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An enhanced protein crosslink identification strategy using CID‐cleavable chemical crosslinkers and LC/MS n analysis
Author(s) -
Liu Fan,
Wu Cong,
Sweedler Jonathan V.,
Goshe Michael B.
Publication year - 2012
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.201100352
Subject(s) - chemistry , peptide , combinatorial chemistry , monomer , reagent , mass spectrometry , chromatography , biochemistry , organic chemistry , polymer
We describe a novel two‐step LC/MS n strategy to effectively and confidently identify numerous crosslinked peptides from complex mixtures. This method incorporates the use of our gas‐phase cleavable crosslinking reagent, disuccinimidyl‐succinamyl‐aspartyl‐proline (SuDP), and a new data‐processing algorithm CXLinkS (Cleavable Crosslink Selection), which enables unequivocal crosslink peptide selection and identification on the basis of mass measurement accuracy, high resolving power, and the unique fragmentation pattern of each crosslinked peptide. We demonstrate our approach with well‐characterized monomeric and multimeric protein systems with and without database searching restrictions where inter‐peptide crosslink identification is increased 8‐fold over our previously published data‐dependent LC/MS 3 method and discuss its applicability to other CID‐cleavable crosslinkers and more complex protein systems.