Fluorogenic Peptide Substrates for Serine and Threonine Phosphatases

A new fluorescent assay for Ser/Thr protein phosphatases has been developed. Hydrolysis of a phosphoSer residue liberates the Ser hydroxyl group, which induces a cyclization reaction on the N-terminal carbamate and releases a fluorescent reporter. Sequence selectivity is observed using several peptide substrates against alkaline phosphatase (ALP), bacteriophage λ protein phosphatase (λ-PPase), and vaccinia H1 related phosphatase (VHR). These studies suggest that the assay could be a useful tool for profiling the substrate specificities of medicinally important phosphatases.