Synthesis of Diazirinyl Photoprobe Carrying a Novel Cleavable Biotin

Photoaffinity labeling is a powerful tool for the identification of receptor proteins and their binding sites. The major drawback of this method, however, is the reliability of the labeling and the purity of labeled peptides after purification from a large number of unlabeled fragments. The recent development of diazirine-based biotinylated probes provides an efficient solution to this problem. In these probes, the diazirine photophore contributes to the formation of a stable cross-link that greatly improves the efficiency. The application of an N-acetylglucosamine photoprobe carrying biotinylated diazirine provided the first information regarding acceptor-site peptides of b-1,4-galactosyltransferase. We have also developed a novel method for the one-step introduction of a biotinylated diazirine photophore into unprotected carbohydrate ligands. Since biotin–avidin binding is essentially irreversible (Kd = 10 15 m), several approaches have been investigated to achieve efficient recovery of biotinylated products from an immobilized avidin matrix. Although the use of monomeric avidin contributes to the isolation of biotinylated products because of the lower affinity to biotin (Kd = 10 8 m), it requires a high