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• W2478485447 abstract "Cross-linking combined with mass spectrometry (MS) has evolved as an alternative strategy in structural biology for characterizing three-dimensional structures of protein assemblies and for mapping protein–protein interactions. Here, we describe an integrated workflow for an automated identification of cross-linked products that is based on the use of a tandem mass spectrometry (MS/MS) cleavable cross-linker (containing a 1,3-bis-(4-oxo-butyl)-urea group, BuUrBu) generating characteristic doublet patterns upon fragmentation. We evaluate different fragmentation methods available on an Orbitrap Fusion mass spectrometer for three proteins and an E. coli cell lysate. An updated version of the dedicated software tool MeroX was employed for a fully automated identification of cross-links. The strength of our cleavable cross-linker is that characteristic patterns of the cross-linker as well as backbone fragments of the connected peptides are already observed at the MS/MS level, eliminating the need for conducting MS3 or sequential CID (collision-induced dissociation)- and ETD (electron transfer dissociation)-MS/MS experiments. This makes our strategy applicable to a broad range of mass spectrometers with MS/MS capabilities. For purified proteins and protein complexes, our workflow using CID-MS/MS acquisition performs with high confidence, scoring cross-links at 0.5% false discovery rate (FDR). The cross-links provide structural insights into the intrinsically disordered tetrameric tumor suppressor protein p53. As a time-consuming manual inspection of cross-linking data is not required, our workflow will pave the way for making the cross-linking/MS approach a routine technique for structural proteomics studies." @default.
• W2478485447 created "2016-08-23" @default.
• W2478485447 creator A5053102600 @default.
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• W2478485447 date "2016-07-28" @default.
• W2478485447 modified "2023-10-17" @default.
• W2478485447 title "Integrated Workflow for Structural Proteomics Studies Based on Cross-Linking/Mass Spectrometry with an MS/MS Cleavable Cross-Linker" @default.
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• W2478485447 doi "https://doi.org/10.1021/acs.analchem.5b04853" @default.
• W2478485447 hasPubMedId "https://pubmed.ncbi.nlm.nih.gov/27428000" @default.
• W2478485447 hasPublicationYear "2016" @default.
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