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Ubiquitination Affinity Beads

Ubiquitination Affinity Beads (40 assays)
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UBA01B-beads
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MSDS
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As part of the Signal-Seeker™ product line, Ubiquitin affinity beads have been optimized in order to detect endogenous levels of mono- and poly-ubiqutinated proteins, which often represent <1% of the target protein. UBA01B-Beads can enrich all ubiquitinated species with a high affinity, including mono-, multi- and polyubiquitin chains (see Application 1 below). The affinity moeities have been cross-linked to beads for low leaching and cleaner detection of the protein of interest. A comprehensive Signal-Seeker™ Ubiquitination Detection Kit is also available (BK161) and is recommended for first time users.

Note: The product #s have been updated from UBA01 and CUB02 to UBA01B and CUB02B beads due to a reformulation of the bead preservation buffer. Reformulation results in an enhanced bead performance post-lyophilization. The affinity protein formulation on the beads has not changed.

Validation Data: Ubiquitination Affinity Beads White Paper

Each lot of affinity-bead is quality controlled  to provide high batch to batch consistency, see COA documents.

Click Here for Detailed Methods

Validated Applications

Application 1: Investigate mono- and poly-ubiquitination
The activity of the Rho family of proteins, including Rac1, is known to be regulated, in part, through ubiquitination (also termed ubiquitylation) events that can lead to signaling pathway regulation via degradation and/or localization of the modified protein (Visvikis, O. et al. 2010. Biol. Cell 102: 377-389. Nethe, M., & Hordijik, P.2010. J. Cell Sci. 123: 4011-4018). In many cases, the GTP-bound active form of Rac1 is the preferred substrate for ubiquitination. For example, it has been shown that cells treated with the bacterial toxin CNF1 leads to constitutive activation of Rac1 and subsequent mono- and poly-ubiquitination (Pop, M. et al. 2004. J. Biol.Chem. 279: 35840-35848).
Using ubiquitin affinty beads (Cat. # UBA01B-beads) as part of the Signal Seeker™ Ubiquitination Detection Kit (Cat # BK161) we examined the ubiquitination of endogenous Rac1 in 3T3 cells treated with CNF1 toxin (Cat # CN04) and found that both mono- and polyubiquitination of Rac1 could be detected from 300 μg of 3T3 cell lysate. The kit offers a user friendly tool to examine mono- and poly-ubiquitination for any target protein.
CUB02_product_page_figure

Figure 1: Swiss 3T3 cells were pre-treated with MG-132, then either untreated or treated with CNF1 (CN04) for 3 hours prior to lysis with BlastR buffer. The BK161 kit was utilized to perform the IP on 300 μg of lysate per condition. Lane 1: 3 μg input untreated lysate, Lane 2: Ubiquitin Affinity beads (UBA01) plus untreated lysate, Lane 3: UBA01 plus CN04 treated lysate, Lane 4: ubiquitin control beads (CUB02) plus untreated lysate, Lane 5: CUB02 plus CN04 treated lysate. Samples were analyzed for Rac1 ubiquitination using an anti-Rac1 antibody.

Amount:
Each package contains enough ubiquitin beads for 40 reactions and sufficient control beads for 5 reactions.

 

For more information contact:  signalseeker@cytoskeleton.com

Associated Products:

Signal-Seeker™ Ubiquitination Detection Kit (Cat. # BK161)

Signal-Seeker™ SUMOylation 2/3 Detection Kit (Cat. # BK162)

Signal-Seeker™ Phosphotyrosine Detection Kit (Cat. # BK160)

Signal-Seeker™: BlastR™ Rapid Lysate Prep Kit (Cat. # BLR01)

Signal-Seeker™: PTMtrue™ Ubiquitin Antibody (Cat.# AUB01)

 

    AuthorTitleJournalYearArticle Link
    Otani, Takahito et al.The role of adhesion molecules in osteocalcin-induced effects on glucose and lipid metabolism in adipocytesBiochimica et Biophysica Acta (BBA) - Molecular Cell Research2024
    Ruggeri, Edoardo et al.Protein kinase CK2α is overexpressed in classical hodgkin lymphoma, regulates key signaling pathways, PD-L1 and may represent a new target for therapyFrontiers in Immunology2024
    Shi, Shujie et al.Control of ENaC ubiquitinationRenal Physiology2024
    Kotani, Hiroshi et al.Dual inhibition of SUMOylation and MEK conquers MYC-expressing KRAS-mutant cancers by accumulating DNA damageJournal of Biomedical Science2024
    Hołda, Mateusz K. et al.Autophagy and ubiquitin-dependent proteolysis processes in left ventricular mass loss in pulmonary arterial hypertensionScientific Reports2024
    Liu, Shiwei et al.O-GlcNAcylated RALY Contributes to Hepatocellular Carcinoma Cells Proliferation by Regulating USP22 mRNA Nuclear ExportInternational Journal of Biological Sciences2024
    Yu, Hui et al.MELK aggravates lung adenocarcinoma by regulating EZH2 ubiquitination and H3K27me3 histone methylation of LATS2Journal of Cellular and Molecular Medicine2024
    Steinbach, Adriana et al.Cross-family small GTPase ubiquitination by the intracellular pathogen Legionella pneumophilaMolecular biology of the cell2024
    Hotani, Takuma et al.Involvement of Protein Kinase R in Double-Stranded RNA-Induced Proteasomal Degradation of Hypoxia Inducible Factor-1αInflammation2023
    Chen, Shanlin et al.Aloe-emodin targets multiple signaling pathways by blocking ubiquitin-mediated degradation of DUSP1 in nasopharyngeal carcinoma cellsPhytotherapy Research2023
    Guo, Qianjin et al.TMEM127 suppresses tumor development by promoting RET ubiquitination, positioning, and degradationCell reports2023
    Huang, Xiaogang et al.Unfolded protein response inhibits KAT2B/MLKL-mediated necroptosis of hepatocytes by promoting BMI1 level to ubiquitinate KAT2BOpen Medicine2023
    Tahaei, Ebrahim et al.Pendrin regulation is prioritized by anion in high-potassium dietsAmerican Journal of Physiology - Renal Physiology2023
    Wang, Yuan et al.MTSS1 curtails lung adenocarcinoma immune evasion by promoting AIP4-mediated PD-L1 monoubiquitination and lysosomal degradationCell Discovery 2023
    Román-Fernández, Alvaro et al.Spatial regulation of the glycocalyx component podocalyxin is a switch for prometastatic functionScience Advances2023
    Nguyen, Kha The et al.The MARCHF6 E3 ubiquitin ligase acts as an NADPH sensor for the regulation of ferroptosisNature Cell Biology 2022
    Dieter, Sebastian M. et al.Degradation of CCNK/CDK12 is a druggable vulnerability of colorectal cancerCell Reports2021
    Gu, Chunming et al.Identification of berberine as a novel drug for the treatment of multiple myeloma via targeting UHRF1BMC Biology2020
    Horita, Henrick et al.Utilizing a comprehensive immunoprecipitation enrichment system to identify an endogenous post-translational modification profile for target proteinsJournal of Visualized Experiments2018
    Horita, Henrick et al.Utilizing optimized tools to investigate PTM crosstalk: Identifying potential PTM crosstalk of acetylated mitochondrial proteinsProteomes2018
    Horita, Henrick et al.A simple toolset to identify endogenous post-translational modifications for a target protein: A snapshot of the EGFR signaling pathwayBioscience Reports2017

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