Extracellular domains of the neurokinin-1 receptor: structural characterization and interactions with substance P

The technical difficulties associated with the structure determination of membrane proteins have limited the structural information available for the ligand binding to G-protein coupled receptors (GPCRs). Here, we describe a reductionist approach to GPCR structure determination in which the extracellular domains of the receptor are examined by high-resolution NMR in the presence of a membrane mimetic. The resulting structural features are then incorporated into a molecular model of the receptor, utilizing the x-ray structure of rhodopsin to generate the topological orientation of the transmembrane helices. The results of our study of the neurokinin-1 receptor (NK-1R) and its interactions with substance P (SP) are detailed here. The structure of the N-terminus, NK-1R(1-39), and of the third extracellular loop, NK-1R(264-290), in the presence of dodecylphosphocholine micelles is described. Our findings provide a structural basis for the interpretation of the results from other methods including mutagenesis, fluorescence, and photoaffinity labeling experiments, resulting in an experimentally based, high-resolution model of SP binding to NK-1R.