Proteinase K, RNase/DNase free

Proteinase K is a highly reactive nonspecific serine protease that belongs to the subtilisin family of proteins. It cleaves at the carboxylic acid side of aliphatic, aromatic, or hydrophobic amino acids. Proteinase K is capable of inactivating RNases and DNases and is used in the isolation or preparation of high molecular weight nucleic acids. Proteinase K is also useful for helping to characterize enzymes, due to its cleavage specificity. This enzyme was designated proteinase K because of its ability to hydrolyze keratin. Proteinase K is stable in a wide variety of detergents and buffer salts and at various temperatures and pH. The isoelectric point of proteinase K is 8.9.

Proteinase K is typically used at a concentration of 50-100 μg/ml. It is active with or without the presence of SDS, urea, EDTA or various metal ions, but the activity of proteinase K can be increased by adding the denaturing agents and the structure of proteinase K can be stabilized by addition of Ca ²⁺. Proteinase K is inactivated by heating to 95°C for 10 minutes or using an inhibitor such as PMSF, AEBSF or DFP.

GoldBio's Proteinase K undergoes two sterile filtration steps during the separation and purification processes. The final product is further analyzed by plate culture to ensure the removal of all living organisms was 100% effective.

Unit Definition: One unit is defined as the amount of enzyme that will liberate 1.0 μmol tyrosine (Folin-positive amino acid) from casein per minute at 37°C, pH 7.5.

Applications

• Digestion of unwanted proteins
• Removal of endotoxins bound to cationic proteins such as lysozyme and RNaseA
• Removal of nucleases for in situ hybridization
• Prion research with respect to TSE (transmissible spongiform encephalopathies)
• Protease footprinting Mitochontrial isolation
• Isolation of genomic DNA
• Isolation of cytoplasmic RNA
• Isolation of highly native DNA or RNA

Storage/Handling

Store at -20°C.
Reconstitute in 50mM Tris-HCl (pH 8.0), 3mM CaCl₂.

Proteinase K Quick Answers:

How do you inactivate proteinase K?

Inactivate proteinase K by heating it to 95˚C for 10 minutes. You can also permanently inactivate proteinase K by using protease inhibitors such as PMSF and AEBSF (Pefabloc®).

What is the best temperature for proteinase K activation?

The best temperature for proteinase K activity is between 50-65˚C. Higher temperatures help with protein unfolding, which makes it easier for proteinase K to breakdown proteins.

How do you store proteinase K?

Aliquot proteinase K stock solution and store at -20˚C for up to 1 year.

Store lyophilized proteinase K powder desiccated at -20˚C for up to 2 years.

How do you dissolve proteinase K?

Proteinase K is water soluble and it can also be dissolved in Tris or PBS. Dissolving proteinase K with PBS is a little challenging, which may be due to pH. It's best to add proteinase K a little bit at a time while mixing into solution.

What is the optimal pH when working with proteinase K?

The optimal pH when working with proteinase K is between 7.5 - 12.0