Proteomics

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An optimised fragmentation regime for cross-linking/mass spectrometry


ABSTRACT: In this study we compared three different fragmentation techniques and two combined fragmentation schemes available on a novel tribrid mass spectrometer (Orbitrap Fusion Lumos, Themro Fisher Scientific) CID, HCD, ETD, ETD with supplemental CID (ETciD) and ETD with supplemental HCD (EThcD) on cross-linked peptides obtained by tryptic cleavage of SDA-cross-linked Human Serum Albumin (HSA). The three-dimensional structure of HSA has been resolved by X-ray crystallography [35] and is used as ground-truth to evaluate the identification results. Right choice of the fragmentation method allows increasing the number of identified linkage sites, increasing the sequence coverage of both linked peptides thereby reducing the second peptide problem, and increasing the precision of cross-link site calling.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Serum

SUBMITTER: Sven Giese  

LAB HEAD: Juri Rappsilber

PROVIDER: PXD003737 | Pride | 2016-08-02

REPOSITORIES: Pride

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Publications

Optimized Fragmentation Regime for Diazirine Photo-Cross-Linked Peptides.

Giese Sven H SH   Belsom Adam A   Rappsilber Juri J  

Analytical chemistry 20160804 16


Cross-linking/mass spectrometry has evolved into a robust technology that reveals structural insights into proteins and protein complexes. We leverage a new tribrid instrument with improved fragmentation capacities in a systematic comparison to identify which fragmentation method would be best for the identification of cross-linked peptides. Specifically, we explored three fragmentation methods and two combinations: collision-induced dissociation (CID), beam-type CID (HCD), electron-transfer dis  ...[more]

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