High-Throughput Method for N-Terminal Sequencing of Proteins by MALDI Mass Spectrometry

A high-throughput method for sequencing of N termini of proteins by using postsource decay (PSD) of matrix-assisted laser desorption/ionization mass spectrometry has been developed. After a protein blotted on the PVDF membrane was successively reduced, S-alkylated, and guanidinated, its N-amino group was coupled to biotinylcysteic acid. The protein was then extracted from the membrane and digested with trypsin. The derivatized N-terminal fragment was then specifically isolated from the tryptic digest with avidin resins, and its de novo sequencing was successfully performed by PSD utilizing a sulfonic acid group introduced to the N terminus.