PXD004059 is an
original dataset announced via ProteomeXchange.
Dataset Summary
Title | Mass spectrometry insights into a tandem ubiquitin‐binding domain hybrid engineered for the selective recognition of unanchored polyubiquitin |
Description | Unanchored polyubiquitin chains are emerging as important regulators of cellular physiology with diverse roles paralleling those of substrate-conjugated polyubiquitin. However tools able to discriminate unanchored polyubiquitin chains of different isopeptide linkages have not been described. We describe the design of a linker-optimised ubiquitin-binding domain hybrid (t-UBD) containing two UBDs, a ZnF-UBP domain in tandem with a linkage-selective UBA domain, which exploits avidity effects to afford selective recognition of unanchored Lys48-linked polyubiquitin chains. Utilising native MS to quantitatively probe binding affinities we confirm cooperative binding of the UBDs within the synthetic protein, and desired binding specificity for Lys48-linked ubiquitin dimers. Furthermore MS/MS analyses indicate that the t-UBD, when applied as an affinity enrichment reagent, can be used to favour the purification of endogenous unanchored Lys48-linked polyubiquitin chains from mammalian cell extracts. Our study indicates that strategies for the rational design and engineering of polyubiquitin chain-selective binding in non-biological polymers are possible, paving the way for the generation of reagents to probe unanchored polyubiquitin chains of different linkages and more broadly the ‘ubiquitome’. |
HostingRepository | PRIDE |
AnnounceDate | 2016-05-18 |
AnnouncementXML | Submission_2016-05-18_01:50:59.xml |
DigitalObjectIdentifier | https://dx.doi.org/10.6019/PXD004059 |
ReviewLevel | Peer-reviewed dataset |
DatasetOrigin | Original dataset |
RepositorySupport | Supported dataset by repository |
PrimarySubmitter | Andrew Bottrill |
SpeciesList | scientific name: Homo sapiens (Human); NCBI TaxID: 9606; |
ModificationList | Ammonia-loss; LeuArgGlyGly; Dehydrated; Oxidation; Acetyl; Carbamidomethyl; GlyGly |
Instrument | LTQ Orbitrap Velos |
Dataset History
Revision | Datetime | Status | ChangeLog Entry |
0 | 2016-04-26 08:23:08 | ID requested | |
⏵ 1 | 2016-05-18 01:51:00 | announced | |
Publication List
Scott D, Garner TP, Long J, Strachan J, Mistry SC, Bottrill AR, Tooth DJ, Searle MS, Oldham NJ, Layfield R, Mass spectrometry insights into a tandem ubiquitin-binding domain hybrid engineered for the selective recognition of unanchored polyubiquitin. Proteomics, 16(14):1961-9(2016) [pubmed] |
Keyword List
curator keyword: Biological |
submitter keyword: Ubiquitin, polyubiquitin |
Contact List
Rob Layfield |
contact affiliation | School of Life Sciences, Queen’s Medical Centre, University of Nottingham, Nottingham NG7 2UH, UK |
contact email | Robert.Layfield@nottingham.ac.uk |
lab head | |
Andrew Bottrill |
contact affiliation | University of Leicester |
contact email | arb29@le.ac.uk |
dataset submitter | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
[ + ]
[ - ]
- PRIDE
- PXD004059
- Label: PRIDE project
- Name: Mass spectrometry insights into a tandem ubiquitin‐binding domain hybrid engineered for the selective recognition of unanchored polyubiquitin