Product Name

Proteinase K from Tritirachium album

Synonym

Endopeptidase K

Assay

3.0-15.0 unit/mg solid; Protein, 15-40% biuret

CAS Number

39450-01-6

Molecular Weight

mol wt 28.93 kDa

MDL number

MFCD00132129

Suitability

BioReagent

l Physical and Chemical Information

Appearance

lyophilized powder

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 is highly active towards native proteins. It 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 α-amino groups. The optimum pH is between 7.5-9.0 and the isoelectric point is 8.9. Ca₂₊(1-5 mM) is required for activation. Proteinase K is inhibited by diisopropyl fluorophosphate (DFIP), and phenylmethanesulfonyl fluoride (PMSF).

Application

Proteinase K is useful for the proteolytic inactivation of nucleases during the isolation of DNA and RNA. It is used for the removal of endotoxins bound to cationic proteins such as lysozyme and ribonuclease A. It is also useful for the isolation of hepatic, yeast, and mung bean mitochondria and is used to determine enzyme localization on membranes. Furthermore, it is used for the treatment of paraffin embedded tissue sections to expose antigen binding sites and for digestion of proteins from brain tissue samples. Product P8044 is provided as a lyophilized powder. The enzyme from Sigma has been used to degrade complex I (NADH:ubiquinone oxidoreductase) prior to extraction of ubiquinone from Yarrowia lipolytica. It has been used to examine the effect of Proteinase k on Pythium ultimum.It has also been used to maximize the variety of peptide bonds hydrolyzed in the sediment slurry without autolytic production of amino acids from the enzyme itself. 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

This product is soluble in water (1–2 mg/ml).

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.

-20°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