UBA6 [6His-tagged]


Catalogue Number
61-0002-010
Product Size
10 μg
Price £
£100
Accession Number
NP_060697
Residues Expressed
1-1052
Alternate Product Size
50 µg
Certificate of Analysis Size
10 µg
Species
Human
Source
Sf21 insect cell-baculovirus expression
Quantity
10 μg
Storage
-70°C
Concentration
0.5 mg/ml
Formulation
50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol
Molecular Weight
~123 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_060697. For full protein sequence information download the Certificate of Analysis pdf.
QA; Protein Identification
Confirmed by mass spectrometry.
QA Activity

E1-Ubiquitin Thioester Loading Assay: The activity of His-UBA6 was validated by loading ubiquitin onto the active cysteine of His-UBA6. Incubation of the His-UBA6 enzyme 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-UBA6 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 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). UBA6 is a member of the E1 activating enzyme family and the human gene was first described by Jin et al. (2007). UBA6 shares 42% homology with UBE1 and contains all the structural elements required for E1 enzyme activity (Groettrup et al., 2008). UBA6 interacts with a number of E2 and E3 enzymes and has been shown to be involved with p53 ubiquitylation in vitro (Groettrup et al., 2008; Pelzer et al., 2007). UBA6 activates ubiquitin and the ubiquitin-like protein human leukocyte antigen F-Associated Transcript 10 (FAT10), both of which may serve as a signal for proteasomal degradation. FAT10, is encoded by the major histocompatibility (MHC) class I locus, and its expression is induced by tumor necrosis factor alpha (TNFα) and interferon-gamma (IFNγ). FAT10 expression is significantly upregulated in hepatocellular carcinoma as well as in gastrointestinal and gynecological cancers (Lee et al., 2003), however its precise biochemical and cellular functions have yet to be determined. Although FAT10 forms covalent conjugates with cellular proteins through its C-terminal diglycine motif (Raasi et al., 2001), substrates remain to be identified. Knockdown of UBA6 results in decreased FAT10 conjugation, indicating that UBA6 is required to activate FAT10 and facilitate its conjugation (Chiu et al., 2007).


References

Chiu YH, Sun Q, Chen ZJ (2007) E1-L2 activates both ubiquitin

Groettrup M, Pelzer C, Schmidtke G, Hofmann K (2008) Activating the ubiquitin family: UBA6 challenges the field. Trends Biochem Sci 33, 230-7.

Jin J, Li X, Gygi SP, Harper JW (2007) Dual E1 activation systems for ubiquitin differentially regulate E2 enzyme charging. Nature 447, 1135-8.

Lee CG, Ren J, Cheong IS, Ban KH, Ooi LL, Yong Tan S, Kan A, Nuchprayoon I, Jin R, Lee KH, Choti M, Lee LA (2003) Expression of the FAT10 gene is highly upregulated in hepatocellular carcinoma and other gastrointestinal and gynecological cancers. Oncogene 22, 2592-603.

Pelzer C, Kassner I, Matentzoglu K, Singh RK, Wollscheid HP, Scheffner M, Schmidtke G, Groettrup M (2007) UBE1L2, a novel E1 enzyme specific for ubiquitin. J Biol Chem 282, 23010-4.

Raasi S, Schmidtke G, Groettrup M (2001) The ubiquitin-like protein FAT10 forms covalent conjugates and induces apoptosis. J Biol Chem 276, 35334-43.