Molecular Biology Grade Proteinase K Solution N9012

Proteinase K Solution

Molecular Biology Grade,For Research Use Only

Cat No. N9012

Components

Storage

-20ºC recommended. The shelf life of proteinase K solution is one year when stored sealed below 4ºC. Proteinase K solution is shipped on Bio ice packs or at room temperature.

Description

Proteinase K is an endolytic protease that cleaves peptide bonds at the carboxylic sides of aliphatic, aromatic or hydrophobic amino acids. The Proteinase K is classified as a serine protease. The smallest peptide to be hydrolyzed by this enzyme is a tetrapeptide.

Features

• Ready-to-use solution

• Recombinant proteinase K

• Active in a wide range of reaction products

Applications

• Isolation of genomic DNA from cultured cells and tissues

• Removal of DNases and RNases when isolating DNA and RNA from tissues or cell lines

• Determination of enzyme localization

• Improving cloning efficiency of PCR products

Quality Control

DNase Activity: None detectable enzyme activity after 6 hrs incubation with λ DNA at 37ºC.

RNase Activity: None detectable ribonuclease activity after 16 hrs incubation with RNA at 25ºC.

Source

From yeast cells with cloned gene encoding genetically engineered Engyodontium album (Tritirachium album) endolytic protease.

Molecular Weight

29.3 kDa monomer.

Definition of Activity Unit

One unit of the enzyme liberates Folin-positive amino acids and peptides corresponding to 1 µmol tyrosine in 1 min at 37°C , pH 7.5 using denatured hemoglobin as substrate.

Enzyme activity is assayed in the following mixture: 0.08 M potassium phosphate (pH 7.5), 5 M urea, 4 mM NaCl, 3 mM CaCl₂ and 16.7 mg/ml hemoglobin.

Storage Buffer

The enzyme is supplied in: 50 mM Tris-HCl (pH 7.5), containing Ca²⁺ and glycerol as stabilizers.

Inhibition and Inactivation

Inhibitors: Proteinase K is not inactivated by metal chelators, by thiol-reactive reagents or by specific trypsin and chymotrypsin inhibitors. Phenylmethylsulfonyl fluoride and diisopropyl phosphorofluoridate completely inhibit the enzyme.

Inactivated by heating at 95°C for 10 minutes.

Note

• Optimum activity at 50-55°C.

• Rapid denaturation of enzyme occurs at temperatures above 65°C.

• The recommended working concentration for Proteinase K is 0.05-1 mg/ml. The activity of the enzyme is stimulated by 0.2-1% SDS or by 1-4 M urea.

• Ca²⁺ protects Proteinase K against autolysis, increases the thermal stability and has a regulatory function for the substrate binding site of Proteinase K.

• Stable over a wide pH range: 4.0-12.5, optimum pH 7.5-8.0.

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