UBE2F (NCE2) [6His-tagged]

Ubiquigent

Catalog No: UBI-62-0023-020

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Species Reactivity: Human  
Source: E. coli expression  
Formula: 50 mM HEPES pH 7.5, 150 mM sodium chloride, 2 mM dithiothreitol, 10% glycerol  
Concentration: 1 mg/ml  
Molecular Weight: ~25 kDa  
Protein Sequence: Accession number: NP_542409. For full protein sequence information download the Certificate of Analysis pdf.  
Quality Control Protein Identification: Confirmed by mass spectrometry.  
Quality Control Activity: E2-NEDD8 Thioester Loading Assay: The activity of His-UBE2F was validated by loading E1 APP-BP1/Uba3 activated NEDD8 onto the active cysteine of the His-UBE2F E2 enzyme via a transthiolation reaction. Incubation of the APP-BP1/Uba3 and His-UBE2F enzymes in the presence of NEDD8 and ATP at 30 °C was compared at two time points, T0 and T10 minutes. Sensitivity of the NEDD8/His-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 proteosomal 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 ubiquitin-like 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).

Reference:

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.