Universität zu Köln

Ihling, Christian H., Springorum, Patrizia, Iacobucci, Claudio ORCID: 0000-0001-9592-3606, Hage, Christoph, Goetze, Michael, Schaefer, Mathias ORCID: 0000-0002-5943-4335 and Sinz, Andrea ORCID: 0000-0003-1521-4899 (2020). The Isotope-Labeled, MS-Cleavable Cross-Linker Disuccinimidyl Dibutyric Urea for Improved Cross-Linking/Mass Spectrometry Studies. J. Am. Soc. Mass Spectrom., 31 (2). S. 183 - 196. WASHINGTON: AMER CHEMICAL SOC. ISSN 1879-1123

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Abstract

Previous studies have shown the benefits of the amine reactive, CID-MS/MS-cleavable cross-linker disuccinimidyl dibutyric urea (DSBU) for structural proteomics studies via cross-linking/MS (XL-MS). To further facilitate the automation of XL-MS experiments, we synthesized a deuterated (D-12) version of the DSBU cross-linker combining the advantages of MS-cleavable linkers and isotope labeling. The rationale of conducting XL-MS with a mixture of unlabeled and stable isotope-labeled DSBU is to obtain characteristic mass differences at the MS level indicating cross-linked species. These cross-linked species can then be selected for fragmentation by collisional activation. At the MS/MS level, the characteristic 26-u doublets arising from cleavage of the central urea group in DSBU confirm the amino acid sequences of cross-linked peptides as well as the exact cross-linking sites. D-12-labeled DSBU was tested on three systems with increasing complexity: (i) bovine serum albumin as purified protein, (ii) Escherichia coli ribosome as large, multimeric protein assembly, and (iii) Drosophila embryo extract as complete proteome. We demonstrate the benefits arising from the use of isotope-labeled DSBU for an automated assignment of cross-linked products. Combining isotope labeling and MS cleavability in one cross-linker resulted in higher cross-link identification numbers especially for highly complex protein mixtures.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Ihling, Christian H. UNSPECIFIED UNSPECIFIED UNSPECIFIED Springorum, Patrizia UNSPECIFIED UNSPECIFIED UNSPECIFIED Iacobucci, Claudio UNSPECIFIED orcid.org/0000-0001-9592-3606 UNSPECIFIED Hage, Christoph UNSPECIFIED UNSPECIFIED UNSPECIFIED Goetze, Michael UNSPECIFIED UNSPECIFIED UNSPECIFIED Schaefer, Mathias UNSPECIFIED orcid.org/0000-0002-5943-4335 UNSPECIFIED Sinz, Andrea UNSPECIFIED orcid.org/0000-0003-1521-4899 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-346889
DOI:	10.1021/jasms.9b00008
Journal or Publication Title:	J. Am. Soc. Mass Spectrom.
Volume:	31
Number:	2
Page Range:	S. 183 - 196
Date:	2020
Publisher:	AMER CHEMICAL SOC
Place of Publication:	WASHINGTON
ISSN:	1879-1123
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 RESONANCE MASS-SPECTROMETRY; PROTEIN-STRUCTURE ANALYSIS; RELIABLE IDENTIFICATION; STRUCTURAL PROTEOMICS; PRODUCTS; ENRICHMENT; TOOL Multiple languages Biochemical Research Methods; Chemistry, Analytical; Chemistry, Physical; Spectroscopy Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/34688