Impact of Charge State on Characterization of Large Middle-Down Sized Peptides by Tandem Mass Spectrometry

Fragmentation trends of large peptides were characterized by five activation methods, including HCD, ETD, EThcD, 213 nm UVPD, and 193 nm UVPD. Sequence coverages and scores were assessed based on charge site, peptide sequence, and peptide size. The effect of charge state and peptide size on sequence coverage was explored for a Glu-C digest of E. coli ribosomal proteins, and linear regression analysis of the collection of peptides indicated that HCD, ETD, and EThcD have a higher dependence charge state than 193 and 213 nm UV. Four model peptides, neuromedin, glucagon, galanin, and amyloid β, were characterized in greater detail based on charge site analysis and showed a charge state dependence on sequence coverage for collision and electron-based activation methods.