Dataset Information

AtT 20 endogenous peptides - Improving the identification rate of endogenous peptides using electron transfer dissociation and collision-induced dissociation

ABSTRACT: We performed LC-MSMS analysis using both CID and ETD for the identification of endogenous peptides. Endogenous peptides were extracted from mouse AtT 20 cells by acidified methanol method and all large molecules including proteins were removed by centrifugation. The supernatant containing endogenous peptides was freeze-dried. For LC-MSMS analysis, extracted peptides were resuspended and injected to Ultimate 3000 HPLC system and analysed on LTQ Orbitrap XL mass spectrometer. A 60 min gradient from 2% acetonitrile to 50% acetonitrile, both containing 0.1% formic acid was used to separate peptides on C18 column.The LTQ-Orbitrap mass spectrometer was operated in data-dependent mode, automatically switching between MS and MS/MS acquisition for the three most abundant peaks in a given MS spectrum. A chosen precursor ion was first fragmented by CID and ETD. Data processing: The raw data files were processed with Proteome Discoverer 1.3. The CID and ETD spectra were then written to Mascot generic files. OMSSA (version 2.1.9) was used and b- and y- ions were selected for CID data, and c-, y- and z- ions were used for ETD. The spectra were searched by setting the parent ion mass accuracy to +/- 0.02 Da. For fragment ions, the mass tolerance was set to +/- 0.4 Da. For the genome-wide peptide search, the mouse genomic sequence (NCBI build 37.61) was directly translated in its 6 reading frames, and used for spectral searching. No enzymatic cleavage was taken into account during the database searches. No variable PTMs were included.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Mus Musculus (mouse)

SUBMITTER: Eisuke Hayakawa

PROVIDER: PXD000186 | Pride | 2014-06-09

REPOSITORIES: Pride

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Publications

Improving the identification rate of endogenous peptides using electron transfer dissociation and collision-induced dissociation.

Hayakawa Eisuke E Menschaert Gerben G De Bock Pieter-Jan PJ Luyten Walter W Gevaert Kris K Baggerman Geert G Schoofs Liliane L

Journal of proteome research 20131003 12

Tandem mass spectrometry (MS/MS) combined with bioinformatics tools have enabled fast and systematic protein identification based on peptide-to-spectrum matches. However, it remains challenging to obtain accurate identification of endogenous peptides, such as neuropeptides, peptide hormones, peptide pheromones, venom peptides, and antimicrobial peptides. Since these peptides are processed at sites that are difficult to predict reliably, the search of their MS/MS spectra in sequence databases nee ...[more]

PMID: 24032530

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Dataset Information

AtT 20 endogenous peptides - Improving the identification rate of endogenous peptides using electron transfer dissociation and collision-induced dissociation

Publications

Improving the identification rate of endogenous peptides using electron transfer dissociation and collision-induced dissociation.

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