Ligand binding properties of β2ARGsα and β2ARCAMGsα expressed in Sf9 membranes: effect of GTPγS
| Ligand | Kh | Kl | Rh | KhGTPγS | KlGTPγS | RhGTPγS |
|---|---|---|---|---|---|---|
| nM | % | nM | % | |||
| β2ARGsα | ||||||
| (−)-ISO | 1.0 ± 0.6 | 100 ± 38 | 44.3 ± 3.8 | 190 ± 20 | ||
| (+)-ISO | 17 ± 4 | 3400 ± 310 | 38.9 ± 4.4 | 4300 ± 500 | ||
| SAL | 21 ± 5 | 2000 ± 450 | 37.7 ± 4.4 | 2900 ± 650 | ||
| DOB | 62 ± 23 | 2300 ± 600 | 45.0 ± 5.1 | 2800 ± 660 | ||
| EPH | 3000 ± 1100 | 15000 ± 2000 | ||||
| DCI | 96 ± 17 | 140 ± 20 | ||||
| ICI | 1.7 ± 0.3 | 1.9 ± 0.4 | ||||
| β2ARCAMGSα | ||||||
| (−)-ISO | 0.2 ± 0.1 | 3.8 ± 1.3 | 35.4 ± 4.0 | 2.0 ± 1.0 | 13 ± 2.2 | 65.5 ± 8.9 |
| (+)-ISO | 3.8 ± 1.0 | 75 ± 13 | 56.1 ± 5.3 | 135 ± 21 | ||
| SAL | 2.2 ± 1.1 | 76 ± 16 | 43.6 ± 5.1 | 1.9 ± 0.9 | 160 ± 20 | 16.3 ± 3.3 |
| DOB | 16 ± 4 | 1400 ± 430 | 80.0 ± 6.8 | 500 ± 170 | ||
| EPH | 32 ± 12 | 1800 ± 400 | 56.0 ± 8.9 | 2900 ± 600 | ||
| DCI | 12 ± 4 | 240 ± 80 | 48.8 ± 4.9 | 470 ± 120 | ||
| ICI | 3.4 ± 0.4 | 1.4 ± 0.4 | ||||
[3H]DHA binding was determined as described underExperimental Procedures in Sf9 membranes expressing β2ARGsα (3.3–7.5 pmol/mg) or β2ARCAMGsα (2.5–4.9 pmol/mg). The ligand competition binding curves shown in Figs. 2, 5, and 7 were analyzed by nonlinear regression for best fit to single-site or two-site binding.Kh and Kl designate the dissociation constants for high- and low-affinity agonist binding, respectively. %Rh indicates the percentage of receptors displaying high agonist affinity. The corresponding values obtained in the presence of GTPγS (10 μM) are designatedKhGTPγS, KlGTPγS, and %RhGTPγS, respectively. Dissociation constants for ICI are listed below Kl andKlGTPγS, respectively. Data shown represent the means ± S.D. of four to seven independent experiments performed in triplicate.