μMap Photoproximity Labeling Enables Small Molecule Binding Site Mapping.

Area of Science:

• Chemical Biology
• Drug Discovery
• Proteomics

Background:

• Ligand binding site characterization is vital in drug discovery, especially for phenotypic screening where targets are initially unknown.
• Existing methods like X-ray crystallography and photoaffinity labeling (PAL) have limitations, including technical challenges, poor signal, and complex data analysis.

Purpose of the Study:

• To establish a robust and general photocatalytic approach for mapping protein binding sites, identifying residues proximal to the ligand binding pocket.
• To demonstrate the utility of this method for analyzing various protein targets and drug candidates, including those without existing structural data.

Main Methods:

• Development of a catalytic photocatalytic method (μMap) for protein binding site mapping.
• Utilizing catalytic activation to label amino acids in proximity to the ligand binding site.
• Application of the method in vitro to six protein targets and in live cells.

Main Results:

• Successfully mapped binding sites of six diverse protein targets, including kinases and molecular glues.
• Investigated the binding site of the STAT3 inhibitor MM-206, a ligand lacking crystal structure data.
• Demonstrated successful mapping of drug binding sites within live cells.

Conclusions:

• μMap provides a powerful and generalizable method for identifying amino acids near ligand binding pockets.
• The technique generates valuable amino-acid- and peptide-level target engagement data.
• μMap enables binding site characterization in vitro and in live cells, advancing drug discovery efforts.