Reactions of polypeptide ions with electrons in the gas phase

Abstract

Reactions of electrons in the energy range below 70 eV with polypeptide cations and anions are reviewed, as well as their applications for the structural analysis of polypeptides. At very low energies (</= 0.1 eV), the major outcome is electron-capture dissociation (ECD) of S-S and backbone N-C(alpha) bonds, leading to c' and z. fragments. ECD is useful in sequencing and characterization of post-translational modifications (PTMs), because c', z. fragmentation is abundant and the fragments usually retain labile groups. Electron capture at higher energies (3-13 eV) induces secondary fragmentation in radical z. fragments; this hot ECD (HECD) allows one to distinguish between the isomeric leucine and isoleucine residues. If a hot electron is not captured, then the induced electronic excitation converts internally into vibrational energy, resulting in fragmentation of the C(O)bond;N backbone bond (so-called EIEIO process). Above 9-10 eV, further ionization of n-charged cations occurs. If the formed (n + 1)+. cations capture electrons, then the C(alpha)bond;C backbone bond is usually broken. For anions that collide with approximately 20 eV electrons, the ejection of an electron leads to the creation of a radical positive charge (hole) that recombines internally with a negative charge. Such recombination leads to various backbone bond cleavages. This electron-detachment dissociation (EDD) is analogous to ECD for negative ions.

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

“…2). 24,36 Typically, positive ions are produced in an external electrospray ionization (ESI) source of an FTICR mass spectrometer and accumulated in an hexapole ion trap to increase the sensitivity and duty cycle of MS and MS/MS analysis. 37 -39 Subsequently, ion packages are transported by an ion transfer line to the ICR trap, located in a high magnetic field.…”

“…The electron capture dissociation (ECD) technique, discovered in 1998 by McLafferty's group, 24 became a unique MS/MS technique, providing structural information complementary to slow-heating fragmentation methods. The ECD technique is based on fragmentation of multiply charged peptide and protein cations due to interaction with electrons.…”

“…2 Early applications of ECD reactions for peptide and protein MS demonstrated its unique abilities and great potential to solve important problems in peptide and protein structural elucidation, not achieved by slow-heating fragmentation methods. 24 However, despite first efforts to optimize the performance of ECD/MS, the fragmentation process was not well controlled and sequence information of only a limited number of peptides was obtained using commercially available instruments. 25 One of the main drawbacks of the technique was the long period of electron irradiation required (up to 30 s and longer).…”

“…1), the electron flux, beam dimensions and electron energy control were optimized by the implementation of electron-injection systems based on indirectly heated dispenser cathodes. 24,26 As a result, improvements in the ECD rate and performance were achieved, enabling the combination of ECD/FTICR-MS with on-line liquid separation techniques, such as LC and CE, for structural analysis of peptides in peptide mixtures. 27 -29 From earlier experiments with directly heated filaments it was known that increased fragmentation efficiency of large proteins (up to 45 kDa) can be obtained using vibrational activation prior to electron irradiation.…”

“…18 The second employed an on-axis hollow electron beam for high-rate ECD, while the IR laser beam was kept on-axis. 19 The application area of high-rate ECD/FTICR-MS is growing rapidly, currently covering the de novo sequencing and sequence verification of peptides and proteins, 24 characterization of post-translational modifications in peptides and proteins, 24,31 revealing the secondary and tertiary structures of proteins, 32 studies of protein folding/unfolding dynamics, 33,34 and analysis of complex biological mixtures by on-line and off-line combination of ECD and LC(CE)/FTICR-MS. 27 -29,35 The present paper presents an overview of selected applications of high-rate ECD/FTICR-MS in peptide and protein characterization, preceded by a short description of the basic features of ECD and current experimental configurations.…”