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Membrane-mimetic thermal proteome profiling (MM-TPP) toward mapping membrane protein-ligand dynamic interactions
Rupinder Singh Jandu1, Ashim Bhattacharya1, Frank Antony1
1Department of Biochemistry and Molecular Biology, Life Sciences Institute, University of British Columbia, Vancouver, Canada.
Elife
|November 12, 2025
Summary
We developed membrane-mimetic thermal proteome profiling (MM-TPP) to study membrane protein interactions. This detergent-free method accurately maps small molecule interactions with integral membrane proteins, revealing on- and off-target effects.
Area of Science:
- Biochemistry
- Chemical Biology
- Proteomics
Background:
- Integral membrane proteins (IMPs) are crucial therapeutic targets, but studying their interactions is challenging due to limitations in current methods.
- Existing techniques often rely on detergents, which can disrupt protein structure and function, hindering unbiased analysis.
- Previous work introduced the Peptidisc membrane mimetic (MM) for stabilizing membrane proteins in a soluble state.
Purpose of the Study:
- To establish membrane-mimetic thermal proteome profiling (MM-TPP), a novel method for proteome-wide mapping of membrane protein-ligand interactions.
- To demonstrate the capacity of MM-TPP to detect specific on- and off-target effects of small molecules on IMPs.
- To provide a robust, detergent-free platform for exploring the druggable membrane proteome.
Main Methods:
- Integration of the Peptidisc membrane mimetic (MM) with thermal proteome profiling (TPP).
- Application of MM-TPP to a mouse liver membrane protein library to assess ligand interactions.
- Comparison of MM-TPP with detergent-based TPP (DB-TPP) using specific ligands like ATP and orthovanadate.
Main Results:
- MM-TPP successfully mapped ligand-induced thermal stability changes in ATP-binding cassette (ABC) transporters and G protein-coupled receptors (GPCRs).
- The method revealed specific interactions, such as ATP and its by-products with ABC transporters and GPCRs.
- MM-TPP demonstrated superior specificity compared to DB-TPP in detecting ATP-binding proteins and identified selective ligand binding (e.g., 2-methylthio-ADP with P2RY12 receptor).
Conclusions:
- MM-TPP is a powerful, detergent-free platform for studying membrane protein-ligand interactions.
- The method enables the discovery of both on- and off-target effects of small molecules on IMPs.
- MM-TPP offers valuable insights into the druggable membrane proteome and its dynamic stability.
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