Label-Free Target Identification and Confirmation Using Thermal Stability Shift Assays

Abstract

Target identification presents one of the biggest challenges to chemical genomic approaches. In recent years, several methods have been applied for target identification and validation in plant cells. Here, we describe a label-free method based on the thermodynamic stabilization of a protein by interaction with a small-molecule ligand. With increasing temperature, proteins undergo thermal denaturation resulting in irreversible aggregation and precipitation. The binding of a small molecule to its target can enhance protein stability resulting in an increased temperature of aggregation (Tagg). This distinct increase in the temperature of aggregation known as a thermal shift can identify a compound–target protein interaction in high-throughput assays or, validate a predicted interaction.

Keywords

Small-molecule inhibitors; Target protein; Label-Free Target Identification; Thermal Stability Shift Assay

Published in

Methods in Molecular Biology
2021, volume: 2213, number: 2213, pages: 163-173
Title: Plant Chemical Genomics : Methods and Protocols
Publisher: Springer

SLU Authors

• Hicks, Glenn

  • Department of Molecular Sciences, Swedish University of Agricultural Sciences
    
  • University of California, Riverside (UCR)

UKÄ Subject classification

Biochemistry and Molecular Biology

Publication identifier

• DOI: https://doi.org/10.1007/978-1-0716-0954-5_14
• ISBN: 978-1-0716-0953-8
• eISBN: 978-1-0716-0954-5

Permanent link to this page (URI)

https://res.slu.se/id/publ/129795