PXD043709-2

PXD043709 is an original dataset announced via ProteomeXchange.

Title	Chemical acetylation of ligands and two-step digestion protocol for reducing co-digestion in affinity purification-mass spectrometry
Description	We present an effective, fast and user-friendly method to reduce co-digestion of bead-bound ligands, such as antibodies or streptavidin, in affinity purification-mass spectrometry experiments. A short pre-incubation of beads with Sulfo-NHS-Acetate leads to chemical acetylation of lysine residues, making ligands insusceptible to Lys-C-mediated proteolysis. In contrast to similar approaches, our procedure offers the advantage of exclusively using non-toxic chemicals and employing mild chemical reaction conditions. After binding of bait proteins to Sulfo-NHS-Acetate treated beads, we employ a two-step digestion protocol with the sequential use of Lys-C protease for on-bead digestion followed by in-solution digestion of the released proteins with trypsin. The implementation of this protocol results in a strong reduction of contaminating ligand peptides, which allows significantly higher amounts of sample to be subjected to LC-MS analysis, improving sensitivity and quantitative accuracy.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_09:06:10.595.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Wolfgang Reiter
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606; scientific name: Saccharomyces cerevisiae (Baker's yeast); NCBI TaxID: 4932;
ModificationList	No PTMs are included in the dataset
Instrument	Q Exactive HF-X

Revision	Datetime	Status	ChangeLog Entry
0	2023-07-12 03:50:23	ID requested
1	2023-10-23 05:15:44	announced
⏵ 2	2023-11-14 09:06:16	announced	2023-11-14: Updated project metadata.

Bordons M, Zulueta MA, [Evaluation of the scientific activity through bibliometric indices]. Rev Esp Cardiol, 52(10):790-800(1999) [pubmed]

submitter keyword: chemical acetylation, immunopurification, streptavidin, ligand,affinity purification, Sulfo-NHS-Acetate, nanobody, GFP

Markus Hartl
contact affiliation	Mass Spectrometry Facility, Max Perutz Labs Vienna, University of Vienna, Vienna Biocenter, Dr.-Bohr-Gasse 7, 1030 Vienna
contact email	markus.hartl@univie.ac.at
lab head
Wolfgang Reiter
contact affiliation	Mass Spectrometry Facility, Max Perutz Labs Vienna, University of Vienna, Vienna Biocenter, Dr.-Bohr-Gasse 7, 1030 Vienna
contact email	wolfgang.l.reiter@univie.ac.at
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2023/10/PXD043709

PRIDE project URI

• PRIDE
  1. PXD043709
     1. Label: PRIDE project
     2. Name: Chemical acetylation of ligands and two-step digestion protocol for reducing co-digestion in affinity purification-mass spectrometry