Exploring GPCR‐arrestin interfaces with genetically encoded crosslinkers. (14th September 2020)

• Record Type: Journal Article
• Authors: Böttke, Thore; Ernicke, Stefan; Serfling, Robert; Ihling, Christian; Burda, Edyta; Gurevich, Vsevolod V; Sinz, Andrea; Coin, Irene
• Is Part Of: EMBO reports. Volume 21:Number 11(2020)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2020-09-14
• Subjects: G protein‐coupled receptor -- genetically encoded crosslinkers -- GPCR–arrestin complexes -- live cells -- β‐arrestins; Molecular biology -- Periodicals; Molecular Biology -- Periodicals; Molecular biology; Periodicals; 572.8
• DOI: 10.15252/embr.202050437
• Languages: English
• ISSNs: 1469-221X

Abstract

Abstract: β‐arrestins (βarr1 and βarr2) are ubiquitous regulators of G protein‐coupled receptor (GPCR) signaling. Available data suggest that β‐arrestins dock to different receptors in different ways. However, the structural characterization of GPCR‐arrestin complexes is challenging and alternative approaches to study GPCR‐arrestin complexes are needed. Here, starting from the finger loop as a major site for the interaction of arrestins with GPCRs, we genetically incorporate non‐canonical amino acids for photo‐ and chemical crosslinking into βarr1 and βarr2 and explore binding topologies to GPCRs forming either stable or transient complexes with arrestins: the vasopressin receptor 2 (rhodopsin‐like), the corticotropin‐releasing factor receptor 1, and the parathyroid hormone receptor 1 (both secretin‐like). We show that each receptor leaves a unique footprint on arrestins, whereas the two β‐arrestins yield quite similar crosslinking patterns. Furthermore, we show that the method allows defining the orientation of arrestin with respect to the GPCR. Finally, we provide direct evidence for the formation of arrestin oligomers in the cell. Synopsis: Chemical and photo‐crosslinkers genetically incorporated into β‐arrestins allow the mapping of footprints of different receptors on the arrestin surface in living cells, and to identify intermolecular pairs of proximal amino acids in GPCR‐arrestin complexes. Genetically encoded crosslinkers can capture arrestin‐GPCR interactions with high spatial precision. Each G protein‐coupled receptor leaves a unique footprint on arrestins. Arrestins βarr1 and βarr2 may bind to a GPCR in a similar way. Abstract : Chemical and photo‐crosslinkers genetically incorporated into β‐arrestins allow the mapping of footprints of different receptors on the arrestin surface in living cells, and to identify intermolecular pairs of proximal amino acids in GPCR‐arrestin complexes.