Fig. 1: Model class A and B GPCRs, β2 adrenergic receptor (b2AR) and Vasopressin receptor 2 (V2R), induce distinct patterns of β-arrestin conformational patterns.

a Schematic representation of NanoBRET-based assay to measure β-arrestin2-NanoLuciferase (Nluc) recruitment to the GPCR-Halo-Tag in Control cells. For the b2AR measurements, the BRET pair was switched. b, c β-arrestin2 recruitment to b2AR (b) and V2R (c), as published in Drube et al.²⁶. Here, the Δ net BRET fold change was analyzed over time when cells were stimulated with 10 µM Isoprenaline (Iso) (b) or 3 µM [Arg8]-Vasopressin (AVP) (c). Timepoint 0 is defined as midpoint of the average time needed for ligand addition, considering all analyzed biological and technical replicates. All data are shown ±SEM of n = 3 independent experiments. d Schematic visualization of the utilized intramolecular NanoBRET-based β-arrestin2 sensors, as described in detail in Haider et al.²⁵. e, f Conformational change of β-arrestin2-FlAsH (F) 5-Nluc in Control cells, co-transfected with untagged b2AR (e)²⁶ or V2R (f). Data were analyzed over time as in (b, c) and are shown as Δ net BRET change in percent. g–j Fingerprint of β-arrestin2 conformational change sensors measured in Control cells in presence of untagged b2AR (g, i) or V2R (h, j). The Δ net BRET change at 10 µM Iso (g) or 3 µM AVP (h) are shown as bar graphs. All data are shown ±SEM of n = 3 independent experiments. Sensor conditions, which did not fulfill the set pharmacological parameters (Hill slope and EC₅₀ analysis as described in methods) were classified as non-responding conditions and assigned zero. These data were visualized onto the surface of the inactive β-arrestin2 crystal structure (PDB: 3P2D) by coloring the respective loop (-fragments) of the labeled FlAsH positions (i, j) ranging from blue to red. For each receptor, the Δ net BRET change was normalized to the maximally reacting sensor (red). Non-responding conditions are shown in gray. Source data are provided as a Source Data file.