Open AccessInternal Fragments Generated from Different Top-Down Mass Spectrometry Fragmentation Methods Extend Protein Sequence Coverage
Author(s) -
Muhammad A. Zenaidee,
Benqian Wei,
Carter Lantz,
Hoi Ting Wu,
Tyler R. Lambeth,
Jolene K. Diedrich,
Rachel R. Ogorzalek Loo,
Ryan R. Julian,
Joseph A. Loo
Publication year - 2021
Publication title -
journal of the american society for mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.961
H-Index - 127
eISSN - 1879-1123
pISSN - 1044-0305
DOI - 10.1021/jasms.1c00113
Subject(s) - chemistry , mass spectrometry , protein sequencing , sequence (biology) , fragmentation (computing) , complete sequence , electron capture dissociation , sequence analysis , protein primary structure , myoglobin , peptide sequence , fragment (logic) , tandem mass spectrometry , biochemistry , dna , chromatography , algorithm , gene , biology , ecology , genome , computer science
Top-down mass spectrometry (TD-MS) of intact proteins results in fragment ions that can be correlated to the protein primary sequence. Fragments generated can either be terminal fragments that contain the N- or C-terminus or internal fragments that contain neither termini. Traditionally in TD-MS experiments, the generation of internal fragments has been avoided because of ambiguity in assigning these fragments. Here, we demonstrate that in TD-MS experiments internal fragments can be formed and assigned in collision-based, electron-based, and photon-based fragmentation methods and are rich with sequence information, allowing for a greater extent of the primary protein sequence to be explained. For the three test proteins cytochrome c , myoglobin, and carbonic anhydrase II, the inclusion of internal fragments in the analysis resulted in approximately 15-20% more sequence coverage, with no less than 85% sequence coverage obtained. Combining terminal fragment and internal fragment assignments results in near complete protein sequence coverage. Hence, by including both terminal and internal fragment assignments in TD-MS analysis, deep protein sequence analysis, allowing for the localization of modification sites more reliably, can be possible.