NMR identification of hydrophobic cavities with low water occupancies in protein structures using small gas molecules.

Magnetization transfer through dipole-dipole interactions (nuclear Overhauser effects, NOEs) between water protons and the protons lining two small hydrophobic cavities in hen egg-white lysozyme demonstrates the presence of water molecules with occupancies of approximately 10-50%. Similarly, NOEs were observed between the cavity protons and the protons of hydrogen, methane, ethylene or cyclopropane applied at 1-200 bar pressure. These gases can thus be used as general NMR indicators of empty or partially hydrated hydrophobic cavities in proteins. All gases reside in the cavities for longer than 1 ns in marked contrast to common belief that gas diffusion in proteins is not much slower than in water. Binding to otherwise empty cavities may be a major aspect of the anesthetic effect of small organic gas molecules.