PROPOSED LIGAND-BINDING SITE OF THE TRANSMEMBRANE RECEPTOR FOR NEUROTENSIN(8-13)

We report here the first proposed ligand binding site of the transmemb rane receptor for neurotensin (8-13) in human and rat, the correspondi ng bound conformation of the peptide ligand, and site directed mutagen esis studies that support the binding site model, These three-dimensio nal structures were generated by using a heuristic approach in conjunc tion with experimental data. The proposed neurotensin(8-13) binding si te is primarily composed of eight residues (i.e., Phe(326), Ile(329), Trp(334), Phe(337), Tyr(339), Phe(341), Tyr(342), and Tyr(344) in the human receptor; Phe(331), Ile(334), Trp(339), Phe(342), Phe(344), Phe( 346), Tyr(347), and Tyr(349) in the rat receptor) located in the third extracellular loop, The seven aromatic residues form an aromatic pock et on the extracellular surface of the neurotensin receptor to accommo date its ligands apparently by cation-pi, pi-pi, and hydrogen bonding interactions, The neurotensin(8-13) ligand adopts a compact conformati on at the proposed binding site, In the bound conformation of neuroten sin(8-13), the backbone of Arg(9)-Pro(10)-Tyr(11)-Ile(12) forms the pr oline type I turn, and the hydroxy group of Tyr(11) interacts with the two guanidinium groups of Arg(8) and Arg(9). These guanidinium groups are curled toward the hydroxy group so that they interact electrostat ically with the hydroxy group, and that the guanidinium group of Arg(9 ) forms an intra-hydrogen bond with the hydroxy group. The proposed th ree-dimensional structure may not only provide a basis for rationalizi ng mutations of the neurotensin receptor gene but also offer insights into understanding the binding of many neurotensin analogs, biological functions of the neurotensin receptors, and structural elements for s pecies specificity of the neurotensin receptors, and may expedite deve loping nonpeptidic neurotensin mimetics for the potential treatment of the neuropsychiatric diseases.