BD Ac-DEVD-AFC Caspase-3 Substrate

Description

• The caspase family of cysteine proteases was discovered following a search of human cDNA libraries for sequences homologous to ced -3, a cell death gene described in the nematode worm C
• elegans
• The first mammalian homologue of ced-3 to be identified was ICE (interleukin-1α converting enzyme)
• Subsequently, numerous mammalian ced -3 homologues have been discovered and have each been given a variety of names
• To achieve consistency, the term “caspase” was adopted as a root name for all family members
• The name reflects the catalytic properties of these enzymes, the “c” denotes their cysteine protease mechanism and “aspase” refers to their ability to cleave after aspartic acid residues
• These proteases are expressed as inactive proenzymes, which are proteolytically cleaved into large and small subunits, which form the active enzyme
• Active caspase-3 consists of 17 and 12 kDa subunits which are derived from a 32 kDa proenzyme (pro-caspase-3)
• Activecaspase-3 has been shown to cleave PARP [poly (ADP ribose) polymerase], an enzyme that is involved in DNA repair and genomic maintenance
• Proteolysis of the 116 kDa intact form of PARP into 85 and 25 kDa subunits results in loss of normal PARP function
• The cleavage site in PARP is C-terminal to Asp-216
• The upstream sequence of the PARP cleavage site, DEVD (Asp-Glu-Val-Asp), is utilized as a basis for the highly specific caspase-3 substrate Ac(N-acetyle)-DEVD-AFC (7-amino-4-trifluoromethylcoumarin)
• Caspase-3 cleaves the tetrapeptide between D and AFC, thus releasing the fluorogenic AFC which can be quantified by U.V
• spectrofluorometry
• When coupled to an aldehyde group (CHO), the DEVD peptide functions as a potent inhibitor of caspase-3 activity and can be used to block caspase-3 mediated cleavage of Ac-DEVD-AFC
• These tetrapeptide substrates can be used to identify and quantitate caspase-3 activity in apoptotic cell lysates
• Format: Purified Molecular weight: 729 Dalton Storage buffer: Lyophilized powder Purity: ≥98% Functional assay