An intrabody sensor to monitor conformational activation of β-arrestins

Hemlata Dwivedi-Agnihotri, Parishmita Sarma, S. Deeksha, Kouki Kawakami, Asuka Inoue, Arun K. Shukla

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

5 Citations (Scopus)


Agonist-induced interaction of β-arrestins with GPCRs is critically involved in downstream signaling and regulation. This interaction is associated with activation and major conformational changes in β-arrestins. Although there are some assays available to monitor the conformational changes in β-arrestins in cellular context, additional sensors to report β-arrestin activation, preferably with high-throughput capability, are likely to be useful considering the structural and functional diversity in GPCR-β-arrestin complexes. We have recently developed an intrabody-based sensor as an integrated approach to monitor GPCR-β-arrestin interaction and conformational change, and generated a luminescence-based reporter using NanoBiT complementation technology. This sensor is derived from a synthetic antibody fragment referred to as Fab30 that selectively recognizes activated and receptor-bound conformation of β-arrestin1. Here, we present a step-by-step protocol to employ this intrabody sensor to measure the interaction and conformational activation of β-arrestin1 upon agonist-stimulation of a prototypical GPCR, the complement C5a receptor (C5aR1). This protocol is potentially applicable to other GPCRs and may also be leveraged to deduce qualitative differences in β-arrestin1 conformations induced by different ligands and receptor mutants.

Original languageEnglish
Title of host publicationBiomolecular Interactions Part B
EditorsArun K. Shukla
PublisherAcademic Press Inc.
Number of pages12
ISBN (Print)9780128233535
Publication statusPublished - 2022 Jan

Publication series

NameMethods in Cell Biology
ISSN (Print)0091-679X


  • Antibody fragments
  • Biosensors
  • C5aR1
  • Cellular signaling
  • Complement C5a
  • GPCRs
  • Intrabody
  • Protein-protein interactions
  • β-Arrestins


Dive into the research topics of 'An intrabody sensor to monitor conformational activation of β-arrestins'. Together they form a unique fingerprint.

Cite this