Proteinase K Recombinant

Properties

Quality level: 100,

Form

buffered aqueous solution (18 ± mg / mL; pH 7.5)

Specific activity

~ 2.5 units/mg protein (Approximately 2.5 U / mg (Chromozym assay); haemoglobin assay ≥30 U / mg. See Certificate of Analysis for specific values ​​for this lot).

Packaging

  • 1.25 mL package (03115887001)
  • 25 mL package (03115844001)
  • 5 mL package (03115828001)

Manufacturer / trade name: Roche

Optimum pH: 6.5 and 9.5

pH range: 4.0 – 12.5

Sent in: wet ice

Storage temperature: 2-8 ° C

General description

Proteinase K is a subtilisin-related serine protease that does not have a pronounced cleavage specificity. The enzyme is also available as a lyophilisate. The recombinant enzyme is identical to the native protease originally isolated from the Tritirachium album fungus.

The specifications of the recombinant enzyme are the same as those of the native protease. The amino acid sequence (molecular weight) and the structure of the molecule are identical. However, the recombinant preparation is much purer than the native enzyme, as it is free of DNA, according to current quality control procedures, and therefore very suitable for isolating PCR and RT-PCR templates.

Application

  • This PCR grade proteinase K is extremely efficient on native proteins and therefore can be used to rapidly inactivate endogenous nucleases such as RNases and DNases. This property makes Proteinase K particularly suitable for the isolation of native RNA and DNA from tissues or cell lines.
  • The enzyme promotes cell lysis by activating a bacterial autolytic factor.
  • Proteinase K is also used for the analysis of membrane structures by modifying proteins and glycoproteins on the cell surface.
  • The enzyme is particularly suitable for isolating nucleic acids for amplification reactions.
  • Proteinase K can be used to remove cell debris during colony lift preparation and to treat tissue sections to ensure efficient probe infiltration during in situ hybridization.

Features and Benefits

Proteinase K cleaves proteins between amino acids X and Y (X- ↓ -Y) when X = an aliphatic, aromatic, or hydrophobic amino acid, and Y = any amino acid. The enzyme is extremely effective on native proteins and therefore can be used to rapidly inactivate endogenous nucleases such as RNases and DNases.

  • Choose an effective template preparation tool. Inactivate DNases and RNases of most species.
  • Count on consistent quality and performance. Stringent quality tests ensure optimal stability and high-level batch-to-batch performance.
  • Prepare samples under a wide range of conditions. The robust enzyme is stable over a wide pH range and is ideal for various applications.
  • Benefit from a contamination-free enzyme. The enzyme is tested for the absence of RNases and DNases and is virtually DNA-free. It is especially suitable for the isolation of PCR templates.

Quality

This preparation is free of RNases, DNases and DNA, according to current quality control procedures.

  • Nuclease-free – Each lot is tested on multiple substrates to ensure the absence of endonucleases, exonucleases, ribonucleases, and cleavage activity.
  • DNA content: ≤10 pg/mg enzyme (determined by threshold)
  • Biological load: ≤5 CFU / g (determined by the strictest European Pharmacopoeia test, which identifies the total number of viable aerobic bacteria, yeast and fungi)

Definition of unit

  • Volume activity: Approximately 50 U / ml solution (chromosome assay); approximately 600 U / ml solution (hemoglobin test). One unit is the enzyme activity that cleaves at +25 ° C in 1 min 18 mmol of Chromozym TRY (equivalent to 600 U / ml with the haemoglobin assay). See Certificate of Analysis for current lot-specific values.

Preparation note

  • Activator: to stimulate proteinase K activity, denaturing agents (SDS and urea) can be added. For example, SDS at a final concentration of 2% can significantly increase proteinase K activity. Optimization using denaturing agents can increase proteinase activity up to sevenfold.
  • Working Solution: Suggested Buffers: The most appropriate buffer for proteinase K will vary from application to application. Always follow the pH and temperature guidelines in the parameter file. As a general rule, proteinase K is stable and highly active in buffers containing denaturing reagents such as urea, sodium dodecyl sulfate (SDS), and guanidinium salts.
  • Inhibitors: Pefabloc® SC inactivates the enzyme. However, it is not inactivated by metal ions, chelating agents (eg, EDTA), sulfhydryl reagents, or trypsin and chymotrypsin inhibitors.

Other notes

  1. For the rapid inactivation of RNases and DNases.
  2. The enzyme can reduce protein to free amino acids if it is present in large excess during long incubation periods.
  3. For life science research only. It should not be used in diagnostic procedures.

Legal information

Pefabloc is a registered trademark of Pentapharm

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