Enabling Protein Degradation Drug Discovery

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  • Name
    Catalogue Number
    Size
    Price
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  • Name:
    UBE2D1 (UbcH5a) [6His-tagged]
    Catalogue Number:
    62-0008-100
    Size:
    100 µg
    Price:
    £325
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  • Species
    Human
  • Source
    E. coli expression
  • Quantity
    100 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
    ~20 kDa
  • Stability
    12 months at -70°C; aliquot as required
  • Protein Sequence
    Accession number: NP_003329. 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-UBE2D1 was validated by loading E1 UBE1 activated ubiquitin onto the active cysteine of the His-UBE2D1 E2 enzyme via a transthiolation reaction. Incubation of the UBE1 and His-UBE2D1 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-UBE2D1 thioester bond to the reducing agent DTT was confirmed.

     

The enzymes of the ubiquitylation pathway play a pivotal role in a number of cellular processes including regulated and targeted proteosomal 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). UBE2D1 is a member of the E2 ubiquitin-conjugating enzyme family and cloning of the human gene was first described by Scheffner et al. (1994). UBE2D1 shares 89% sequence identity with its Drosophila homologue and mediates E6/UBE3A (E6AP)-induced ubiquitylation of p53 (Jensen et al., 1995; Scheffner et al., 1994). Ubiquitylation of the yeast PTS1 import receptor (pex5p) has been demonstrated in an in vitro assay in the presence of the human UBE2D1 in combination with the ring domain of the yeast E3 ligase pex10p (Williams et al., 2008). Sequence encoding the stimulated Iron transport gene SFT overlaps with intron 7 and exon 6 of UBE2D1, and RT/PCR has shown significantly upregulated levels of UBE2D1 in livers of iron-overloaded patients with hereditary hemochromatosis (Gehrke et al., 2003). 

References:

Gehrke SG, Riedel HD, Herrmann T, Hadaschik B, Bents K, Veltkamp C, Stremmel W (2003) UbcH5A, a member of human E2 ubiquitin-conjugating enzymes, is closely related to SFT, a stimulator of iron transport, and is up-regulated in hereditary hemochromatosis. Blood 101, 3288-93.

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.

Scheffner M, Huibregtse JM, Howley PM (1994) Identification of a human ubiquitin-conjugating enzyme that mediates the E6-AP-dependent ubiquitination of p53. Proc Natl Acad Sci USA 91, 8797-801.

Williams C, van den Berg M, Geers E, Distel B (2008) Pex10p functions as an E3 ligase for the Ubc4p-dependent ubiquitination of Pex5p. Biochem Biophys Res Commun 374, 620-4.