Structural descriptions of ligands in their binding site of integral membrane proteins at near physiological conditions using solid-state NMR

• Published in: European biophysics journal 1998-01, Vol.28 (1), p.84-90
• Main Authors: Watts, A., Burnett, I. J., Glaubitz, Clemens, Gröbner, Gerhard, Middleton, David A., Spooner, Paul J. R., Williamson, Philip T. F.
• Format: Article
• Language: English
• Subjects: Binding sites; Biophysics; Membranes; NMR; Nuclear magnetic resonance; Pharmacology; Physiology; Proteins
• ISSN: 0175-7571; 1432-1017
• DOI: 10.1007/s002490050187

Using solid-state NMR approaches, it is now possible to define the structure and dynamics of binding for a small, isotopically (^sup 2^H, ^sup 13^C, ^sup 19^F, ^sup 15^N) labelled ligand, prosthetic group or solute in its binding site of a membrane-bound protein at near physiological conditions in natural membrane fragments or in reconstituted complexes. Studies of oriented membranes permit the orientational bond vectors of labelled groups to be determined to good precision, as shown for retinal in bacteriorhodopsin and bovine rhodopsin. Using novel magic angle spinning NMR methods on membrane dispersions, high-resolution NMR spectra can be obtained. Dipolar couplings can be reintroduced into the spectrum of labelled ligands in their binding sites of membrane-bound proteins to give interatomic distances to high precision (±0.5 Å). Relaxation and cross-polarization data give estimates for the kinetics for on-off rates for binding. In addition, chemical shifts can be measured directly to help provide details of the binding environment for a bound ligand, as shown for analogues of drugs used in peptic ulcer treatment in the gastric ATPase, and for acetylcholine in the acetylcholine receptor.[PUBLICATION ABSTRACT]