Proteinase K

Details for Product No. ABIN6383959, Supplier: Log in to see
Antigen
  • proteinase K
  • MCYG_04955
  • MGYG_07924
Reactivity
Tritirachium album
Host
Yeast (Pichia pastoris)
Antibody Type
Recombinant
Biological Activity
Active
Application
DNA Isolation (DNAIso), Endotoxin Removal (ER), Immunohistochemistry (IHC)
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Purpose Proteinase K is commonly used in molecular biology to remove protein contamination.
Specificity DNase is not detected in quality control procedure of incubation 40 µg Proteinase K with 1 µg λ DNA for 6 hours at 37°C.
RNase is not detected in quality control procedure of incubation 40 µg Proteinase K with 2 µg RNA for 2 hours at 37°C. This preparation is considered as RNase and DNase free.
Each lot was tested to ensure the absence of Nucleases and DNA.
Characteristics Molecular Biology Grade, Recombinant, from yeast cells with cloned gene encoding genetically engineered Engyodontium album (Tritirachium album) endolytic protease. The recombinant enzyme is a mutant to the native protease, which gains higher specific activity and yield as well as wider pH and temperature range . The large scale recombinant preparation has advantage in lot-to-lot consistency, superior purity and cost-efficiency. DNA-free nature of recombinant Proteinase K made it well-suited in isolating PCR and RT-PCR templates.
Purification chromatography
Grade Molecular Biology Grade
Biological Activity Comment Specific activity: ≥ 34 units/mg of protein
Unit Definition One unit is defined as the enzyme activity that produce1 µmol of tyrosine per minute from casein at 37°C at pH 7.5. Refer to the certificate of analysis for specific values for the present lot.
Antigen
Background Proteinase K is a broad-spectrum serine protease originally isolated from fungus Engyodontium album. The protease was named “Proteinase K” for its ability to digest Keratin. Crystal and molecular structure studies suggest that the enzyme belongs to the subtilisin family characterized with a catalytic triad (Asp39-His69-Ser224) in active site. Proteinase K has no pronounced cleavage specificity and preferential cleavage site is the peptide bond adjacent hydrophobic amino acids.
Molecular Weight 29.3 kDa
CAS-No 39450-01-6
Application Notes Nucleic Acids Isolation, Immunohistochemistry, Endotoxin Removal, Nucleases Removal, Protein Contamination Removal
Comment

Inhibitor: The enzyme is inactivated by DIFP or PMSF (PMSF used at final concentration 5 mM. ). However, it is not inhibited by EDTA , iodoacetic acid, trypsin-specific inhibitor TLCK, chymotrypsin-specific inhibitor TPCK, and p-chloromercuribenzoate.
Activator: To stimulate proteinase K activity, 1-5 mM Ca2+ can be added. Optimization using activators can increase proteinase activity significantly. Enzyme activity will be reduced by 25% when calcium is removed by addition of EDTA. Enzyme activity will be reduced by 80% if the EDTA-Ca2+ complex is removed from the enzyme solution by gel filtration, while it can be partially restored by addition of excess Ca2+.

Protocol Proteinase K is commonly used in molecular biology to remove protein contamination from preparations of highly native undamaged nucleic acid because it rapidly and effectively inactives nuclease that might degrade the DNA or RNA even in the presence of denaturing reagents.
Proteinase K is active in 1% Tritonä X-100 and fully active in 0.5% (w/v) SDS which denatures protein substrates to increase digestion rates. The enzyme works best at 50-200 mg/mL at pH 7.5-8.0, 37 °C and is usually denatured by subsequent phenol extractions. Incubation times vary from 30 minutes to 18 hours and proteinase K can auto-digest during long incubation.
Restrictions For Research Use only
Format Liquid
Concentration 20 mg/mL
Buffer 20 mM Tris-HCl, 1 mM CaCl2, 50 % Glycerol, pH 7.4
Storage 4 °C
Storage Comment 4°C
Supplier Images
 image for Proteinase K (ABIN6383959) Shelf life of proteinase K solution: Proteinase K solution stored 26 months at temper...
Enzyme Activity Assay (EAA) image for Proteinase K (ABIN6383959) Enzyme Activity Assay: One unit is defined as the enzyme activity that produce1 µmol ...
Agarose Gel Electrophoresis (AGE) image for Proteinase K (ABIN6383959) DNA and RNA contamination Assay: A sample of 5ul solution in 20mg/ml is analyzed with...
Agarose Gel Electrophoresis (AGE) image for Proteinase K (ABIN6383959) DNase contamination Assay: Quality control procedure of incubation 40 µg Proteinase K...
Agarose Gel Electrophoresis (AGE) image for Proteinase K (ABIN6383959) RNase contamination Assay: Quality control procedure of incubation 40 µg Proteinase K...