UBE1 [6His-tagged]


Catalogue Number
61-0001-050
Product Size
50 µg
Price £
£175
Accession Number
NP_003325
Residues Expressed
2-1058
Alternate Product Size
10 μg
Certificate of Analysis Size
50 μg
Species
Human
Source
Sf21 insect cell-baculovirus expression
Quantity
50 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
~121 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_003325. 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-UBE1 was validated by loading ubiquitin onto the active cysteine of His-UBE1. Incubation of the His-UBE1 enzyme in the presence of ubiquitin and ATP at 30oC was compared at two time points, T0 and T10 minutes. Sensitivity of the ubiquitin/His-UBE1 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 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). UBE1 is a member of the E1 activating enzyme family and cloning of the human gene was first described by Handley et al. (1991). The UBE1 gene has been mapped to Xp11.3-p11.23 by highresolution fluorescence in situ hybridization (Takahashi et al., 1992). UBE1 ‘activates' ubiquitin through catalysing a C-terminal ATP dependent adenylation of the protein which results in it forming a high-energy thioester bond with the sulfhydryl group of UBE1. UBE1 is monomeric and there are two active sites within the UBE1 protein allowing it to bind two ubiquitin moieties at a time, with a new ubiquitin forming an adenylate intermediate as the previous one is transferred to the thiol site (Jin et al., 2007; Zheng et al., 2009). Defects in UBE1 are known to cause spinal muscular atrophy X-linked type 2 (SMAX2) also known as X-linked lethal infantile spinal muscular atrophy, distal X-linked arthrogryposis multiplex congenita or X-linked arthrogryposis type 1 (AMCX1). Spinal muscular atrophy refers to a group of neuromuscular disorders characterized by degeneration of the anterior horn cells of the spinal cord, leading to symmetrical muscle weakness and atrophy. SMAX2 is a lethal infantile form presenting with hypotonia, areflexia, and multiple congenital contractures (Ramser et al., 2008).


References

Handley PM, Mueckler M, Siegel NR, Ciechanover A, Schwartz AL (1991) Molecular cloning, sequence, and tissue distribution of the human ubiquitin-activating enzyme E1. Proc Natl Acad Sci USA 88, 258-62.

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

Ramser J, Ahearn ME, Lenski C, Yariz KO, Hellebrand H, von Rhein M, Clark RD, Schmutzler RK, Lichtner P, Hoffman EP, Meindl A, Baumbach-Reardon L. (2008) Rare missense and synonymous variants in UBE1 are associated with X-linked infantile spinal muscular atrophy. Am J Hum Genet 82, 188-93.

Takahashi E, Ayusawa D, Kaneda S, Itoh Y, Seno T, Hori T (1992) The human ubiquitin-activating enzyme E1 gene (UBE1) mapped to band Xp11.3----p11.23 by fluorescence in situ hybridization. Cytogenet Cell Genet 59, 268-9.

Zheng M, Liu J, Yang Z, Gu X, Li F, Lou T, Ji C, Mao Y (2009) Expression, purification and characterization of human ubiquitin-activating enzyme, UBE1. Mol Biol Rep 37, 1413-9.