Material Data Sheet
Fluorogenic tri-peptide substrate for measuring the trypsin-like peptidase activity of the 20S proteasome. The 20S complex is composed of 28 subunits, arranged in an α7β7β7α7 stoichiometry. Each of the two internal β-type rings harbors three different proteolytically active sites, provided by the amino-terminal residues of three constitutive subunits: β1 (post-glutamyl peptide hydrolase site), β2 (trypsin-like site) and β5 (chymotrypsin-like site). Ac-Arg-Leu-Arg-AMC is particularly useful due to the low Km, high specific activity and the fact that it is cleaved exclusively at the amide R-AMC bond to release fluorescent product.
Formula: C30H46N10O6 Formula Weight: 642.76
Purity:> 97% by reverse-phase HPLC.
Solubility:Soluble in DMSO, methanol and aqueous solutions. For best results, pellet dry compound prior to reconstitution. Solubilize at desired stock concentration.
Use & Storage
Add from DMSO stock directly to in vitro or in vivo assay at desired concentration. Activity with purified proteasome: Km = 78μM. Typical concentrations range from 10-100 μM. (λex: 380nm; λem: 460 nm)
Store lyophilized powder at -20°C for at least one year. Stock solution at -20°C for up to 1 month. Avoid multiple freeze/thaw cycles.
Kisselev, A.F., et al. (2006) J. Biol. Chem. 281:8582-8590.
Kisselev, A.F. and A.L.Goldberg. (2005) Meth. Enzy. 398:364-378
Rogders K.J. and Dean R.T. (2003) Intl. J. Biochem.Cell.Biol. 35:716-727