folding. Here, we report a complete thermodynamic analysis of the reversible, two-state, thermal unfolding of the metastable, 7-kDa N-terminal src-homology 3 domain of the Drosophila signal transduction protein drk in H O and D O using one-dimensional F NMR spectroscopy. The stabilizing effect of D O compared with H O is enthalpic and has a small to insignificant effect on the temperature of maximum stability, the entropy, and the heat capacity of unfolding. We also provide a concise summary of the literature about the effects of D O on protein stability and integrate our results into this body of data.

Direct observation of protein structural transitions through entire amyloid aggregation processes in water using 2D-IR spectroscopy

“…Accordingly, D 2 O effectively increases the hydrophobic interactions between amino acid residues in INS, thereby increasing the thermodynamic stability (Δ G u , free energy of unfolding) and enthalpy of unfolding (Δ H u ) of INS. 24,26 The helical propensity of INS in D 2 O lasts longer than that in H 2 O, which was observed in the time-resolved CD spectra and supports this supposition (Tables S1, S2 and Fig. S5 † ).…”

Selectivity for water isotopologues within metal organic nanotubes

“…1 Heavy water enriched in 2 D and/or 3 T forms stronger hydrogen bonds, which causes smaller amplitude intermolecular motions and more ordering within the solution phase. 2 This effect imparts differences in the physical properties of light and heavy water, including variations in the viscosity, 3 self-diffusion coefficient, 4 phase behavior, 5 hydration energies, 6 and exchange kinetics 7 that have observable effects on protein stability, 8 biological activity, 7 toxicity, 9 and chemical reactivity. 7 Nuclear quantum effects of water isotopologues are an active area of research and the effects on kinetics and thermodynamics in complex systems are critical if difficult to untangle.…”

Danio rerio Oocytes for Eukaryotic In-Cell NMR

“…Fluorine-labeled SH3 was prepared as described, − except that expression was allowed to proceed for 12–14 h at 18.5 °C. Fluorine-labeled N51K SH3 was expressed in the same way, except it was cloned into a pET28b (Novagen) vector with an N-terminal hexahistidine and SUMO tags, transformed into BL21 Star (DE3) One Shot cells (Invitrogen).…”