A Novel Approach for Affinity-Based Screening of Target Specific Ligands:  Application of Photoreactive <scp>d</scp>-Glyceraldehyde-3-phosphate Dehydrogenase

Abstract

A novel application of the photoaffinity technique has been developed for the efficient discovery of small ligand and macromolecule interaction. The approach, photoaffinity capture, uses a photoreactive protein together with immobilized ligand for the rapid screening of competitive inhibitors. The set of photoreactive glyceraldehyde-3-phosphate dehydrogenase (photo-GAPDH) and immobilized dye ligand was prepared and examined as a model system. The photo-GAPDH was shown to efficiently capture the immobilized ligand. When nonimmobilized competitive ligands were included in the system, the capture was prevented in accordance with the affinity of the ligands. The present approach would provide an efficient tool for affinity-based screening of ligand libraries.