abstract
- The binding of full and partial agonist ligands (L) to G protein-coupled receptors (GPCRs) initiates the formation of ternary complexes with G proteins [ligand-receptor-G protein (LRG) complexes]. Cyclic ternary complex models are required to account for the thermodynamically plausible complexes. It has recently become possible to assemble solubilized formyl peptide receptor (FPR) and beta(2)-adrenergic receptor (beta(2)AR) ternary complexes for flow cytometric bead-based assays. In these systems, soluble ternary complex formation of the receptors with G proteins allows direct quantitative measurements which can be analyzed in terms of three-dimensional concentrations (molarity). In contrast to the difficulty of analyzing comparable measurements in two-dimensional membrane systems, the output of these flow cytometric experiments can be analyzed via ternary complex simulations in which all of the parameters can be estimated. An outcome from such analysis yielded lower affinity for soluble ternary complex assembly by partial agonists compared with full agonists for the beta(2)AR. In the four-sided ternary complex model, this behavior is consistent with distinct ligand-induced conformational states for full and partial agonists. Rapid mix flow cytometry is used to analyze the subsecond dynamics of guanine nucleotide-mediated ternary complex disassembly. The modular breakup of ternary complex components is highlighted by the finding that the fastest step involves the departure of the ligand-activated GPCR from the intact G protein heterotrimer. The data also show that, under these experimental conditions, G protein subunit dissociation does not occur within the time frame relevant to signaling. The data and concepts are discussed in the context of a review of current literature on signaling mechanism based on structural and spectroscopic (FRET) studies of ternary complex components.