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C8051 Sigma

Collagenase from Clostridium histolyticum

Sigma Blend Type H, ≥1.0 FALGPA units/mg solid

Synonym: Clostridiopeptidase A

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Description

Biochem/physiol Actions

Collagenase is activated by four gram atom calcium per mole enzyme. It is inhibited by ethylene glycol-bis(beta-aminoethyl ether) - N, N, N′,N′-tetraacetic acid, beta-mercaptoethanol, glutathione, thioglycolic acid and 8-hydroxyquinoline.

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. The pH optimum is 6.3-8.8. The enzyme is typically used to digest the connective components in tissue samples to liberate individual cells. Collagenase treatment can cause some cells to die. Typically, concentrations varying from 0.1 to 5 mg/mL is used for digestion. The duration of reaction varyies from 15 minutes to several hours to yield a satisfactory efficiency of cell dissociation, without causing too much cell death. Krebs Ringer buffer with calcium and BSA is preffered and Zn2+ is required for activity. This enzyme is tested for the release of hepatocytes at approximately 1 mg/mL conc., in a total volume of 100 mL for each rat liver.

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.

Application

Clostridiopeptidase A has been used in a study to investigate the synthesis and in vitro evaluation of macromolecular antitumour derivatives based on phenylenediamine mustard. Clostridiopeptidase A has also been used in a study to investigate the strain-controlled enzymatic cleavage of collagen in loaded matrix.

The enzyme has been used for the partial digestion of mouse abdominal muscle feed artery (AFA) during endothelial cell (EC) tube isolation and superfusion. This study investigated the electrical conduction along EC tubes from mouse feed arteries. The product has also been used during the preparation of bovine pulmonary artery endothelial cells. This bovine study investigated the contribution of nitric oxide to 20-hydroxyeicosatetraenoic acid-induced relaxation of pulmonary arteries.

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Laboratory Gloves

Biomedical Applications
Safety & Documentation

Safety Information

Symbol 
Signal word 
Danger
Hazard statements 
Precautionary statements 
Personal Protective Equipment 
RIDADR 
NONH for all modes of transport
WGK Germany 
1
Protocols & Articles

Articles

Collagenase Tissue Digestion/Dissocation Trouble Shooting and References

* The separately prepared collagenase and protease enzymes in the “Sigma Blend” products (Cat. No. C7926, C8051 or C8176) give reproducible control of how much of each is used.
BioFiles 2006, 1.2, 6.
Keywords: Adhesion, Apoptosis, Applications, Buffers, Catalog, Cell biology, Digestions, Methods

Protocols

Enzymatic Assay of Collagenase

The continuous spectrophotometric rate determination (A345, Light path = 1 cm) is based on the following reaction:
Keywords: Extinction coefficient

Related Content

Enzymes & Proteins

Application Index | Enzyme Index | Substrate Index | Inhibitor Index | Cofactor Index | Lectin Index
Keywords: Cell signaling, Diagnostic, Drug discovery, Molecular biology

Peer-Reviewed Papers
15

References

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substrate

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F5135 N-[3-(2-Furyl)acryloyl]-Leu-Gly-Pro-Ala
C0627 Z-Gly-Pro-Leu-Gly-Pro

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