The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Specific Activity: 0.00049 pmol/min/µg
Assay conditions: 50 µl reaction mix (4 mM ATP, 5mM MgCl2, 1mM DTT, E1 (0.5 µg), E2 (2.8 µg), Biotin-Ubiquitin (0.5 µg) and SMURF 2 (0-2 µg) in 50mM Tris pH 7.4) incubate for 30min at 37°C, then add to streptavidin coated plates and incubate for 30min at 37°C. Add antibody against SMURF 2, incubate 1h. Finally, incubate 1h with the Europeum-labeled secondary antibody. Time resolved fluorescence measured at room temperature (Lambda exc=330 nm, Lambda em=620 nm).
% . Purity is >80% by SDS-PAGE.
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Preparation and Storage
Stability and Storage
Shipped on Dry Ice. Store at -80°C.
pH: 8.00 Constituents: 20% Glycerol, 0.58% Sodium chloride, 0.395% Tris HCl, 0.05% Tween, 0.0462% DTT Note: 102 µg/ml FLAG peptide
This product is an active protein and may elicit a biological response in vivo, handle with caution.
E3 ubiquitin-protein ligase SMURF2
Smad specific E3 ubiquitin ligase 2
SMAD specific E3 ubiquitin protein ligase 2
SMAD ubiquitination regulatory factor 2
SMAD-specific E3 ubiquitin-protein ligase 2
Ubiquitin protein ligase SMURF2
E3 ubiquitin-protein ligase which accepts ubiquitin from an E2 ubiquitin-conjugating enzyme in the form of a thioester and then directly transfers the ubiquitin to targeted substrates. Interacts with SMAD1 and SMAD7 in order to trigger their ubiquitination and proteasome-dependent degradation. In addition, interaction with SMAD7 activates autocatalytic degradation, which is prevented by interaction with SCYE1. Forms a stable complex with the TGF-beta receptor-mediated phosphorylated SMAD2 and SMAD3. In this way, SMAD2 may recruit substrates, such as SNON, for ubiquitin-mediated degradation. Enhances the inhibitory activity of SMAD7 and reduces the transcriptional activity of SMAD2. Coexpression of SMURF2 with SMAD1 results in considerable decrease in steady-state level of SMAD1 protein and a smaller decrease of SMAD2 level.
The second and third WW domains are responsible for interaction with the PY-motif of R-SMAD (SMAD1, SMAD2 and SMAD3). The C2 domain is involved in autoinhibition of the catalytic activity by interacting with the HECT domain.
Auto-ubiquitinated and ubiquitinated in the presence of RNF11 and UBE2D1.
Nucleus. Cytoplasm. Cell membrane. Membrane raft. Cytoplasmic in the presence of SMAD7. Co-localizes with CAV1, SMAD7 and TGF-beta receptor in membrane rafts.