Protocol for the comprehensive biochemical and cellular profiling of small-molecule degraders using CoraFluor TR-FRET technology.

STAR protocols
Authors
Keywords
Abstract

Comprehensive characterization of small-molecule degraders, including binary and ternary complex formation and degradation efficiency, is critical for bifunctional ligand development and understanding structure-activity relationships. Here, we present a protocol for the biochemical and cellular profiling of small-molecule degraders based on CoraFluor time-resolved fluorescence resonance energy transfer (TR-FRET) technology. We describe steps for labeling antibodies and proteins, tracer saturation binding, binary target engagement, ternary complex profiling, and off-rate determination. We then detail procedures for the quantification of endogenous and GFP fusion proteins in cell lysates. For complete details on the use and execution of this protocol, please refer to Ichikawa et al..

Year of Publication
2024
Journal
STAR protocols
Volume
5
Issue
2
Pages
103129
Date Published
06/2024
ISSN
2666-1667
DOI
10.1016/j.xpro.2024.103129
PubMed ID
38857155
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