Proteinase K

E.C. 3.4.21.64 Synonym: Endoproteinase K

Isolated from the fungus Engyodontium album (formerly Tritirachium album), Proteinase K exhibits broad substrate specificity and degrades many proteins in the native state, even in the presence of detergents.2,3,4 Proteinase K is a stable S8 family serine alkaline protease containing two disulfide bridges and one free Cys near His at the active site.24 The predominant site of cleavage is the peptide bond adjacent to the carboxyl group of aliphatic and aromatic amino acids with blocked alpha amino groups.

 

Physical properties of Proteinase K

Molecular weight: 28,930 Da (amino acid sequence)21; 28,500 Da (SDS-PAGE)22
pH range: 7.5 to 12.02,3
Temperature profile: Maximum activity at 37 °C3
pI: 8.92
Extinction coefficient: E1% = 14.2 (280 nm, 10 mM NaCl and 5 mM CaCl2, pH 8.0)2
Unit definition: One unit will hydrolyze urea-denatured hemoglobin to produce color equivalent to 1.0 mmol (181 mg) of tyrosine per minute at pH 7.5 at 37 °C.
Activators: 1-5 mM Ca2+ is required for activation. Proteinase K is active in 1% TRITON™ X-100 and fully active in 0.5% (w/v) SDS.3,19,20,23
Inhibitors: Proteinase K is inhibited by DIFP or PMSF. Proteinase K is partly inactivated, but not inhibited, by EDTA
Solubility: 1 mg/mL (aqueous)
Storage: Store lyophilized powders at –20 °C. Store solutions at 4 °C.
Stability: Proteinase K solutions are stable over a broad pH range (4.0-12.5, optimum pH 8.0) and temperature range (25 to 65 °C). Lyophilized powders are stable for 2 years when stored at –20 °C. pH 8.0 proteinase K solutions are stable for at least 12 months at 4 °C3

 

Proteinase K Applications

  • Mitochondria isolation
  • Protein digestion for nucleic acid (DNA and RNA) purification3, 5-11
  • Removal of endotoxins bound to cationic proteins, such as lysozyme and ribonuclease A 12
  • Determination of enzyme localization on membranes 13
  • Exposure of antigen binding sites on paraffin embedded tissue sections for antibody labeling14
  • Removal nucleases for in situ hybridization15
  • Digestion of proteins from brain tissue samples16,17
  • Protease footprinting to reveal protein-protein surface interactions18

Proteinase K Products

Product No.
Product Name Add to Cart
70663 Proteinase K from Tritirachium album, lyophilized powder, ≥30 units/mg

P2308 Proteinase K from Tritirachium album, lyophilized powder, BioUltra, ≥30 units/mg

SRE0005 Proteinase K from Tritirachium album, cGMP, suitable for use in an IVD application
P6556 Proteinase K from Tritirachium album, lyophilized powder, ≥30 units/mg
P4850 Proteinase K from Tritirachium album, lyophilized powder, 3.0-15.0 units/mg
P5568 Proteinase K from Tritirachium album, buffered aqueous glycerol solution, ≥500 units/mL
SAE0009 Proteinase K from Tritirachium album, ≥30 units/mg
SRE0047 Proteinase K from Tritirachium album, lyophilized powder, ≥30 units/mg, suitable for manufacturing of diagnostic kits and reagents
71049 Proteinase K Solution, 600 mAU/mL

Proteinase K is available in BioUltra Grade. Learn more about our BioUltra grade.

We are a large-scale manufacturer of Proteinase K. We supply custom formulated preparations to diagnostic manufacturing and biotechnology customers. Inquire with your local SAFC representative.

