A chemical strategy for protease substrate profiling

Here, we introduce a simple method, termed CHOPS, for the discovery of protease substrates. CHOPS exploits a 2-pyridinecarboxaldehyde (2PCA)-biotin probe, which selectively biotinylates protein N-termini except those with proline in the second position. CHOPS can, in theory, discover substrates for any protease, but is particularly well-suited to discover canonical DPP substrates, as cleaved but not intact DPP substrates can be identified by gel electrophoresis or mass spectrometry. This work delineates a practical technology for identifying protease substrates, which we anticipate will complement available “N-terminomic” approaches.