Effects of human GLP-1R 149 or 333 mutation on agonist signaling via pERK1/2
Data were analyzed using a three-parameter logistic equation as defined in eq. 1. pEC50 values represent the negative logarithm of the concentration of agonist that produces half the maximal response. Emax represents the maximal response normalized to that elicited by 10% fetal bovine serum. All mutants were analyzed with an operational model of agonism (eq. 2) to determine logτ values. All logτ values were then corrected to specific [125I]exendin(9–39) binding (logτc). Values are expressed as mean ± S.E.M. of four to six independent experiments, conducted in duplicate. Data were analyzed with one-way analysis of variance and Dunnett’s post-test.
| pERK1/2 | |||||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| GLP-1(1–36)NH2 | GLP-1(7–36)NH2 | Exendin-4 | Oxyntomodulin | Compound 2 | |||||||||||
| pEC50 | Emax | Logτc (τc) | pEC50 | Emax | Logτc (τc) | pEC50 | Emax | Logτc (τc) | pEC50 | Emax | Logτc (τc) | pEC50 | Emax | Logτc (τc) | |
| Wild-type | 7.3 ± 0.2 | 1.3 ± 0.1 | −0.72 ± 0.16 (0.19) | 8.0 ± 0.2 | 3.6 ± 0.2 | −0.10 ± 0.15 (0.79) | 8.1 ± 0.2 | 4.2 ± 0.3 | 0.02 ± 0.16 (1.05) | 7.6 ± 0.1 | 5.0 ± 0.2 | 0.21 ± 0.16 (1.62) | 5.9 ± 0.2 | 0.45 ± 0.1 | −1.21 ± 0.19 (0.06) |
| M149a | ND | 0.6 ± 0.1 | NA | 7.8 ± 0.3 | 4.1 ± 0.5 | NA | 8.0 ± 0.2 | 3.4 ± 0.3* | NA | 7.1 ± 0.2 | 3.7 ± 0.3* | NA | 5.9 ± 0.3 | 2.0 ± 0.4 | NA |
| A149 | ND | ND | ND | 6.9 ± 0.4 | 1.7 ± 0.4* | −0.58 ± 0.41 (0.26) | 7.6 ± 0.4 | 3.0 ± 0.5 | −0.30 ± 0.32 (0.50) | 7.0 ± 0.2 | 2.6 ± 0.2* | −0.38 ± 0.32 (0.42) | 4.7 ± 0.7 | 0.99 ± 0.7 | −0.95 ± 0.38 (0.11) |
| C149 | 6.4 ± 0.5 | 1.6 ± 0.6 | −0.90 ± 0.43 (0.13) | 7.3 ± 0.4 | 2.0 ± 0.4* | −0.82 ± 0.31 (0.15) | 7.7 ± 0.3 | 4.0 ± 0.4 | −0.36 ± 0.31 (0.44) | 6.9 ± 0.2 | 3.3 ± 0.4* | −0.48 ± 0.31 (0.33) | 5.6 ± 0.4 | 0.33 ± 0.1 | −1.67 ± 0.38 (0.02) |
| F149 | ND | ND | ND | ND | ND | ND | 7.7 ± 0.2 | 1.3 ± 0.1* | −0.71 ± 0.32 (0.19) | 7.1 ± 0.2 | 1.3 ± 0.1* | −0.64 ± 0.28 (0.23) | 4.8 ± 0.4 | 0.87 ± 0.4 | −0.78 ± 0.45 (0.17) |
| I149 | ND | ND | ND | ND | ND | ND | 7.6 ± 0.3 | 1.1 ± 0.2* | −1.02 ± 0.39 (0.10) | 7.3 ± 0.2 | 1.3 ± 0.1* | −0.91 ± 0.26 (0.12) | 5.2 ± 0.4 | 0.71 ± 0.2 | −1.21 ± 0.26 (0.06) |
| S149 | 6.8 ± 0.4 | 1.4 ± 0.3 | −0.82 ± 0.26 (0.15) | 7.5 ± 0.2 | 3.3 ± 0.4 | −0.27 ± 0.25 (0.54) | 7.9 ± 0.2 | 3.7 ± 0.3 | −0.19 ± 0.26 (0.65) | 7.3 ± 0.2 | 4.9 ± 0.5 | 0.09 ± 0.25 (1.23) | 5.2 ± 0.4 | 0.79 ± 0.2 | −1.04 ± 0.29 (0.09) |
| V149 | ND | ND | ND | 7.5 ± 0.6 | 1.1 ± 0.3* | −0.97 ± 0.34 (0.11) | 7.2 ± 0.8 | 0.8 ± 0.4* | −1.12 ± 0.95 (0.08) | 7.1 ± 0.3 | 1.5 ± 0.3* | −0.77 ± 0.31 (0.17) | 5.5 ± 0.6 | 0.29 ± 0.1 | −1.50 ± 0.42 (0.03) |
| Y149 | ND | ND | ND | ND | ND | ND | 6.7 ± 0.6 | 1.0 ± 0.3* | −0.85 ± 0.67 (0.14) | 7.0 ± 0.5 | 0.6 ± 0.2* | −1.02 ± 0.36 (0.10)* | 4.8 ± 0.8 | 0.49 ± 0.4 | −1.15 ± 0.41 (0.07) |
| C333a | 7.7 ± 0.2 | 1.7 ± 0.2 | NA | 8.7 ± 0.2 | 5.9 ± 0.4 | NA | 9.1 ± 0.3 | 6.1 ± 0.5 | NA | 7.7 ± 0.2 | 7.0 ± 0.5 | NA | 6.0 ± 0.2 | 1.4 ± 0.2 | NA |
| A333 | 7.0 ± 0.2 | 1.7 ± 0.2 | −0.66 ± 0.17 (0.22) | 7.8 ± 0.2 | 6.1 ± 0.6* | 0.40 ± 0.18 (2.51) | 8.0 ± 0.3 | 6.7 ± 0.8* | 0.60 ± 0.21 (3.98) | 7.5 ± 0.2 | 7.5 ± 0.7* | 1.07 ± 0.36 (11.75) | 6.2 ± 0.3 | 0.70 ± 0.1 | −1.10 ± 0.17 (0.08) |
| V333 | 7.0 ± 0.2 | 1.4 ± 0.2 | −0.70 ± 0.28 (0.20) | 7.8 ± 0.1 | 4.1 ± 0.2 | 0.02 ± 0.27 (1.05) | 8.0 ± 0.1 | 4.7 ± 0.3 | 0.13 ± 0.27 (1.35) | 7.6 ± 0.2 | 6.3 ± 0.5 | 0.57 ± 0.28 (3.72) | 5.9 ± 0.4 | 1.11 ± 0.2 | −0.79 ± 0.27 (0.16) |
NA, data not experimentally determined in Koole et al., 2011; ND, data unable to be experimentally defined or with incomplete curves.
↵a Data obtained from Koole et al., 2011. Reported Emax values are normalized to 10% fetal bovine serum.
↵* Statistically significant at P < 0.05, one-way analysis of variance and Dunnett’s post-test in comparison with wild-type control.