PT - JOURNAL ARTICLE AU - Jo Ann Janovick AU - Irina D Pogozheva AU - Henry I. Mosberg AU - P. Michael Conn TI - Salt Bridges Overlapping the GnRHR Agonist Binding Site Reveal a Coincidence Detector for GPCR Activation AID - 10.1124/jpet.111.180869 DP - 2011 Jan 01 TA - Journal of Pharmacology and Experimental Therapeutics PG - jpet.111.180869 4099 - http://jpet.aspetjournals.org/content/early/2011/04/28/jpet.111.180869.short 4100 - http://jpet.aspetjournals.org/content/early/2011/04/28/jpet.111.180869.full AB - Abstract GPCRs play central roles in most physiological functions and mutations in them cause heritable diseases. While crystal structures provide details about the structure of GPCRs, there is little information that identifies structural features which permit receptors to pass the cellular quality control system or which are involved in transition from the ground state to the ligand-activated state. The GnRHR, because of its small size among GPCRs, is amenable to molecular biological approaches and to computer modeling. These techniques and interspecies comparisons are used to identify structural features which are important for both intracellular trafficking and GnRHR activation, yet distinguish between these processes. Our model features two salt (R38-D98 and E90-K121) and two disulfide (C14-C200 and C114-C196) bridges, all of which are required for the hGnRHR to traffic to the plasma membrane. This study reveals that both constitutive and ligand-induced activation are associated with a "coincidence detector" which occurs when agonist binds. The observed constitutive activation of receptors lacking E90-K121, but not R38-D98 ionic bridge suggests the role of the former connection is holding the receptor in the inactive conformation. Both the aromatic ring and hydroxyl group of Y284, as well as hydrogen bonding of S217, are important for efficient receptor activation. Our modeling results, supported by the observed influence of K191 from extracellular loop 2 (EL2) and a four-residue motif surrounding this loop on ligand binding and receptor activation, suggest that the positioning of EL2 within the seven α-helical bundle regulates receptor stability, proper trafficking, and function.