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P5147 Sigma

Protease from Streptomyces griseus

Type XIV, ≥3.5 units/mg solid, powder

Synonym: Actinase E, Pronase E

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Description

Frequently Asked Questions

Frequently Asked Questions are available for this Product.

Features and Benefits

• highly stable in pH range of 5.0 to 9.0, with peak activity at pH 8.8
• compatible with many DNA and RNA isolation buffers
• broad substrate specificity

Packaging

1, 5 g in glass bottle

100 mg in glass bottle

Preparation Note

Collected from culture broth of S. griseus.

Quality

Contains calcium acetate.

Specificity

A mixture of at least three proteolytic activities including an extracellular serine protease. In general, serine proteases display a wide range of substrate specificities, which are believed to be mediated by an active site composed of one Asp, one His, and a Ser residue in the molecule. This enzyme prefers to hydrolyze peptide bonds on the carboxyl side of glutamic or aspartic acid.

Unit Definition

One unit will hydrolyze casein to produce color equivalent to 1.0 μmole (181 μg) of tyrosine per min at pH 7.5 at 37 °C (color by Folin-Ciocalteu reagent).

Application

Protease is an enzyme used to break down proteins by hydrolyzing peptide bonds. Protease is used to degrade proteins, to study protease inhibitors and to study thermal inactivation kinetics. Protease is used in nucleic acid isolation procedures in incubations. Protease from Streptomyces griseus has been used in crystallographic and kinetic investigations of the covalent complex formed by tetrapeptide aldehydes and serine proteases.

Protease is typically used in nucleic acid isolation procedures in incubations of 0.5-3.0 hours supplemented with 0.2% sodium dodecyl sulfate and 10 mM EDTA.

The enzyme from Sigma has been used for the digestion and analysis of antithrombin-heparin complexes. It has also been used for the isolation of enzyme-resistant starch.

This enzyme is more active at a higher pH range than the known alkaline protease, showing the proteolytic activity even in 0.2N NaOH solution. This enzyme is useful for proteolysis of insoluble protein and for structure investigation of protein.

Biochem/physiol Actions

Protease catabolizes proteins by hydrolysis of peptide bonds. Proteases are inactivated by serine active-site inhibitors, such as phenylmethylsulfonyl fluoride (PMSF) and diisopropylfluorophosphate. Protease from Streptomyces griseus is a mixture of at least three proteolytic activities including an extracellular serine protease. Serine proteases display a wide range of substrate specificities, which are believed to be mediated by an active site composed of one Asp, one His, and a Ser residue in the molecule. This enzyme prefers to hydrolyze peptide bonds on the carboxyl side of glutamic or aspartic acid.

This product is a mixture of at least three caseinolytic activities and one aminopeptidase activity. The caseinolytic enzymes were named as Streptomyces griseus Protease A, Streptomyces griseus Protease B and Streptomyces griseus Trypsin. This product may be used when extensive or complete degradation of protein is required. This protease mixture is highly nonspecific and can digest casein to the extent of >70% as mono-amino acids.

Physical properties

Completely inactivated by heating above 80 °C for 15-20 minutes.

Protocols & Applications

Non-specific Protease Activity Assay Video

Price and Availability


Amplified Detection

Biomedical Applications
Safety & Documentation

Safety Information

Symbol 
Signal word 
Danger
Hazard statements 
Precautionary statements 
RIDADR 
NONH for all modes of transport
WGK Germany 
2
RTECS 
UK9595000

