Pharmacological Characterization of D2R Mutants using Go and CAMYEL BRET Assays.
EC50 and IC50 values were derived as described in Figs. 9 and 10 via nonlinear regression of individual experiments normalized to the dopamine-stimulated wild-type (WT) D2R control. For each D2R mutant, the EC50 or IC50 value was divided by the wild-type EC50 or IC50 value to derive the corresponding wild-type ratio. Data are expressed as geometric means [95% confidence interval].
| WT | L81A | V91A | E95A | T165A | A177S | Y192A | V196A | I397A | ||
|---|---|---|---|---|---|---|---|---|---|---|
| Dopamine | Go EC50 | 9.9 nM [6.3–15] | 5.7 nM [1.5–14] | 7.7 nM [4.3–13] | 11 nM [6.8–17] | 10 nM [7.2–14] | 6.5 nM [4.5–9.3] | 5.9 nM [3.8–8.8] | 8.3 nM [5.8–12] | 7 nM [3.4–13] |
| WT Ratio | 1 | 0.6 | 0.8 | 1.1 | 1 | 0.7 | 0.6 | 0.8 | 0.7 | |
| cAMP EC50 | 12 nM [10–16] | 5.8 nM [4.8–7] | 9 nM [6–13] | 13 nM [9.7–17] | 12.7 nM [9.7–17] | 8.7 nM [6.3–12] | 5.1 nM [3.6–7.2] | 10 nM [7.5–14] | 3.3 nM [2–4.8] | |
| WT Ratio | 1 | 0.5 | 0.7 | 1 | 1 | 0.7 | 0.4 | 0.8 | 0.3 | |
| Sulpiride | Go IC50 | 13 nM [10–20] | 34 nM [24–46] | 38 nM [29–50] | 21 nM [29–50] | 9.8 nM [6.6–15] | 12 nM [8–17] | 37 nM [25–55] | 25 nM [19–33] | 51 nM [39–67] |
| WT Ratio | 1 | 2.6 | 2.9 | 1.6 | 0.8 | 0.9 | 2.9 | 2.1 | 3.9 | |
| cAMP IC50 | 17 nM [12–25] | 26 nM [20–35] | 63 nM [42–92] | 40 nM [24–65] | 18 nM [12–26] | 18 nM [13–23] | 43 nM [31–61] | 25 nM [17–38] | 78 nM [59–104] | |
| WT Ratio | 1 | 1.5 | 3.7 | 2.4 | 1 | 1 | 2.5 | 1.5 | 4.6 | |
| ONC201 | Go IC50 | 21 μM [12–40] | 45 μM [29–71] | >100 μM | >100 μM | 32 μM [15–67] | 31 μM [17–60] | 40 μM [25–66] | 50 μM [30–87] | >100 μM |
| WT Ratio | 1 | 2 | >5 | >5 | 1.5 | 1.5 | 1.9 | 2.4 | >5 | |
| cAMP IC50 | 23 μM [16–33] | 46 μM [20–122] | >100 μM | 55 μM [28–126] | 12 μM [9–16] | 29 μM [14–63] | >100 μM | 45 μM [23–97] | >100 μM | |
| WT Ratio | 1 | 2 | >5 | 2.4 | 0.5 | 1.3 | >5 | 2 | >5 |
N = 3–5 experiments run in triplicate.