A highly conserved aspartic acid (Asp-155) anchors the terminal amine moiety of tryptamines and is involved in membrane targeting of the 5-HT2A serotonin receptor but does not participate in activation via a "salt-bridge disruption" mechanism

Discovering the molecular and atomic mechanism(s) by which G-protein-couple d receptors (GPCRs) are activated by agonists remains an elusive goal. Rece ntly, studies examining two representative GPCRs (rhodopsin and alpha(1b)-a drenergic receptors) have suggested that the disruption of a putative "salt -bridge" between highly conserved residues in transmembrane (TM) helix III, involving aspartate or glutamate, and helix VII, involving a basic residue , results in receptor activation. We have tested whether this is a general mechanism for GPCR activation by constructing a model of the 5-hydroxytrypt amine (5-HT)(2A) receptor and characterizing several mutations at the homol ogous residues (Asp-155 and Asn-363) of the 5-HT2A serotonin receptor. All of the mutants (D155A, D155N, D155E, D155Q, and S363A) resulted in receptor s with reduced basal activity; in no case was evidence for constitutive act ivity revealed. Structure-function studies with tryptamine analogs and vari ous Asp-155 mutants demonstrated that Asp-155 interacts with the terminal, and not indole, amine moiety of 5-HT2A agonists. Interestingly, the D155E m utation interfered with the membrane targeting of the 5-HT,2A receptor, and an inverse relationship was discovered when comparing receptor activation and targeting for a series of Asp-155 mutants. This represents the first kn own instance in which a charged residue located in a putative TM helix alte rs the membrane targeting of a GPCR. Thus, for 5-HT,2A receptors, the TMIII aspartic acid (Asp-155) is involved in anchoring the terminal amine moiety of indole agonists and in membrane targeting and not in receptor activatio n by salt-bridge disruption.