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Recombinant Human Parkin pS65 Protein, CF Summary
Reaction conditions will need to be optimized for each specific application. Unlike unphosphorylated Parkin (E3-160), Parkin pS65 has substantial ligase activity in the absence of added activators as determined by its abilityto autoubiquitinate, andubiquitinate a Miro/Rhot1 substrate(SP-490).
Product Datasheets
Carrier Free
CF stands for Carrier Free (CF). We typically add Bovine Serum Albumin (BSA) as a carrier protein to our recombinant proteins.Adding a carrier protein enhances protein stability, increases shelf-life, and allows the recombinant protein to be stored at a more dilute concentration.The carrier free version does not contain BSA.
In general, we advise purchasing the recombinant protein with BSA for use in cell or tissue culture, or as an ELISA standard.In contrast, the carrier free protein is recommended for applications, in which the presence of BSA could interfere.
E3-166
| Formulation | X mg/ml (X μM) in 25 mM Tris pH 8.5, 200 mM NaCl, 0.03% Brij35, 10% (v/v) Glycerol, 5 mM TCEP |
| Shipping | The product is shipped with dry ice or equivalent. Upon receipt, store it immediately at the temperature recommended below. |
| Stability & Storage: | Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
|
Reconstitution Calculator
Background: Parkin
The E3 Ubiquitin ligase Parkin (encoded by the PARK2 gene) is an essential part of the cellular machinery that participates in the removal of damaged mitochondria. Mutations in PARK2 are known to cause a form of Parkinson"s disease known as autosomal recessive juvenile Parkinson"s disease (AR-JP), and the mechanisms by which defective Parkin ligase contributes to the dopaminergic cell death in this disease is an area of intense investigation.
Reported substrates for Parkin include BCL2, GPR37, MIRO1, MFN1, MFN2, TOMM20, USP30, and many others. Parkin (an RBR-class Ubiquitin ligase) structures have recently been reported by multiple groups, and reveal that the ligase is folded upon itself to produce an auto-inhibited state. The auto-inhibition is relieved by interactions with PINK1 kinase (which can phosphorylate both Parkin and Ubiquitin at serine residue number 65) and pS65 phospho-Ubiquitin by mechanisms that are under investigation.
In vitro, Parkin may be activated by treatment with recombinant PINK1, or addition of low concentrations of pS65-phosphoubiquitin. Parkin has been reported to generate poly-Ubiquitin chains in K6, K11, K48, and K63 linkages both in vitro and in vivo. This recombinant protein is phosphorylated on serine 65, and is untagged.
- Bingol, B. et al. (2014) Nature 510: 370
- Ordureau, A. et al. (2014) Mol. Cell 56: 360
- Riley, B.E. et al. (2013) Nat. Comm. 4: doi:10.1038/ncomms2982
- Saraff, S.A. et al. (2013) Nature 496: 372
- Spratt, D.E. et al. (2013) Nat. Comm. 4: doi:10.1038/ncomms2983
- Trempe, J.F. et al. (2013) Science 340: 1451
- Wauer T. et. al. (2015) Nature 524: 370
- Wauer T. & Komander, D. (2013) EMBO J 32: 2099
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