Figure 4.

Comparison of effects of quenchers localized to the micelle on the response of FM-β₂R to ISO. (A) Schematic depicting the structure of CAT-16 and 5-DOX, as well as the putative location of these quenching groups in the micelle. The quenching group on CAT-16 is localized on the polar surface of the micelle. The quenching group on 5-DOX is located within the hydrophobic core of the micelle. (B) Stern–Volmer plots depicting the extent of quenching of FM-β₂AR by increasing concentrations of CAT-16 or 5-DOX. Quenchers were added to labeled receptor, and fluorescence was measured and plotted as in Fig. 3 and Materials and Methods. The total lipid concentration was kept constant at 100 μM with stearic acid. The quenching constant K_sv was 2.4 ± 0.1 mM⁻¹ in the presence of CAT-16 and 1.4 ± 0.2 mM⁻¹ in the presence of 5-DOX. (C) Differing effects of CAT-16 and 5-DOX on agonist-induced fluorescence change of FM-β₂AR. The extent of response to ISO is presented as a percentage of control ISO response, calculated as in Fig. 3. (D) An example of the experiments used to generate the ratios in C. In this example, FM-β₂AR was incubated with either 100 μM CAT-16 or 100 μM stearic acid. The response to agonist was monitored as described in Fig. 2. In the presence of the quencher CAT-16, ISO induced a 24.2 ± 0.3% decrease in fluorescence vs. 4.1 ± 0.6% in the presence of the stearic acid. All values are mean ± SEM, n = 3.