Frequently Asked Questions

Is Product P5147, Protease from Streptomyces griseus, also described as "Pronase"?
"Pronase E" is the name given to a group of proteolytic enzymes produced by Streptomyces griseus K-1. At least 10 proteases are in the mixture, five serine-type proteases, two zinc endopeptidases, two zinc leucine aminopeptidases, and one zinc carboxypeptidase. To view this information on the P5147 data sheet click on the following link:  http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/1/p5147pis.Par.0001.File.tmp/p5147pis.pdf
Does Product P5147, Protease from Streptomyces griseus, contain calcium acetate?
This product contains calcium acetate buffer salt. The amount of calcium acetate present will vary for each lot number and has been observed to range from 2 - 30% (w/w).
How should Product P5147, Protease from Streptomyces griseus, be reconstituted and how should a stock solution be prepared?
This product can be dissolved in 0.01 M sodium acetate with 0.005 M calcium acetate at pH 7.5 at 37°C; at 0.2 mg/mL, this yields a clear solution which ranges from colorless to light tan. Calcium ion is recommended for protection from autolysis. The activity of a dilute enzyme solution containing 0.01 to 0.1 M calcium ion was stable over 24 hours at neutral pH at 2-8°C.  Pronase E is stable at 4°C for at least six months.  Stock solutions of 5 to 20 mg/mL in water are usually stored at -20°C. To view this information on the P5147 data sheet click on the following link: http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/1/p5147pis.Par.0001.File.tmp/p5147pis.pdf
Under what conditions can Product P5147, Protease from Streptomyces griseus, be used to digest protein?
For DNA isolation, Pronase E is usually prepared as a stock solution, and prior to storage at -20°C, the solution is first heated to 56°C for about 15 minutes, then incubated at 37°C for 1 hour. This encourages self-digestion, to eliminate DNAse and RNAse contamination. The enzyme is added to a DNA sample (in the presence of 0.5-1%(w/v) SDS to disrupt DNA protein interactions) typically at 250-500 μg protein/mL, then incubated at 37°C for 1-4 hours. For protein hydrolysis, dissolve about 0.2 umol of protein in 0.2 mL of 50 mM ammonium bicarbonate buffer at pH 8 (or phosphate buffer pH 7). Add Pronase to 1% (w/w) and incubate at 37°C for 24 hours. It may be necessary to add aminopeptidase M at 4% (w/w) and incubate at 37°C for another 18 hours. To view this information on the P5147 data sheet click on the following link: http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Product_Information_Sheet/1/p5147pis.Par.0001.File.tmp/p5147pis.pdf
How is the activity of Product P5147, Protease from Streptomyces griseus, measured?
The method of enzymatic analysis which is utilized by Sigma to measure the activity of this protease can be viewed by clicking on the following link:  http://www.sigmaaldrich.com/etc/medialib/docs/Sigma/Enzyme_Assay/p5147enz.Par.0001.File.tmp/p5147enz.pdf
Which document(s) contains shelf-life or expiration date information for a given product?
If available for a given product, the recommended re-test date or the expiration date can be found on the Certificate of Analysis. These documents are located on the product detail page under Useful Links & Tools. Click on the following link to search for a Certificate of Analysis. Please click the following link to see the details on our Product Dating Information.
How do I get lot-specific information or a Certificate of Analysis?
A Certificate of Analysis is available by lot number and can be obtained through our Advanced Search Option: http://www.sigmaaldrich.com/catalog/AdvancedSearchPage.do
How do I find price and availability?
There are several ways to find pricing and availability for our products.  Once you log onto our website, you will find the price and availability displayed on the product detail page. You can contact any of our Customer Sales and Service offices to receive a quote.  USA customers:  1-800-325-3010 or view local office numbers. 
What is the Department of Transportation shipping information for this product?
Transportation information can be found in Section 14 of the product's (M)SDS. To access the shipping information for this material, use the link on the product detail page for the product, or search here. 
My question is not addressed here, how can I contact Technical Service for assistance?
Use the option to the right to "Ask a Question" by email of a Technical Service Scientist.
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Protocols & Articles

Protocols

Enzymatic Assay of Protease Using Casein As a Substrate

To standardize a procedure for the enzymatic assay of Protease using Casein as a substrate at Sigma-Aldrich St. Louis.

Peer-Reviewed Papers
15

References

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