Active Caspase 2 (CASP2)

Caspase-2 (CASP2) is a 3032 kDa member of the peptidase C14A/IL1 beta-converting family of enzymes. It is widely expressed and is an integral component of the apoptotic cascade. Based on the length of its prodomain, caspase-2 has been considered to be an initiator caspase. Human procaspase-2 is a 4851 kDa, 452 amino acid (aa) protein. It is known to exist as a disulfide-linked homodimer via covalent linkage at Cys436. But this dimeric state may not be sufficient for (auto) activation. Actual activation may occur following oligomerization within the context of activating platforms such as DISC (death-inducing signaling complex) or the PIDDosome. The activity assay of recombinant human CASP2 was measured by its ability to cleave the fluorogenic peptide substrate Ac-VDVAD-AFC. The reaction was performed in 25 mM HEPES, 0.1% (w/v) CHAPS, 10 mM dithiothreitol (DTT), pH 7.5 ( Assay Buffer). The CASP2 was diluted to 3 ug/ml by assay buffer and incubated at room temperature for 15min. The reaction was initiated by adding 50 ul 3 ug/ml CASP2 to 50 ul of 200 uM substrate and then read at excitation and emission wavelengths of 400 nm and 505 nm (top read), respectively, in kinetic mode for 5 minutes. The specific activity of recombinant human CASP2 is 2100 pmol/min/µg.