sepharose and carbene

Small molecule-immobilized affinity matrices are indispensable tools in chemical genomics to screen and purify protein targets for biologically active small molecules. Usually, prior to immobilization, small -molecules have to be derivatized at a position that does not significantly abrogate the intrinsic biological activity, or chemically synthesized to have an appropriate functional group for the immobilization chemistry. Here, we describe a photo-cross-linking technique to immobilize biologically active small molecules for protein target screening, without the need for chemical derivatization.

Topics: Chromatography, Affinity; Cross-Linking Reagents; Cyclosporine; Mass Spectrometry; Methane; Microspheres; Photochemical Processes; Proteins; Sepharose; Small Molecule Libraries

The introduction of a cleavable site in a photoactivatable linker, which is used to immobilize small molecules on an affinity matrix via a site-nonselective carbene addition/insertion reaction, makes it possible to verify the presence of the immobilized small molecule on the affinity matrix. It also permits the efficient detection of proteins covalently bound to the immobilized small molecule.

Topics: Binding Sites; Blotting, Western; Chromatography, Liquid; Cross-Linking Reagents; Disulfides; Enzymes, Immobilized; Ligands; Mass Spectrometry; Methane; Microspheres; Organophosphorus Compounds; Photoaffinity Labels; Photochemistry; Protein Binding; Protein Phosphatase 2; Sepharose