The G-Protein-Coupled Neuropeptide Y Receptor Type 2 is Highly Dynamic in Lipid Membranes as Revealed by Solid-State NMR Spectroscopy

In spite of the recent success in crystallizing several G-protein-coupled receptors (GPCRs), a comprehensive biophysical characterization of these molecules under physiological conditions also requires the study of the molecular dynamics of these proteins. The molecular mobility of the human neuropeptideY receptor type2 reconstituted into dimyristoylphosphatidylcholine (DMPC) membranes was investigated by means of solid-state NMR spectroscopy. Static (NNMR)-N-15 spectra show that the receptor performs axially symmetric motions in the membrane, and several residues undergo large amplitude fluctuations. This was confirmed by quantitative measurements of the motional H-1,C-13 order parameter of the CH, CH2, and CH3 groups. In directly polarized (CNMR)-C-13 experiments, these order parameters showed astonishingly low values of S-CH=0.55, SCH2=0.33, and SCH3=0.17, which corresponds to segmental amplitudes of approximately 50 degrees in the backbone and approximately 50-60 degrees in the side chain. At physiological temperature, (HNMR)-H-2 spectra of the deuterated receptor showed a narrow component that is indicative of molecular order parameters of S0.3 superimposed with a very broad spectrum that could stem from the transmembrane -helices. These results suggest that the crystal structures of GPCRs only represent a static snapshot of these highly mobile molecules, which undergo significant structural fluctuations with relatively large amplitudes in a liquid-crystalline membrane at physiological temperature.