Product Details
| MW: | ~9.5kDa |
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| Source: | Produced in E. coli. |
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| UniProt ID: | P0CG47 (UBB), P0CG48 (UBC), P62979 (RPS27A), P62987 (UBA52) |
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| Formulation: | Lyophilized. |
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| Purity: | ≥95% (SDS-PAGE) |
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| Purity Detail: | Purified by multi-step chromatography. |
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| Application Notes: | For use in ubiquitinylation studies. |
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| Reconstitution: | Reconstitute in minimum volume of DMSO (5-10µL) followed by dilution to required concentration with aqueous buffer of choice (e.g. 1mg/mL in PBS). |
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| Shipping: | Blue Ice |
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| Short Term Storage: | -20°C |
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| Long Term Storage: | -20°C |
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| Use/Stability: | Stable for at least 1 year after receipt when stored at -20°C. Reagent is light sensitive; store in the dark or in amber vials. Stable to freeze/thaw but this should be minimized wherever possible. |
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| Handling: | Protect from light. |
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| Scientific Background: | The covalent attachment of ubiquitin to proteins (ubiquitinylation) and their subsequent proteasomal degradation plays a fundamental role in the regulation of cellular function through biological events involving cell cycle, differentiation, immune responses, DNA repair, chromatin structure, and apoptosis. |
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| Technical Info/Product Notes: | Fluorescein labelled ubiquitin is a useful reagent for studying ubiquitinylation in vitro, where it represents a readily detectable alternative to radio-iodinated ubiquitin. Fluorescein labelled ubiquitin is activated by ubiquitin E1, is a competent substrate for known ubiquitin E2/E3 enzymes (thioester bond formation) and can be conjugated to ubiquitin target proteins (isopeptide bond formation).
This product contains a number of multiply fluorescein-labelled ubiquitin species, with [fluorescein]2-4ubiquitin being the predominant species.
Typical thioester assay set-up: 2.5µM Fluorescein-Ub (BML-UW1240), 100nM Ub-E1 (BML-UW9410), 2.5µM E2 (e.g. His6-UbcH5a, BML-UW9050), 1U inorganic pyrophosphatase, 5mM Mg-ATP in 20mM TRIS-HCl, pH 7.5, 1mM DTT. Incubate at 37ºC for 30-60 mins. Analyse by SDS-PAGE followed by fluorescence detection. |
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| Regulatory Status: | RUO - Research Use Only |
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Product Literature References
Mechanisms underlying ubiquitination: C.M. Pickart; Annu. Rev. Biochem.
70, 503 (2001),
Abstract;
The ubiquitin-proteasome system and endocytosis: G.J. Strous & R. Govers; J. Cell Sci.
112 (Pt 10), 1417 (1999),
Abstract;
The ubiquitin system: A. Hershko et al.; Annu. Rev. Biochem.
67, 425 (1998),
Abstract;
Pathways of ubiquitin conjugation: A.L. Haas & T.J. Siepmann; FASEB J.
11, 1257 (1997),
Abstract;
