Table 2.
Rates of reaction of MTSEA (M−1 s−1) with cysteine-substitution mutants of the CB2 receptor stably expressed in HEK293 cells.
| Receptor | kMTSEA (M−1 s−1) | kMUT/kWT |
|---|---|---|
| CB2* | 1.22 ± 0.03 | 1.00 |
| C2.59S | 0.23 ± 0.03 | 0.19 |
| (T6.36C) | 0.34 ± 0.09 | 0.28 |
| V6.51C* | 1.20 ± 0.02 | 0.98 |
| L6.52C* | 1.21 ± 0.02 | 0.99 |
| L6.54C* | 1.29 ± 0.02 | 1.06 |
| M6.55C* | 1.24 ± 0.03 | 1.02 |
| (H6.57C) | 0.28 ± 0.01 | 0.23 |
| L6.59C* | 1.24 ± 0.03 | 1.02 |
| (T6.61C) | 0.24 ± 0.01 | 0.20 |
| T6.62C* | 1.23 ± 0.01 | 1.01 |
Second-order rate constants were determined as described in materials and methods. Cells were treated with four concentrations of MTSEA (1, 3, 10 and 30 mM) and [3H]CP55940 binding performed. Rate constants of all accessible mutants were determined. Data represent the means±SEM of at least four independent experiments performed in duplicate. kMUT/kWT was obtained by dividing the k value obtained for each cysteine mutant by the k value of the CB2 wildtype receptor (not the C2.59S background which does not react). The asterisk (*) denotes a statistically significant difference (p < 0.05) compared to the C2.59S background mutant, by one-way ANOVA plus Newman-keuls post test. The brackets ( ) show selected “insensitive” mutants that were not significantly more inhibited than the background (C2.59S) mutant.