PXD004234-1

PXD004234 is an original dataset announced via ProteomeXchange.

Title	Quantification of Lysine Acetylation and Succinylation Stoichiometry in Proteins Using Mass Spectrometric Data-Independent Acquisitions (SWATH)
Description	Posttranslational modification of lysine residues by Nepsilon-acylation is an important regulator of protein function. Many large-scale protein acylation studies have assessed relative changes of lysine acylation sites using mass spectrometry-based proteomics. While relative acylation fold-changes are important, this does not reveal site occupancy, or stoichiometry, of individual modification sites, which is critical to understand functional consequences. Recently, methods for determining lysine acetylation stoichiometry have been proposed based on ratiometric analysis of endogenous levels to those introduced after quantitative per-acetylation of proteins using stable isotope-labeled acetic anhydride. However, in our hands we find that these methods can over-estimate acetylation stoichiometries due to signal interferences when endogenous levels of acylation are very low, which is especially problematic when using MS1 scans for quantification. In this study, we sought to improve the accuracy of determining acylation stoichiometry from data-independent acquisitions (DIA). Specifically, we use SWATH acquisitions to comprehensively collect both precursor and fragment ion intensity data. The use of fragment ions for stoichiometry quantification not only reduces interferences but also allows for determination of site-level stoichiometry from peptides with multiple lysine residues. We also demonstrate the novel extension of this method to measurements of succinylation stoichiometry by using deuterium labeled succinic anhydride. Proof of principle SWATH acquisition studies were first performed using BSA for both acetylation and succinylation occupancy measurements, followed by the analysis of more complex samples of E. coli cell lysates. While overall site occupancy was low (<1 percent), some proteins contained lysines with relatively high acetylation occupancy.
HostingRepository	MassIVE
AnnounceDate	2016-08-19
AnnouncementXML	Submission_2016-08-19_17:28:59.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Birgit Schilling
SpeciesList	scientific name: Bos taurus; common name: cattle; NCBI TaxID: 9913; scientific name: Escherichia coli; NCBI TaxID: 562;
ModificationList	Acetyl; Succinyl
Instrument	TripleTOF 6600; TripleTOF 5600

Revision	Datetime	Status	ChangeLog Entry
0	2016-05-26 20:04:13	ID requested
⏵ 1	2016-08-19 17:28:59	announced

79) J.G. Meyer, A.K. D?Souza, D.J. Sorensen, M.J. Rardin, A.J. Wolfe, B.W. Gibson, B. Schilling. Quantification of Lysine Acetylation and Succinylation Stoichiometry in Proteins Using Mass Spectrometric Data-Independent Acquisitions (SWATH). J. Amer. Soc. of Mass Spectrom. 2016, in press.

submitter keyword: Data-Independent Acquisitions , SWATH, Stoichiometry, chemical peracylation, stable isotope labeling, Skyline

Brad Gibson Birgit Schilling
lab head
Birgit Schilling
contact affiliation	Buck Institute
contact email	bschilling@buckinstitute.org
dataset submitter

MassIVE dataset URI

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