 

References

  1. Betzel, C., Three Dimensional Structure of Proteinase K at 0.15 nm Resolution. Eur. J. Biochem., 178, 155-171 (1988).
  2. Ebeling, W., et al., Proteinase K from Tritirachium album Linder, Eur. J. Biochem., 47, 91 (1974).
  3. Enzymes of Molecular Biology, vol. 16, Burrell, M.M., ed. Humana Press (Totowa, NJ: 1993), p. 307. Kraus, E., and Femfert, U., Proteinase K from the Mold Tritirachium album Limber, Specificity and Mode of Action. Z. Physiol. Chem., 357, 937 (1976).
  4. Lizardi, P.M., and Engelberg, A., Rapid Isolation of RNA Using Proteinase K and Sodium Perchlorate. Anal. Biochem., 98, 116 (1979).
  5. Gross-Bellard, et al., Isolation of High Molecular Weight DNA from Mammalian Cells, Eur. J. Biochem., 36, 32-38 (1973).
  6. Molecular Cloning: A Laboratory Handbook, 2nd ed., Sambrook et al., eds., Cold Spring Harbor Press (Cold Spring Harbor, NY: 1989) p. 1.61 and p. B.16.
  7. Kasche, V., et al., A Two-step Procedure for Quantitative Isolation of Pure Double-strand DNA from Animal Tissues and Cell Cultures. Prep. Biochem., 11, 233 (1981).
  8. Hansen, J.N., Isolation of Higher Molecular Weight DNA from Bacillus cereus T Using Proteinase K. Prep. Biochem., 4, 473 (1974).
  9. Holm, C., et al., A Rapid, Efficient Method for Isolating DNA from Yeast. Gene, 42, 169 (1986).
  10. La Claire, J.W., and Herrin, D.L., Co-isolation of High-Quality DNA and RNA from Coenocytic Green Algae. Plant Mol. Biol. Reporter, 15, 263 (1997).
  11. Petsch, P., et al., Proteinase K Digestion of Proteins Improves Detection of Bacterial Endotoxins by the Limulus Amebocyte Assay: Application for Endotoxin removal from Cationic Proteins. Anal. Biochem., 259, 42 (1998).
  12. Brdiczyka, D., and Krebs, W., Localization of Enzymes by Means of Proteases. Biochem. Biophys. Acta, 297, 203 (1973).
  13. Short, B.G., et al., Automated Double Labeling of Proliferation and Apoptosis in Glutathione S-transferase-positive Hepatocytes in Rats. J. Histochemistry and Cytochemistry, 45, 1299 (1997).
  14. Angerer, L.M., et al., Identification of Tissue-Specific Gene Expression by in-situ Hybridization. Methods in Enzymology, 152, 649 (1987).
  15. Sakaguchi, S., et al., Accumulation of Proteinase K-Resistant Prion Protein (PrP) is Restricted by the Expression Level of Normal PrP in Mice Inoculated with a Mouse-Adapted strain of the Creutzfeldt-Jakob Disease Agent. J. Virology, 69, 7586 (1995).
  16. Bennion, B.J., and Daggett, V., Protein Conformation and Diagnostic Tests: the Prion Protein. Clinical Chemistry, 48, 2105 (2002).
  17. Hori, R., and Carey, M., Protease Footprinting Analysis of Ternary Complex Formation by Human TFIIA. J. Biol. Chem., 272, 1180 (1997).
  18. Hilz, H., et al., Stimulation of Proteinase K action by Denaturing Agents: Application to the Isolation of Nucleic Acids and the degradation of “Masked” Proteins. Eur. J. Biochem., 56, 103 (1975).
  19. Methods of Enzymatic Analysis, 3rd Edition, Bergmeyer, H.U., ed., Academic Press (New York, NY: 1983) vol. 2, p. 299.
  20. Jany, K.D., et al., Amino Acid Sequence of Proteinase K from the Mold, Tritirachium album Linder. Proteinase K; a Subtilisin-related Enzyme with Disulfide Bonds. FEBS Letters, 199, 139 (1986).
  21. Jany, K.D., and Mayer, B., Proteinase K from Tritirachium album linder, Molecular Mass and Sequence Around the Active Serine Residue. Biol. Chem. Hoppe-Seyler, 366, 485 (1985).
  22. Bajorath, J., et al., The Enzymatic Efficiency of Proteinase K is Controlled by Calcium. Eur. J. Biochem., 176, 441-447 (1988).
  23. IUBMB Enzyme Nomenclature: http://www.chem.qmul.ac.uk/iubmb/enzyme/EC3/4/21/64.html

 

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