Proteinase K

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  • Available formats: Lyophilized Powder and Solution

Proteinase K

The Master Protease designed by Nature
For more than 25 years, Merck has manufactured Proteinase K in large quantities and high quality by processing fermenter supernatants of the fungus Tritirachium album. Proteinase K degrades almost every protein and inactivates rapidly enzymatic activities like nucleases under a broad range of conditions, even in the presence of SDS, Urea or at extreme pH. Due to its non-specific proteolytic activity Proteinase K is used as the standard protease.

Inhibitors: The enzyme belongs to the group of serine proteases and is inactivated by sulfonyl fluoride analogs like PMSF or Pefabloc SC. Complexing agents of ions like EDTA do not interfere with the activity of the enzyme but Proteinase K needs calcium ions for long term stability.

Specificity: Proteinase K cleaves peptide bonds with some preference next to the carboxyl group of N-substituted hydrophobic aliphatic and aromatic amino acids. Proteinase K also cleaves peptide amides. The specificity of esterolytic cleavage is high.

Activity: Substrates like hemoglobin demonstrate a very high specific activity. Proteinase K is 6 times more active than Streptomyces protease mixture "Pronase E" and about 3 times more active than bovine Trypsin. A remarkable property of Proteinase K is its ability to rapidly deactivate native proteins by cleavage of accessible peptide bonds. Addition of 0.5-1% of SDS or 1-4 M Urea increase its activity due to denaturation of the native proteins. The pH optimum is between pH 7.5 and 10 (>50% activity at pH 4 or pH11). Increasing the temperature up to 50°C also increases the proteolytic activity.

Inactivation: Proteinase K is inactivated by heat, eg. incubating at >55 °C for 10 minutes, depending on the buffer conditions. Autoproteolytic digestion of the enzyme is induced by depletion of calcium (EGTA) and is further accelerated with the ionic detergents like SDS.

Stability: Lyophilized Proteinase K is extremely stable and a shelf life more than 36 months is guaranteed when stored dry at 4-8°C. The enzyme is soluble at least at 20 mg/ml in water. The ready-to-use solution of the enzyme contains calcium ions as well as glycerol and no loss of activity has been detected after 24 months. Short-term storage at ambient temperature does no harm to the enzyme activity and stability.

Quality: Proteinase K is available in two kinds of formulations, lyophilized and in solution. Both formulations guarantee a long shelf life of the enzyme (>>24months). In the manufacturing process for Proteinase K solution an extra purification step is included to intercept any contaminating nucleic acid. The final product is filtered and packed under laminar air in an aerosol free environment. Proteinase K is quality controlled with highly sensitive tests:

  • Proteolytic activity: degradation of hemoglobin, 37 °C = > 600 U/ml or >35 U/mg
  • Nicking activity: no change of native forms of plasmid DNA after incubation: 16 h at 37 °C
  • RNases: no degradation of flourescent labeled RNA after incubation: 4 h at 37 °C
  • DNA: no host DNA derived PCR amplicon detected
  • BioBurden: no bacterial rDNA derived PCR amplicon detected

© Merck KGaA, Darmstadt, Germany, 2014

All references to Merck refer to Merck KGaA, Darmstadt, Germany.


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