One-step peptide backbone dissociations in negative-ion free radical initiated peptide sequencing mass spectrometry

Jihye Lee, Hyeyeon Park, Hyuksu Kwon, Gyemin Kwon, Aeran Jeon, Hugh I. Kim, Bong June Sung, Bongjin Moon, Han Bin Oh

Research output: Contribution to journalArticle

26 Citations (Scopus)

Abstract

Peptide dissociation behavior in TEMPO (2,2,6,6-tetramethylpiperidine-1- oxyl)-based FRIPS (free radical initiated peptide sequencing) mass spectrometry was analyzed in both positive-and negative-ion modes for a number of peptides including angiotensin II, kinetensin, glycoprotein IIb fragment (296-306), des-Pro2-bradykinin, and ubiquitin tryptic fragment (43-48). In the positive mode, the ·Bz-C(O)-peptide radical species was produced exclusively at the initial collisional activation of o-TEMPO-Bz-C(O)-peptides, and two consecutive applications of collisional activation were needed to observe peptide backbone fragments. In contrast, in the negative-ion mode, a single application of collisional activation to o-TEMPO-Bz-C(O)-peptides produced extensive peptide backbone fragmentations as well as ·Bz-C(O)-peptide radical species. This result indicates that the duty cycle in the TEMPO-based FRIPS mass spectrometry can be reduced by one-half in the negative-ion mode. In addition, the fragment ions observed in the negative-ion experiments were mainly of the a-, c-, x-, and z-types, indicating that radical-driven tandem mass spectrometry was mainly responsible for the TEMPO-based FRIPS even with a single application of collisional activation. Furthermore, the survival fraction analysis of o-TEMPO-Bz-C(O)-peptides was made as a function of the applied normalized collision energy (NCE). This helped us to better understand the differences in FRIPS behavior between the positive-and negative-ion modes in terms of dissociation energetics. The duty-cycle improvement made in the present study provides a cornerstone for future research aiming to achieve a single-step FRIPS in the positive-ion mode.

Original languageEnglish
Pages (from-to)7044-7051
Number of pages8
JournalAnalytical chemistry
Volume85
Issue number15
DOIs
Publication statusPublished - 2013 Aug 6
Externally publishedYes

ASJC Scopus subject areas

  • Analytical Chemistry

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