Protocol for clickable photoaffinity labeling and quantitative chemical proteomics.

Area of Science:

• Chemical Biology
• Proteomics
• Molecular Biology

Background:

• Identifying specific protein targets of small molecules is crucial for understanding drug mechanisms.
• Traditional methods for target deconvolution can be complex and time-consuming in live cellular environments.

Purpose of the Study:

• To present a novel photoaffinity labeling probe strategy for efficient target deconvolution in live cells.
• To detail a protocol applicable to identifying targets of nuclear receptors like Liver X Receptor beta (LXRβ).

Main Methods:

• Synthesis of a chemical probe incorporating a photoreactive group.
• Application of UV irradiation to induce covalent cross-linking between the probe and adjacent amino acid residues.
• Utilization of click chemistry for enrichment of labeled proteins prior to proteomic analysis.

Main Results:

• Successful demonstration of the photoaffinity labeling probe strategy in live cells.
• Establishment of a protocol for identifying targets of LXRβ.
• The protocol shows potential for broad application in target deconvolution for various proteins.

Conclusions:

• The described photoaffinity labeling probe strategy offers a robust method for target deconvolution.
• This protocol facilitates the identification of direct protein interactions within a cellular context.
• The methodology is adaptable for discovering targets of diverse proteins beyond LXRβ.