Evaluation and optimization of electron capture dissociation efficiency in fourier transform ion cyclotron resonance mass spectrometry.
Electron capture dissociation (ECD) efficiency has typically been lower than for other dissociation techniques. Here we characterize experimental factors that limit ECD and seek to improve its efficiency. Efficiency of precursor to product ion conversion was measured for a range of peptide (approximately 15% efficiency) and protein (approximately 33% efficiency) ions of differing sizes and charge states. Conversion of precursor ions to products depends on electron irradiation period and maximizes at approximately 5-30 ms. The optimal irradiation period scales inversely with charge state. We demonstrate that reflection of electrons through the ICR cell is more efficient and robust than a single pass, because electrons can cool to the optimal energy for capture, which allows for a wide range of initial electron energy. Further, efficient ECD with reflected electrons requires only a short (approximately 500 micros) irradiation period followed by an appropriate delay for cooling and interaction. Reflection of the electron beam results in electrons trapped in or near the ICR cell and thus requires a brief (approximately 50 micros) purge for successful mass spectral acquisition. Further electron irradiation of refractory precursor ions did not result in further dissociation. Possibly the ion cloud and electron beam are misaligned radially, or the electron beam diameter may be smaller than that of the ion cloud such that remaining precursor ions do not overlap with the electron beam. Several ion manipulation techniques and use of a large, movable dispenser cathode reduce the possibility that misalignment of the ion and electron beams limits ECD efficiency.
McFarland
Melinda A
MA
Ion Cyclotron Resonance Program, National High Magnetic Field Laboratory, Florida State University, Tallahassee, Florida 32310-4005, USA.
Chalmers
Michael J
MJ
Quinn
John P
JP
Hendrickson
Christopher L
CL
Marshall
Alan G
AG
D017357:Cyclotrons; D018698:Glutamic Acid / Q000737:chemistry; D008555:Melitten / Q000737:chemistry; D010455:Peptides / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*; D013373:Substance P / Q000737:chemistry; D025801:Ubiquitin / Q000737:chemistry
D010455:Peptides; D025801:Ubiquitin; D008555:Melitten; D013373:Substance P; D018698:Glutamic Acid