Protocol for the comprehensive biochemical and cellular profiling of small-molecule degraders using CoraFluor TR-FRET technology.
STAR Protoc
; 5(2): 103129, 2024 Jun 21.
Article
em En
| MEDLINE
| ID: mdl-38857155
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.1.
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Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Transferência Ressonante de Energia de Fluorescência
Limite:
Humans
Idioma:
En
Revista:
STAR Protoc
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Estados Unidos