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Ubiquitin aldehyde, (recombinant)

High purity inhibitor of deubiquitinylating enzymes (DUBs).
BML-UW8450-0050 50 µg 238.00 USD
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Replaces Prod. #: ALX-200-603

Synthetically modified recombinant ubiquitin. The modification of the C-terminal glycine carboxyl into an aldehyde results in a potent, highly specific inhibitor of all ubiquitin deconjugating enzymes, including ubiquitin C-terminal hydrolases (UCHs), ubiquitin-specific proteases (USPs) and deubiquitinylating enzymes (DUBs).

Product Details

Alternative Name:Ub-H
Sequence:Recombinant Ubiquitin (corresponding to UniProt sequence P62988). The C-terminal glycine carboxyl is synthetically modified to an aldehyde.
Source:Produced in E.coli.
UniProt ID:P0CG47 (UBB), P0CG48 (UBC), P62979 (RPS27A), P62987 (UBA52)
Formulation:Liquid. In aqueous solution containing 0.15M HCl.
Purity:≥95% (HPLC)
Quality Control:Ubiquitin Aldehyde (recombinant) activity was confirmed by its inhibition of human UCH-L1 and deubiquitinylation of Ubiquitin-AMC (Prod. No. BML-SE211).
Activity:Ubiquitin Aldehyde (recombinant) activity was confirmed by its inhibition of UCH-L3 (BML-UW9745) and USP2 (BML-UW9850) deubiquitinylation of Ubiquitin-AMC (BML-SE211).
Specific Activity:Ki=2.5nM vs. UCH isopeptidase-T.
Application Notes:Ubiquitin Aldehyde (recombinant) is useful in the stabilisation of ubiquitin-protein conjugates in vitro, enhancing their accumulation in cell lysates and tissue extracts. Inhibition of deubiquitinylating enzyme activity by Ubiquitin Aldehyde (recombinant) can be used to identify and confirm such activity and to determine the inhibition kinetics for a particular enzyme. Recommended concentration for maximal inhibition is 2-5µM.
Co-crystallisation of ubiquitin aldehyde with specific deubiquitinylating enzymes (the inhibitor mimics the natural ubiquitin substrate) has also been used to probe enzyme:substrate interactions.
Shipping:Blue Ice Not Frozen
Long Term Storage:+4°C
Handling:Do not neutralise until immediateley prior to use. Do not lyophilize. Avoid presence of amino-containing compounds. Soluble and stable in aqueous solution at pH <7.0.
Technical Info/Product Notes:Typical assay set-up:
Substrate concentration: 0.01-1.0µM.
Enzyme concentration, UCH-L1 (human) (recombinant) (His) (BML-UW9740): 10-100nM.
Inhibitor concentration: 0.01-1.0µM.
Release of AMC fluorescence by DUB enzymes can be monitored using 380nm excitation and 460nm emission wavelengths.
Regulatory Status:RUO - Research Use Only
Ubiquitin aldehyde, (recombinant) image
Figure 1: Typical HPLC analysis of Ubiquitin Aldehyde (recombinant) (Prod. No. BML-UW8450). Method: Column: VYDAC 218TP54; A: 0.1% TFA/H2O; B: 0.1% TFA/CH3CN; Gradient: 25-45% B/20 mins. Monitoring: 230nm.
Ubiquitin aldehyde, (recombinant) image
Figure 2: Typical DUB Assay.Method: 12.5nM UCH-L1 (PGP9.5) (human) (recombinant) (His) (Prod. No. BML-UW9740); 500nM Ubiquitin-AMC; 500nM Ubiquitin Aldehyde (recombinant); 50mM HEPES pH7.8, 0.5mM EDTA, 1mM DTT."
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Product Literature References

Stability of SARS-CoV-2-Encoded Proteins and Their Antibody Levels Correlate with Interleukin 6 in COVID-19 Patients: W. Li, et al.; mSystems 7, e0005822 (2022), Abstract;
Lipopolysaccharide modulates p300 and Sirt1 to promote PRMT1 stability via an SCFFbxl17-recognized acetyldegron: Y. Lai, et al.; J. Cell Sci. 130, 3578 (2017), Abstract; Full Text
PEPD is a pivotal regulator of p53 tumor suppressor: L. Yang, et al.; Nat. Commun. 8, 2052 (2017), Abstract; Full Text
Ubiquitin Carboxy-Terminal HydrolaseL3 Correlates with Human Sperm Count, Motility and Fertilization: M. Wang, et al.; PLoS One 11, e0165198 (2016), Application(s): Determination of UCH enzymatic activity of spermatozoa , Abstract; Full Text
F-box protein Fbxl18 mediates polyubiquitylation and proteasomal degradation of the pro-apoptotic SCF subunit Fbxl7: Y. Liu, et al.; Cell Death Dis. 6, e1630 (2015), Application(s): Assay, Western Blotting, Immunoprecipitation, Abstract; Full Text
Pro-apoptotic F-box protein Fbxl7 regulates mitochondrial function by mediating the ubiquitylation and proteasomal degradation of survivin: Y. Liu, et al.; J. Biol. Chem. 290, 11843 (2015), Application(s): Assay, Abstract; Full Text
Differential effects of ubiquitin aldehyde on ubiquitin and ATP-dependent protein degradation.: J. R. Shaeffer & R. E. Cohen; Biochemistry 35, 10886 (1996), Abstract;

General Literature References

Structure and mechanisms of the proteasome-associated deubiquitinating enzyme USP14. : M. Hu et al.; EMBO J. 24 , 3747 (2005), Abstract; Full Text
The polyglutamine neurodegenerative protein ataxin-3 binds polyubiquitylated proteins and has ubiquitin protease activity. : B. Burnett et al.; Hum. Mol. Genet. 12 , 3195 (2003), Abstract; Full Text
A novel active site-directed probe specific for deubiquitylating enzymes reveals proteasome association of USP14. : A.Borodovsky et al.; EMBO J. 20, 5187 (2001), Abstract; Full Text
Structural basis for the specificity of ubiquitin C-terminal hydrolases.: S. C. Johnston et al.; EMBO J. 18, 3877 (1999), Abstract; Full Text
Ubiquitin-proteasome-dependent degradation of apolipoprotein B100 in vitro.: N. Sakata & J. L. Dixon; Biochim. Biophys Acta. 1437, 71 (1999), Abstract;
Kinetic and mechanistic studies on the hydrolysis of ubiquitin C-terminal 7-amido-4-methylcoumarin by deubiquitinating enzymes. : L. C. Dang et al.; Biochemistry 37, 1868 (1998), Abstract;
Kinetic studies on the inhibition of isopeptidase T by ubiquitin aldehyde.: F. Melandri et al.; Biochemistry 35 , 12893 (1996), Abstract;
Recognition of modified forms of ribonuclease A by the ubiquitin system.: R. L. Dunten et al.; J. Biol. Chem. 264, 16739 (1989), Abstract; Full Text
Ubiquitin-aldehyde: a general inhibitor of ubiquitin-recycling processes.: A. Hershko & I. A. Rose; PNAS 84, 1829 (1987), Abstract; Full Text

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