PXD040302 is an
original dataset announced via ProteomeXchange.
Dataset Summary
Title | Control of TurboID-dependent biotinylation intensity in proximity ligation screens |
Description | Proximity biotinylation screens are a widely used strategy for the unbiased identification of interacting or vicinal proteins. The latest generation biotin ligase TurboID has broadened the range of potential applications, as this ligase promotes an intense and faster biotinylation, even in subcellular compartments like the endoplasmic reticulum. On the other hand, the uncontrollable high basal biotinylation rates deny the system´s inducibility and are often associated with cellular toxicity precluding its use in proteomics. We report here an improved method for TurboID-dependent biotinylation reactions based on the tight control of free biotin levels. Blockage of free biotin with a commercial biotin scavenger reversed the high basal biotinylation and toxicity of TurboID, as shown by pulse-chase experiments. Accordingly, the biotin-blockage protocol restored the biological activity of a bait protein fused to TurboID in the endoplasmic reticulum and rendered the biotinylation reaction inducible by exogenous biotin. Importantly, the biotin-blockage protocol was more effective than biotin removal with immobilised avidin and did not affect the cellular viability of human monocytes over several days. The method presented should be useful to researchers interested in exploiting the full potential of biotinylation screens with TurboID and other high-activity ligases for challenging proteomics questions. |
HostingRepository | PRIDE |
AnnounceDate | 2023-11-14 |
AnnouncementXML | Submission_2023-11-14_09:00:06.260.xml |
DigitalObjectIdentifier | |
ReviewLevel | Peer-reviewed dataset |
DatasetOrigin | Original dataset |
RepositorySupport | Unsupported dataset by repository |
PrimarySubmitter | Thomas Krüger |
SpeciesList | scientific name: Homo sapiens (Human); NCBI TaxID: 9606; |
ModificationList | biotinylated residue; acetylated residue; monohydroxylated residue; iodoacetamide derivatized residue |
Instrument | Orbitrap Exploris 480 |
Dataset History
Revision | Datetime | Status | ChangeLog Entry |
0 | 2023-02-21 02:17:53 | ID requested | |
⏵ 1 | 2023-11-14 09:00:06 | announced | |
Publication List
Dataset with its publication pending |
Keyword List
submitter keyword: Biotinylation, Proximity ligation screens, Protein-Protein Interactions, Proteomics |
Contact List
Axel A. Brakhage |
contact affiliation | Department of Molecular and Applied Microbiology, Leibniz Institute for Natural Product Research and Infection Biology, Hans Knöll Institute Jena, Leibniz-HKI, Adolf-Reichwein-Str. 23, 07745 Jena, Germany |
contact email | axel.brakhage@leibniz-hki.de |
lab head | |
Thomas Krüger |
contact affiliation | Leibniz Institute for Natural Product Research and Infection Biology - Hans Knöll Institute |
contact email | thomas.krueger@leibniz-hki.de |
dataset submitter | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
[ + ]
[ - ]
- PRIDE
- PXD040302
- Label: PRIDE project
- Name: Control of TurboID-dependent biotinylation intensity in proximity ligation screens