UBE2D3 (UbcH5c) [6His-tagged]


Catalogue Number
62-0074-020
Product Size
20 µg
Price £
£100
Accession Number
NP_003331
Residues Expressed
2-147
Alternate Product Size
100 µg
Certificate of Analysis Size
20 µg
Species
Human
Source
E. coli expression
Quantity
20 μg
Storage
-70°C
Concentration
1 mg/ml
Formulation
50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol
Molecular Weight
~19 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_003331. For full protein sequence information download the Certificate of Analysis pdf.
QA; Protein Identification
Confirmed by mass spectrometry.
QA Activity

E2-Ubiquitin Thioester Loading Assay: The activity of His-UBE2D3 was validated by loading E1 UBE1 activated ubiquitin onto the active cysteine of the His-UBE2D3 E2 enzyme via a transthiolation reaction. Incubation of the UBE1 and His-UBE2D3 enzymes in the presence of ubiquitin and ATP at 30°C was compared at two time points, T0 and T10 minutes. Sensitivity of the ubiquitin/His-UBE2D3 thioester bond to the reducing agent DTT was confirmed.


Background

The enzymes of the ubiquitylation pathway play a pivotal role in a number of cellular processes including the regulated and targeted proteasomal degradation of substrate proteins. Three classes of enzymes are involved in the process of ubiquitylation; activating enzymes (E1s), conjugating enzymes (E2s) and protein ligases (E3s). UBE2D3 is a member of the E2 ubiquitin-conjugating enzyme family and cloning of the gene was first described by Jensen et al. (1995). Human UBE2D3 shares 94% and 79% sequence identity with the Drosophila and S. cerevisiae homologues respectively. The E3 ligase E6AP mediates the conjugation of ubiquitin to targets such as p53 via UBE2D3 (Jensen et al., 1995). Upregulation of UBE2D3 following treatment with Retinoic Acid (RA) has been shown to induce differentiation and growth arrest in NB4 human promyelocytic cells. UBE2D3 also associates with Cyclin D1 and mediates retinoic acid induced cyclin D1 degradation (Hattori et al., 2007). Activation of the IKK complex is mediated by unanchored polyubiquitin chains formed by UBE2D3 and TRAF6 (Xia et al., 2009). Zipper-Interacting Protein Kinase (ZIPK) is a serine/threonine kinase implicated in cell death and transcriptional regulation, UBE2D3 induces ZIPK accumulation in promyelocytic leukaemia protein nuclear bodies resulting in their ubiquitylation (Ohbayashi et al., 2008). Meibomian Cell Carcinoma (MCC) is a malignant tumour of the meibomian glands located in the eyelids. UBE2D3 has been identified by RT/PCR as one of five genes found to be upregulated in MCC tumours (Kumar et al., 2007).


References

Hattori H, Zhang X, Jia Y, Subramanian KK, Jo H, Loison F, Newburger PE, Luo HR (2007) RNAi screen identifies UBE2D3 as a mediator of all-trans retinoic acid-induced cell growth arrest in human acute promyelocytic NB4 cells. Blood 110, 640-50.

Jensen JP, Bates PW, Yang M, Vierstra RD, Weissman AM (1995) Identification of a family of closely related human ubiquitin conjugating enzymes. J Biol Chem 270, 30408-14.

Kumar A, Kumar Dorairaj S, Prabhakaran VC, Prakash DR, Chakraborty S (2007) Identification of genes associated with tumorigenesis of meibomian cell carcinoma by microarray analysis. Genomics 90, 559-66.

Ohbayashi N, Okada K, et al. (2008) Physical and functional interactions between ZIP kinase and UbcH5. Biochem Biophys Res Commun 372, 708-12.

Xia ZP, Sun L, Chen X, Pineda G, Jiang X, Adhikari A, Zeng W, Chen ZJ (2009) Direct activation of protein kinases by unanchored polyubiquitin chains. Nature 461, 114-9.