Peptide and protein characterization by high‐rate electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry

Abstract

The analytical utility of the electron capture dissociation (ECD) technique, developed by McLafferty and co‐workers, has substantially improved peptide and protein characterization using Fourier transform ion cyclotron resonance mass spectrometry (FTICR‐MS). The limitations of the first ECD implementations on commercial instruments were eliminated by the employment of low‐energy electron‐injection systems based on indirectly heated dispenser cathodes. In particular, the ECD rate and reliability were greatly increased, enabling the combination of ECD/FTICR‐MS with on‐line liquid separation techniques. Further technique development allowed the combination of two rapid fragmentation techniques, high‐rate ECD and infrared multiphoton dissociation (IRMPD), in a single experimental configuration. Simultaneous and consecutive irradiations of trapped ions with electrons and photons extended the possibilities for ion activation/dissociation and led to improved peptide and protein characterization. The application of high‐rate ECD/FTICR‐MS has demonstrated its power and unique capabilities in top‐down sequencing of peptides and proteins, including characterization of post‐translational modifications, improved sequencing of peptides with multiple disulfide bridges and secondary fragmentation (w‐ion formation). Analysis of peptide mixtures has been accomplished using high‐rate ECD in bottom‐up mass spectrometry based on mixture separation by liquid chromatography and capillary electrophoresis. This paper summarizes the current impact of high‐rate ECD/FTICR‐MS for top‐down and bottom‐up mass spectrometry of peptides and proteins. Copyright © 2004 John Wiley & Sons, Ltd.