ceptors (GPCR 4 ). The b 2 receptor was first cloned in A modified human b 2 receptor, designated 0K-b 2 , was 1986 (1) and serves as a model for ligand binding and developed for site-specific labeling at the amino termisignal transduction for other neurotransmitter and nus with amine reactive fluorescent probes. 0K-b 2 has hormone receptors in the GPCR superfamily (2). The b 2 the following modifications: (1) all 16 lysines in the wildreceptor is an integral membrane protein with primary type b 2 receptor were mutated to arginines, (2) a FLAG amino acid sequence homology to the light-activated epitope preceded by a cleaved hemagglutinin signal sephotoreceptor, rhodopsin. Based on the hydropathy quence was fused to the amino terminus, and (3) a hexaprofile similarity of primary amino acid sequences, the histidine tail was added to the carboxyl terminus. The topology of the b 2 receptor is similar to bacteriorhodop-FLAG epitope and hexahistidine tail were added to fasin (3) and rhodopsin (4) with seven membrane-spancilitate purification while lysine to arginine mutations ning helices. Both biochemical (5, 6) and immunologic eliminate potential labeling sites for amine-reactive (7) evidence supports this proposed topology. fluorescent probes. The remaining primary amines in We are interested in developing approaches for the 0K-b 2 receptor, the amino terminal amine and the studying the structure and dynamics of the human b 2 e-amine of Lys 3 , both reside in the amino-terminal FLAG adrenergic receptor. Protein fluorescence spectroscopy epitope. The 0K-b 2 receptor expressed in Sf9 insect cells is a sensitive technique that can be used to obtain both exhibited ligand binding and G-protein coupling charstructural and dynamic information from purified proacteristics similar to the wild-type b 2 receptor. The modteins (see (8) for review). Intrinsic protein fluorescence ified receptor was labeled with fluorescamine, an is primarily due to tryptophan, and the cloned human amine-reactive fluorescent probe. Proteolysis with facb 2 receptor has eight tryptophans distributed throughtor Xa showed that labeling was confined to the amino out the protein (9). Because of the number and distributerminus of the 0K-b 2 receptor. Our results demonstrate site-specific fluorescamine labeling at the amino termi-tion of tryptophans in the b 2 receptor, tryptophan fluonus of the 0K-b 2 receptor, a lysine-depleted b 2 receptor rescence is not useful for detecting conformational that retains functional characteristics of the wild-type changes in specific structural domains. As an alternareceptor.