Hydrogen bond strength in membrane proteins probed by time-resolved 1 H-detected solid-state NMR and MD simulations.

1 H-detected solid-state NMR in combination with 1 H/2 D exchange steps allows for the direct identification of very strong hydrogen bonds in membrane proteins. On the example of the membrane-embedded potassium channel KcsA, we quantify the longevity of such very strong hydrogen bonds by combining time-resolved 1 H-detected solid-state NMR experiments and molecular dynamics simulations. In particular, we show that the carboxyl-side chain of the highly conserved residue Glu51 is involved in ultra-strong hydrogen bonds, which are fully-water-exposed and yet stable for weeks. The astonishing stability of these hydrogen bonds is important for the structural integrity of potassium channels, which we further corroborate by computational studies.