Description

• General description: Collagenase from Clostridium histolyticum comprises of collagenases G and H. Collagenase has N-terminal activator domain and C-terminal peptidase domain. The collagen binding domains (CBD) and polycystic kidney disease-like (PKD-like) domain resides in the peptidase domain.[1] The collagenase module undergoes structural changes upon calcium binding.[2]
• Application: Collagenase from Clostridium histolyticum has been used:for the recovery of neutrophils from collagen gels[3]for the dissociation of breast tumor biopsies into single cells[4]for the isolation of primary human endometrial stromal cells(HESC) from endometrial biopsies[5]
• Biochem/physiol Actions: Crude collagenases are mixtures of enzymes (mostly proteases) secreted by C. histolyticum. Effective release of cells from tissue requires the action of both collagenase enzymes and the neutral protease. Collagenase is activated by four gram atom calcium (Ca2+) per mole enzyme. The culture filtrate is thought to contain at least 7 different proteases ranging in molecular weight from 68-130 kDa.
• Packaging: Product sold on the basis of collagen digestion units (CDU).
• Unit Definition: One collagen digestion unit (CDU) liberates peptides from collagen from bovine achilles tendon equivalent in ninhydrin color to 1.0 μmole of leucine in 5 hours at pH 7.4 at 37 °C in the presence of calcium ions. One FALGPA hydrolysis unit hydrolyzes 1.0 μmole of furylacryloyl-Leu-Gly-Pro-Ala per min at 25°C. One Neutral Protease unit hydrolyzes casein to produce color equivalent to 1.0 μmole of tyrosine per 5 hr at pH 7.5 at 37°C. One Clostripain Unit hydrolyzes 1.0 μmole of BAEE per min at pH 7.6 at 25°C in the presence of DTT.
• Preparation Note: Collagenase may be dissolved in TESCA buffer (50 mM TES, 0.36 mM Calcium chloride, pH 7.4 at 37 °C) at 0.05-0.1 mg/mL concentration.