Product Name

Proteinase K from Tritirachium album

Synonym

Endopeptidase K

Assay

≥500 units/mL

CAS Number

39450-01-6

Molecular Weight

mol wt 28.93 kDa

MDL number

MFCD00132129

Suitability

BioReagent

l Physical and Chemical Information

Appearance

buffered aqueous glycerol solution

Physical form

Solution in 40% (v/v) glycerol containing 10 mM Tris-HCl, pH 7.5, with 1 mM calcium acetate.

l Biological Information

Biochem/Physiol Actions

Proteinase K is a stable and highly reactive serine protease. Evidence from crystal and molecular structure studies indicates the enzyme belongs to the subtilisin family with an active-site catalytic triad (Asp39-His69-Ser224). It is stable in a broad range of environments: pH, buffer salts, detergents (SDS), and temperature. In the presence of 0.1-0.5% SDS, proteinase K retains activity and will digest a variety of proteins and nucleases in DNA preparations without compromising the integrity of the isolated DNA. Proteinase K has a broad specificity and degrades many proteins even in the native state. It mainly cleaves the peptide bond adjacent to the carboxyl group of aliphatic and aromatic amino acids with blocked alpha-amino groups.The optimum pH is between 7.5-9.0and the isoelectric point is 8.9. Ca₂₊ (1-5 mM) is required for activation. Proteinase K is inhibited by DIFP or PMSF.

Application

Product has been used to break down cardiac muscle during histopathology studies. It has also been used during the digestion of HEK-293 cells.

The enzyme has been used in the digestion of sealed cytosolic side out ER vesicles. It has been used to deproteinize dissected brain and/or whole pupae sections of honey bee prior to in situ hybridisation. This was done during the study of neuropeptide Y-like signaling, and nutritionally-mediated gene  and behaviour in the honey bee.

Useful for the proteolytic inactivation of nucleases during the isolation of DNA and RNA.

Removes endotoxins that bind to cationic proteins such as lysozyme and ribonuclease A.

Reported useful for the isolation of hepatic, yeast, and mung bean mitochondria

Determination of enzyme localization on membranes

Treatment of paraffin embedded tissue sections to expose antigen binding sites for antibody labeling.

Digestion of proteins from brain tissue samples for prions in Transmissible Spongiform Encephalopathies (TSE) research.

Preparation Note

Solution in 40% (v/v) glycerol containing 10 mM Tris-HCl, pH 7.5, with 1 mM calcium acetate. Proteinase K in solution is stable over a pH range of 4.0-12.5 (optimum pH 8.0), and is also stable over the temperature range of 25°C to 65°C during use. At pH 8.0, solutions will be stable for at least 12 months at 4°C. At pH 4-11.5, solutions containing Ca₂₊(1-6 mM) are expected to be stable for several weeks. An 80% ammonium sulfate suspension stored at 4°C is stable for at least 12 months.

Unit Definition

One unit will hydrolyze urea-denatured hemoglobin to produce color equivalent to 1.0 μmole of tyrosine per min at pH 7.5 at 37 °C (color by Folin-Ciocalteu reagent).

l Storage

Storage temp.

2-8°C

l Precautions and Disclaimer

This product is for R&D use only, not for drug, household, or other uses.

l References

1. http://www.drugbank.ca

2. https://ncit.nci.nih.gov

3. https://www.ncbi.nlm.nih.gov