PXD041674

PXD041674 is an original dataset announced via ProteomeXchange.

Title	Determining DNA-protein binding affinities and specificities from crude lysates using a combined SILAC/TMT labeling strategy
Description	Gene expression is driven by the binding of transcription factors to regulatory elements in the genome, such as enhancers and promoters. A powerful technique to study DNA-protein interactions is affinity purification followed by mass spectrometry. Classic affinity purifications coupled to quantitative mass spectrometry provide information about binding specificity. Binding of transcription factors to regulatory elements in vivo, however, also depends on binding affinity, so the strength of an interaction. To obtain this information, we recently developed a technique called PAQMAN that uses a series of DNA affinity purifications to quantify apparent binding affinities proteome-wide. Here, we expand our PAQMAN workflow to obtain information about binding specificity and affinity in a single experiment. To this end, we combine quantitation at the MS1 level with quantitation at the MS2 level, a strategy that is known as higher order multiplexing. This is, to our knowledge, the first time that higher order multiplexing is applied to affinity purification - mass spectrometry experiments. In the future, we anticipate that this new workflow will be a useful tool to investigate transcription factor biology.
HostingRepository	PRIDE
AnnounceDate	2023-11-14
AnnouncementXML	Submission_2023-11-14_09:05:43.182.xml
DigitalObjectIdentifier
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Unsupported dataset by repository
PrimarySubmitter	Cathrin Graewe
SpeciesList	scientific name: Homo sapiens (Human); NCBI TaxID: 9606;
ModificationList	TMT6plex-126 reporter+balance reagent acylated residue; 6x(13)C labeled residue; acetylated residue; monohydroxylated residue; iodoacetamide derivatized residue
Instrument	Orbitrap Exploris 480; Orbitrap Fusion

Revision	Datetime	Status	ChangeLog Entry
0	2023-04-20 01:18:28	ID requested
1	2023-10-08 03:55:18	announced
⏵ 2	2023-11-14 09:05:47	announced	2023-11-14: Updated project metadata.

Gr, ä, we C, Hernandez-Quiles M, Jansen PWTC, Brimmers A, Vermeulen M, Determining DNA-Protein Binding Affinities and Specificities from Crude Lysates Using a Combined SILAC/TMT Labeling Strategy. J Proteome Res, 22(8):2683-2693(2023) [pubmed]

submitter keyword: TMT, SILAC,DNA pulldowns, PAQMAN

Michiel Vermeulen
contact affiliation	Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Oncode Institute, Radboud University Nijmegen, 6525 GA Nijmegen, The Netherlands.
contact email	michiel.vermeulen@ru.nl
lab head
Cathrin Graewe
contact affiliation	Department of Molecular Biology, Faculty of Science, Radboud Institute for Molecular Life Sciences, Oncode Institute, Radboud University Nijmegen, 6525 GA Nijmegen, The Netherlands.
contact email	c.grawe@science.ru.nl
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/PXD041674

PRIDE project URI

• PRIDE
  1. PXD041674
     1. Label: PRIDE project
     2. Name: Determining DNA-protein binding affinities and specificities from crude lysates using a combined SILAC/TMT labeling strategy