Thermodynamic Characterization of LC3/GABARAP:Ligand Interactions by Isothermal Titration Calorimetry

Overview

Journal Methods Mol Biol

Specialty Molecular Biology

Date 2024 Aug 8

PMID 39115670

Authors

Johannes Dopfer

Martin P Schwalm

Stefan Knapp

Vladimir V Rogov

Affiliations

Soon will be listed here.

Abstract

Isothermal titration calorimetry (ITC) is a widely used technique for the characterization of protein-protein and protein-ligand interactions. It provides information on the stoichiometry, affinity, and thermodynamic driving forces of interactions. This chapter exemplifies the use of ITC to investigate interactions between human autophagy modifiers (LC3/GABARAP proteins) and their interaction partners, the LIR motif-containing sequences. The purpose of this report is to present a detailed protocol for the production of LC3/GABARAP-interacting LIR peptides using E. coli expression systems. In addition, we outline the design of ITC experiments using the LC3/GABARAP:peptide interactions as an example. Comprehensive troubleshooting notes are provided to facilitate the adaptation of these protocols to different ligand-receptor systems. The methodology outlined for studying protein-ligand interactions will help to avoid common errors and misinterpretations of experimental results.

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