Product Class: Other

Proteinase K, Molecular Biology Grade
95 Heat

Product Introduction

Proteinase K is a subtilisin-related serine protease that hydrolyzes a variety of peptide bonds and is frequently used to cleanup enzymatic reactions or cell lysates.

  • Highly characterized for more consistent performance
  • Optimal activity and stability for up to 24 months
  • Active in a wide range of temperatures and buffers with optimal activity between 20 and 60°C and a pH between 7.5 and 12.0.
  • No detectable endonuclease, exonuclease, DNase or RNase contaminating activities
 
Catalog # Size Concentration
P8107S 2.0 ml 800 units/ml

Product Information

Description

Highly characterized for more consistent performance, Proteinase K is a subtilisin-related serine protease that will hydrolyze a variety of peptide bonds. Proteinase K is active in a wide range of temperatures and buffers with optimal activity between 20 and 60°C and a pH between 7.5 and 12.0 (1, 2). Activity is stimulated when up to 2% SDS or up to 4 M urea are included in the reaction (3). Calcium is important for thermostability of Proteinase K but it is not required for catalysis, therefore Proteinase K is also active in buffers containing chelating agents such as EDTA (4).

Lot-to-Lot Variability: Competitor ComparisonLot-to-Lot Variability: Competitor Comparison

Activity measurements from three NEB lots of Proteinase K were compared to three lots from another vendor. Reported activity for all NEB lots is 800 Units/ml ± 10% (indicated as a black line with error bars). Direct measurement of the competitor’s material using an assay reported on the vendor’s literature, demonstrated a dramatic over-assessment of all three lots.

Activity Measurements: Competitor ComparisonActivity Measurements: Competitor Comparison

Activity measurements from a single NEB lot were compared to a single lot from seven other vendors. Proteinase K from NEB is sold at 800 Units/ml ± 10% (indicated as a black line with error bars), whereas most other vendors sell Proteinase K as > 800 or > 600 Units/ml. While Proteinase K from some vendors was close to the stated activity values, others varied significantly from the expected amount.

Protein Concentration
20 mg/ml, approximately, as determined by UV absorption at 280 nm.

Product Source

Engyodontium album (Tritirachium album)
This product is related to the following categories:
Proteases Products,
Nucleic Acid Purification Products,

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration

Properties & Usage

Unit Definition

One unit will digest urea-denatured hemoglobin at 37°C (pH 7.5) per minute to produce equal absorbance as 1.0 μmol of L-tyrosine using Folin & Ciocalteu's phenol reagent (6).

Storage Buffer

20 mM Tris-HCl
1 mM CaCl2
50% Glycerol
pH 7.4 @ 25°C

Heat Inactivation

95°C for 10 minutes

Molecular Weight

Theoretical: 28.9 kDa

Unit Assay Conditions

0.5–2 μg of Proteinase K is incubated with 2% denatured hemoglobin solution for 10 minutes at 37°C (pH 7.5). After precipitation, neutralization and addition of Folin & Ciocalteu's phenol reagent, absorbance of soluble cleavage products are measured at 750 nm. Absorbance is compared to a standard curve of L-tyrosine absorbance prepared similarly.

Application Features

  • Isolation of plasmid and genomic DNA 
  • Isolation of RNA
  • Inactivation of RNases, DNases and enzymes in reactions
  • Removal of enzymes from DNA to improve cloning efficiency (5)
  • PCR purification

Product Notes

  1. Active in a wide range of buffers, including all NEB-specific restriction endonuclease buffers. It is highly active between pH 7.5 and 12 and temperatures 20-60°C. Proteinase K is also active in chelating agents such as EDTA and activity is stimulated in up to 2% SDS or 4M urea.
  2. Proteinase K is stable for at least 2 years at –20°C. No loss of activity is observed after 10 freeze-thaw cycles.

  3. Reaction Conditions
    Proteinase K is active in a wide range of buffers including all NEB specific restriction endonuclease buffers. It is highly active between pH 7.5 and 12.0 and temperatures between 20 and 60°C (1,2). Proteinase K is also active in chelating agents such as EDTA (4) and activity is stimulated in up to 2% SDS or 4 M urea (3).

References

  1. Bajorath, J. et al. (1988). Biochimica et Biophysica Acta. 954, 176-182.
  2. Ebeling, W. et al. (1974). Eur. J. Biochem. 47, 91-97.
  3. Hilz, H. et al. (1975). Eur. J. Biochem. 56, 103-108.
  4. Bajorath, J. et al. (1988). Eur. J. Biochem. 176, 441-447.
  5. Crowe, J.S. et al, (1991). Nucleic Acids Research. 91, 184.
  6. Anson, M.L. (1939). J. Gen. Physiol. 22, 79-89.
  7. Pace, C.N. et al. (1995). Protein Sci. 4, 2411-2423.

Protocols, Manuals & Usage

Protocols

  1. General protocol to release nucleic acids prior to capillary or gel electrophoresis using Proteinase K (P8107)
  2. Protocol to cleanup DNA Glucosylation/restriction digest and Proteinase using Proteinase K (P8107)
  3. Protocol to purify PCR products in preparation for cloning using Proteinase K (P8107)
  4. Using recombinant Cas9 nuclease to assess locus modification in genome editing experiments (#M0386)

FAQs & Troubleshooting

FAQs

  1. The protocol I am using requires a µg/ml concentration of Proteinase K. What is the protein concentration of Proteinase K, Molecular Biology Grade?
  2. What buffer should I use for Proteinase K?
  3. If I need to dilute Proteinase K, Molecular Biology Grade, how should I do this?
  4. Why is the unit definition assay for Thermolabile Proteinase K (NEB #P8111) different from the unit definition assay for Proteinase K, Molecular Biology Grade (NEB # P8107)?
  5. What is the activity if Proteinase K, Molecular Biology Grade if subjected to the unit definition assay used for Thermolabile Proteinase K?

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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