Universität zu Köln

Ihling, Christian, Falvo, Francesco, Kratochvil, Isabel, Sinz, Andrea and Schaefer, Mathias ORCID: 0000-0002-5943-4335 (2015). Dissociation behavior of a bifunctional tempo-active ester reagent for peptide structure analysis by free radical initiated peptide sequencing (FRIPS) mass spectrometry. J. Mass Spectrom., 50 (2). S. 396 - 407. HOBOKEN: WILEY. ISSN 1096-9888

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Abstract

We have synthesized a homobifunctional active ester cross-linking reagent containing a TEMPO (2,2,6,6-tetramethylpiperidine-1-oxy) moiety connected to a benzyl group (Bz), termed TEMPO-Bz-linker. The aim for designing this novel cross-linker was to facilitate MS analysis of cross-linked products by free radical initiated peptide sequencing (FRIPS). The TEMPO-Bz-linker was reacted with all 20 proteinogenic amino acids as well as with model peptides to gain detailed insights into its fragmentation mechanism upon collision activation. The final goal of this proof-of-principle study was to evaluate the potential of the TEMPO-Bz-linker for chemical cross-linking studies to derive 3D-structure information of proteins. Our studies were motivated by the well documented instability of the central NO?C bond of TEMPO-Bz reagents upon collision activation. The fragmentation of this specific bond was investigated in respect to charge states and amino acid composition of a large set of precursor ions resulting in the identification of two distinct fragmentation pathways. Molecular ions with highly basic residues are able to keep the charge carriers located, i.e. protons or sodium cations, and consequently decompose via a homolytic cleavage of the NO?C bond of the TEMPO-Bz-linker. This leads to the formation of complementary open-shell peptide radical cations, while precursor ions that are protonated at the TEMPO-Bz-linker itself exhibit a charge-driven formation of even-electron product ions upon collision activation. MS3 product ion experiments provided amino acid sequence information and allowed determining the cross-linking site. Our study fully characterizes the CID behavior of the TEMPO-Bz-linker and demonstrates its potential, but also its limitations for chemical cross-linking applications utilizing the special features of open-shell peptide ions on the basis of selective tandem MS analysis. Copyright (c) 2015 John Wiley & Sons, Ltd.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Ihling, Christian UNSPECIFIED UNSPECIFIED UNSPECIFIED Falvo, Francesco UNSPECIFIED UNSPECIFIED UNSPECIFIED Kratochvil, Isabel UNSPECIFIED UNSPECIFIED UNSPECIFIED Sinz, Andrea UNSPECIFIED UNSPECIFIED UNSPECIFIED Schaefer, Mathias UNSPECIFIED orcid.org/0000-0002-5943-4335 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-414018
DOI:	10.1002/jms.3543
Journal or Publication Title:	J. Mass Spectrom.
Volume:	50
Number:	2
Page Range:	S. 396 - 407
Date:	2015
Publisher:	WILEY
Place of Publication:	HOBOKEN
ISSN:	1096-9888
Language:	English
Faculty:	Faculty of Mathematics and Natural Sciences
Divisions:	Faculty of Mathematics and Natural Sciences > Department of Chemistry > Institute of Organic Chemistry
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language CHEMICAL CROSS-LINKING; PROTEIN-STRUCTURE ANALYSIS; GAS-PHASE; DIRECTED DISSOCIATION; FRAGMENTATION; LINKER; CLEAVAGE; IDENTIFICATION; ARCHITECTURE; CHAPERONIN Multiple languages Biochemical Research Methods; Chemistry, Analytical; Spectroscopy Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/41401