⮝ Full datasets listing
PXD073487
PXD073487 is an original dataset announced via ProteomeXchange.
Dataset Summary
| Title | Native yeast kinetochore structures identify an essential inner kinetochore interaction. |
| Description | Kinetochores were purified from Saccharomyces cerevisiae (W303) cell lysate after being treated with benzonase and sucrose gradient to remove contaminant proteins. The endogenous AME1 kinetochore gene was C-terminally tagged with 3xM3DK. Harvested yeast were resuspended in Buffer H (25 mM HEPES pH 8.0, 150 mM KCl, 2 mM MgCl2, 0.1 mM EDTA pH 8.0, 0.1% NP-40, 15% glycerol) supplemented with protease inhibitors, phosphatase inhibitors, and 2 mM DTT. After resuspension and re-spinning, yeast pellets were frozen in liquid nitrogen and lysed using a Freezer Mill (SPEX, Metuchen NJ). Lysate was clarified via tabletop centrifugation at maximum speed for 30 minutes. The supernatant was pooled in ultracentrifugation tubes with 3mL sucrose gradient cushion from 25% to 40%. The lysate was ultra spun overnight at 27,000RPM and the clear layer of lysate was extracted with a syringe. This extract was incubated with 625ng/mL 500bp CEN3 DNA at room temperature for 90 minutes and magnetic a-M3DK antibody conjugated Dynabeads (Invitrogen, Waltham MA) were added and incubated for 90 minutes at 4 C with rotation. Dynabeads were washed with Buffer L (25 mM HEPES pH 8.0, 175 mM KGlutamate, 6 mM Mg(OAc)2, 0.1 mM EDTA pH 7.6, 0.5 mM EGTA-KOH, 0.1% NP-40, 15% glycerol, pH7.6) for 5 times (the last 2 washes omitting NP-40, glycerol and phosphatase inhibitors). For mass spectrometry, kinetochores were eluted from Dynabeads with 0.2% RapiGest (Waters Corporation, Milford MA) in 50 mM HEPES pH 8.0. |
| HostingRepository | MassIVE |
| AnnounceDate | 2026-01-27 |
| AnnouncementXML | Submission_2026-01-27_10:38:15.586.xml |
| DigitalObjectIdentifier | |
| ReviewLevel | Non peer-reviewed dataset |
| DatasetOrigin | Original dataset |
| RepositorySupport | Supported dataset by repository |
| PrimarySubmitter | Fred Hutch Proteomics Core |
| SpeciesList | scientific name: Saccharomyces cerevisiae; common name: baker's yeast; NCBI TaxID: 4932; |
| ModificationList | No PTMs are included in the dataset |
| Instrument | Orbitrap Eclipse |
Dataset History
| Revision | Datetime | Status | ChangeLog Entry |
|---|---|---|---|
| 0 | 2026-01-23 16:07:22 | ID requested | |
| ⏵ 1 | 2026-01-27 10:38:16 | announced |
Publication List
| no publication |
Keyword List
| submitter keyword: Mitosis, kinetochore, centromere, cell division, DatasetType:Proteomics |
Contact List
| Susan Biggins | |
|---|---|
| contact affiliation | Fred Hutchinson Cancer Center |
| contact email | sbiggins@fredhutch.org |
| lab head | |
| Fred Hutch Proteomics Core | |
| contact affiliation | Fred Hutchinson Cancer Research Center |
| contact email | pgafken@fredhutch.org |
| dataset submitter | |
Full Dataset Link List
| MassIVE dataset URI |
| 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://massive-ftp.ucsd.edu/v12/MSV000100570/ |
