Efficient approach for profiling photoaffinity labeled peptides with a cleavable biotinyl photoprobe.

Based on the application of our recent biotinyl photoprobe with a cleavable N-acylsulfonamide, an efficient process has been developed for profiling photoaffinity labeled peptides among a large excess of unlabeled concomitants. N-acylsulfonamide group was found to be stable under the usual S-pyridylethylation condition of cysteine residues whereas the group was easily cleaved by N-alkylation with iodoacetic acid in acidic condition. The selective nature between two common protein alkylation reactions was evaluated with L-glutamate dehydrogenase (GDH) using an acidic amino acid photoprobe with biotinylated acylsulfonamide function.

Synthesis and evaluation of novel photoreactive alpha-amino acid analog carrying acidic and cleavable functions.

A novel photoreactive alpha-amino acid bearing an acidic residue and a cleavable diazirine was developed. To mimic common acidic alpha-amino acids, the residue was designed to be N-acylsulfonamide that possesses an acidic proton and is able to dissociate under the physiological conditions. The inhibition assay of its biotin-tagged derivative with glutamyl endopeptidase from Staphylococcus aureus V8 strain revealed a Ki(app) value of 162 microM, which is slightly higher than the K(m) value of a common substrate.