MassIVE MSV000091064

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Improved Performance of Positive-ion mode Free Radical-Initiated Peptide Sequencing with p-TEMPO-Bz

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Description

Free radical-initiated peptide sequencing (FRIPS) is a tandem mass spectrometry (MS/MS) technique that generates sequence informative ions via collisionally-initiated radical chemistry. Collision activation (CA) homolytically cleaves an installed radical precursor, initiates radical formation, extensive hydrogen atom transfer, and peptide backbone dissociation. While FRIPS technique shows great promise, when applied to multiply charged derivatized peptide ions, a series of high abundance mass losses are observed which syphon ion abundance from radically generated sequence ions. This loss of ion abundance reduces the sequence coverage generated by FRIPS fragmentation. In this work, we hypothesized that these mass losses were instigated by the ortho-orientation of the radical precursor undergoing facile conversion into five- or six-membered intermediates and that the para-precursor would not undergo this chemistry. To test this assertion, we synthesized para-TEMPO-Bz, conjugated it to these peptides, and collisionally activated each. And indeed, we see the complete elimination of these undesired collisional processes and the significant increase in radical precursor ion abundance. The increase in ion abundance leads to a significant increase in the sequence coverage generated. These results demonstrate that p-TEMPO-Bz significantly improves the performance of positive-ion mode FRIPS and may be a suitable alternative to the currently utilized ortho-TEMPO-Bz-based FRIPS. [doi:10.25345/C5ZC7S43D] [dataset license: CC0 1.0 Universal (CC0 1.0)]

Keywords: Free radical initiated peptide sequencing

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Principal Investigators:
(in alphabetical order) 		Nicholas Borotto, University of Nevada, United States
Submitting User: nborotto
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Distinct protein accessions are counted across all files submitted in the "Statistical Analysis of Quantified Analytes" category having a "Protein" column in this dataset.

"N/A" means no results of this type were submitted.
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