| A great amount of effort has been devoted to the development of structural models that predict ligand binding and selectivity for G-protein coupled receptors (GPCRs). Site-directed mutagenesis and chimeric studies, in conjunction with structure-activity relationships, have been instrumental in locating contacts of molecular recognition. With a better understanding of how a ligand interacts with its receptor, one can create more accurate molecular models and subsequently develop safer, more effective drugs.; To this end, we have examined the molecular recognition of two GPCR ligands. The first ligand, salvinorin A, is a kappa opioid receptor agonist whose analogs may have therapeutic value as non-addictive opioids. The second ligand, xanomeline, is a muscarinic (M1) agonist that may have therapeutic potential for the treatment of Alzheimer's disease. Due to the differences between these ligands structure and target, this dissertation is divided into two sections.; In part I, chimeric and site-directed mutagenesis data are utilized to develop a pharmacophore for salvinorin A. Collectively, the data indicates that Gln115 and Tyr119 in transmembrane helix II (TM II) and Tyr312, Tyr313, and Tyr320 in TM VII are important for salvinorin A binding. Additionally, the data suggests that extracellular loop-2 (EL-2) plays an indirect role in conferring selectivity. Based on the position of the implicated residues, a molecular model that aligns the ligand vertically between TM II and TM VII is proposed. This places salvinorin A in a unique binding epitope which may have implications for future opioid-based drug design.; In part II, various analogs of xanomeline are synthesized. These analogs, which incorporate polar and ionizable functional groups, have been designed to determine the role of hydrophobicity in xanomeline wash-resistant binding. Results indicate that several analogs with hydrophilic moieties, bind to the m1 receptor in a wash-resistant manner. Since it appears that the location the hydrophilic site can vary, it is suggested that wash resistant binding occurs at a receptor exosite that is flexible in nature. Although it is suggested that this allosteric site may be located in the extracellular loops, it is too premature to create a pharmacophoric model that implicates individual residues. |
