UBE2F (NCE2) [GST-tagged]


Catalogue Number
62-0024-100
Product Size
100 µg
Price £
£325
Accession Number
NP_542409
Residues Expressed
1-185
Alternate Product Size
20 µg
Certificate of Analysis Size
100 μg
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
~48 kDa
Stability
12 months at -70°C; aliquot as required
Protein Sequence
Accession number: NP_542409. For full protein sequence information download the Certificate of Analysis pdf.
QA; Protein Identification
Confirmed by mass spectrometry.
QA Activity

E2-NEDD8 Thioester Loading Assay: The activity of GST-UBE2F was validated by loading E1 APP-BP1/Uba3 activated NEDD8 onto the active cysteine of the GST-UBE2F E2 enzyme via a transthiolation reaction. Incubation of the APP-BP1/Uba3 and GST-UBE2F enzymes in the presence of NEDD8 and ATP at 30oC was compared at two time points, T0 and T10 minutes. The sensitivity of this NEDD8/GST-UBE2F thioester bond to the reducing agent DTT was confirmed.

 

Background

The enzymes of the NEDDylation pathway play a pivotal role in a number of cellular processes including the indirect regulation and targeting of substrate proteins for proteasomal degradation. Three classes of enzymes are involved in the process of NEDDylation; the ubiquitin-like activating enzyme APP-BP1/Uba3 (E1), the ubiquitin-like conjugating enzymes (E2s) and protein ligases (E3s). UBE2F is a member of the E2 conjugating enzyme family and the human gene was first described by Huang et al. (2009). UBE2F acts as a NEDD8 conjugating enzyme both in vitro and in vivo. UBE2F accepts the ubiquitin-like protein NEDD8 from the Uba3-NAE1 (APP-BP1/Uba3) E1 complex and catalyzes its covalent attachment to other proteins. The specific interaction of UBE2F with the E3 ubiquitin ligase RBX2, but not RBX1, suggests that the RBX2-UBE2F complex NEDDylates specific target proteins such as CUL5, a component of one of the many Cullin Ring Ligases (CRLs) (Huang et al., 2009).


References

Huang DT, Ayrault O, Hunt HW, Taherbhoy AM, Duda DM, Scott DC, Borg LA, Neale G, Murray PJ, Roussel MF, Schulman BA (2009) E2-RING expansion of the NEDD8 cascade confers specificity to cullin modification. Mol Cell 33, 483-95.