Table 2 Potency, efficacy, and affinity of the chimeric MCP proteins at CCR2 in β-arrestin recruitment, ERK phosphorylation, and radioligand-binding assays.

125I–MCP-1 competition binding was measured in membrane preparations of c-Myc–FLAG–CCR2 Flp-In T-REx 293 cells. β-arr2 recruitment was assessed by BRET in Flp-In T-REx 293 cells transiently transfected with plasmids encoding CCR2-RLuc8 and β-arr2–YFP. ERK1/2 phosphorylation was measured 3 to 5 min after c-Myc–FLAG–CCR2 Flp-In T-REx 293 cells were stimulated with chemokine. pEC50 and pKi values are the negative log of EC50 and Ki values, respectively, in molar units. Emax values are relative to the positive control. Data are means ± SEM of three or four experiments, each performed in triplicate. *P < 0.05, **,^^P < 0.01, ***,^^^P < 0.001, compared to MCP-1 or MCP-3, respectively. Statistical analysis was by one-way ANOVA with Dunnett’s multiple-comparison text.

β-Arrestin recruitmentERK1/2 phosphorylation125I–MCP-1 binding
pEC50EmaxpEC50EmaxpKi
MCP-18.00 ± 0.130.100 ± 0.006^^^7.87 ± 0.3237.8 ± 3.410.67 ± 0.18^^
MCP1-3117.30 ± 0.210.067 ± 0.006*6.84 ± 0.1740.3 ± 3.29.90 ± 0.18
MCP1-1318.32 ± 0.130.11 2 ± 0.006^^^7.64 ± 0.2643.7 ± 3.710.53 ± 0.17^^
MCP1-1138.12 ± 0.130.111 ± 0.006^^^7.58 ± 0.4646.7 ± 6.810.68 ± 0.16^^
MCP1-1338.45 ± 0.270.103 ± 0.009^^7.86 ± 0.4236.4 ± 5.010.84 ± 0.16^^
MCP1-3337.82 ± 0.580.034 ± 0.009***6.95 ± 0.2924.0 ± 3.110.02 ± 0.17
MCP-37.63 ± 0.170.060 ± 0.004**7.21 ± 0.3134.3 ± 4.09.50 ± 0.19**
MCP3-1338.24 ± 0.110.0135 ± 0.005^^^8.12 ± 0.3325.9 ± 2.310.36 ± 0.14
MCP3-3137.17 ± 0.220.051 ± 0.006***7.43 ± 0.2733.1 ± 3.07.45 ± 0.23***,^^
MCP3-3317.65 ± 0.220.056 ± 0.006**7.66 ± 0.3621.7 ± 2.3*8.77 ± 0.33***
MCP3-3117.13 ± 0.390.050 ± 0.010***7.87 ± 0.1440.0 ± 1.67.32 ± 0.32***,^^
MCP3-1117.61 ± 0.200.134 ± 0.010^^^7.99 ± 0.3621.3 ± 2.4*9.80 ± 0.17