Table 1.
Pharmacological properties of Cys-substituted mutant M3Rs
| Receptor | Ballesteros- Weinstein number | [3H]-NMS binding |
Carbachol binding, Ki (μM) | Carbachol-induced inositol monophosphate production |
|||
|---|---|---|---|---|---|---|---|
| KD (nM) | Bmax (pmol/mg protein) | EC50 (nM) | Emax (fold above basal) | Basal activity (%) | |||
| M3′(3C)-Xa | 1.45 ± 0.17 | 9.29 ± 0.35 | 34.3 ± 4.8 | 47 ± 14 | 4.5 ± 0.7 | 100 | |
| R252C | 5.60 | 1.68 ± 0.33 | 3.51 ± 0.35 | 67.5 ± 14.5 | 127 ± 28 | 3.6 ± 1.5 | 92 ± 13 |
| I253C | 5.61 | 1.84 ± 0.53 | 7.63 ± 2.21 | 74.4 ± 22.7 | 177 ± 56 | 4.8 ± 2.6 | 91 ± 9 |
| Y254C | 5.62 | 1.50 ± 0.51 | 4.83 ± 1.47 | 48.8 ± 5.9 | 338 ± 101 | 4.0 ± 2.6 | 112 ± 34 |
| K255C | 5.63 | 0.91 ± 0.26 | 9.58 ± 1.13 | 41.3 ± 0.2 | 56 ± 13 | 3.9 ± 2.0 | 78 ± 14 |
| E256C | 5.64 | 1.59 ± 0.54 | 6.26 ± 0.64 | 12.2 ± 1.6 | 9.7 ± 2.2 | 4.5 ± 1.6 | 132 ± 42 |
| T257C | 5.65 | 1.05 ± 0.48 | 7.96 ± 1.14 | 33.5 ± 5.7 | 16 ± 6.9 | 2.9 ± 0.7 | 103 ± 9 |
| E258C | 5.66 | 1.02 ± 0.65 | 8.73 ± 1.06 | 22.2 ± 7.9 | 5.3 ± 2.0 | 4.4 ± 0.1 | 203 ± 40 |
| K259C | 5.67 | 0.98 ± 0.47 | 8.25 ± 0.70 | 25.2 ± 3.0 | 24 ± 6.8 | 3.1 ± 0.6 | 86 ± 20 |
| R260C | 5.68 | 0.91 ± 0.48 | 6.57 ± 0.98 | 36.9 ± 6.3 | 436 ± 180 | 2.2 ± 1.0 | 73 ± 17 |
| T261C | 5.69 | 0.86 ± 0.33 | 7.83 ± 0.91 | 33.7 ± 6.6 | 62 ± 13 | 2.9 ± 0.4 | 83 ± 24 |
| K262C | 5.70 | 0.87 ± 0.15 | 5.41 ± 0.16 | 62.4 ± 16.9 | 11 ± 2.6 | 2.8 ± 1.1 | 70 ± 4 |
| E263C | 5.71 | 1.16 ± 0.31 | 6.16 ± 0.54 | 26.8 ± 1.0 | 10 ± 5.6 | 2.7 ± 0.7 | 78 ± 13 |
| L264C | 5.72 | 1.24 ± 0.23 | 6.64 ± 0.22 | 20.9 ± 0.8 | 17 ± 4.6 | 2.8 ± 0.5 | 70 ± 13 |
| A265C | 5.73 | 1.38 ± 0.28 | 8.24 ± 0.58 | 48.2 ± 3.8 | 11 ± 6.5 | 3.1 ± 0.2 | 64 ± 2 |
COS-7 cells were transiently transfected with the indicated M3′(3C)-Xa-derived Cys-substituted mutant M3R (rat) constructs. Radioligand binding and PI assays were performed and analyzed as described in Materials and Methods. Basal inositol monophosphate levels for the M3′ (3C)-Xa construct were 11,043 ± 1891 dpm (=100%). Data are shown as means ± se of 2–4 independent experiments.