Site-directed labeling of β-arrestin with monobromobimane for measuring their interaction with G protein-coupled receptors.

Abstract:

:β-arrestins (βarrs) are multifunctional proteins that interact with activated and phosphorylated G protein-coupled receptors (GPCRs) to regulate their signaling and trafficking. Understanding the intricate details of GPCR-βarr interaction continues to be a key research area in the field of GPCR biology. Bimane fluorescence spectroscopy has been one of the key approaches among a broad range of methods employed to study GPCR-βarr interaction using purified and reconstituted system. Here, we present a step-by-step protocol for labeling βarrs with monobromobimane (mBBr) in a site-directed fashion for measuring their interaction with GPCRs and the resulting conformational changes. This simple protocol can be directly applied to other protein-protein interaction modules as well for measuring interactions and conformational changes in reconstituted systems in vitro.

journal_name

Methods Enzymol

journal_title

Methods in enzymology

authors

Srivastava A,Baidya M,Dwivedi-Agnihotri H,Shukla AK

doi

10.1016/bs.mie.2019.11.009

subject

Has Abstract

pub_date

2020-01-01 00:00:00

pages

271-280

eissn

0076-6879

issn

1557-7988

pii

S0076-6879(19)30466-5

journal_volume

633

pub_type

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