An on-line protein digestion device for rapid peptide mapping by electrospray mass spectrometry

A simple laboratory-constructed device has been developed for fast on-line protein digestion followed by peptide mapping by use of electrospray mass s pectrometry. Taking advantage of its nonsolubility properties at near-neutr al pH values, pepsin could be nonpermanently attached to the PEEK tube comm only employed as transfer capillary between the syringe and the electrospra y ion source. After optimization of experimental conditions such as pH, sol vent, and exposure time, efficient digestion of several model proteins of m olecular weights between 14 000 and 66 000, some having disulfide bridges, was successfully carried out. This technique provided reliable and reproduc ible sequence information by means of C-terminal-specific cleavages of arom atic and hydrophobic residues. As an application, protein identification co uld be achieved using a protein database search software.