Product Class: Other

Endoproteinase GluC
NEBU cloned at NEB recombinant 37

P8100_cutsite

Product Introduction

Endoproteinase GluC is a serine proteinase which preferentially cleaves peptide bonds C-terminal to glutamic acid residues. It also cleaves at aspartic acid residues at a rate 100-300 times slower than at glutamic acid residues.

  • Ideal for proteome analysis
  • Recombinant enzyme with no detectable non-specific protease contaminating activities
  • Free of glycerol and detergents which may interfere with Mass Spectrometry or liquid chromatography methods.
  • Supplied in dry format
  • Optimal activity and stability for up to 12 months
Catalog # Size Concentration
P8100S 50.0 µg

Product Information

Description

Endoproteinase GluC (Staphylococcus aureus Protease V8) is a serine proteinase which selectively cleaves peptide bonds C-terminal to glutamic acid residues (1). Endoproteinase GluC also cleaves at aspartic acid residues at a rate 100-300 times slower than at glutamic acid residues (2,3).

MALDI-TOF Ms
A 16 hour GluC digestion of Equine heart cytochrome c using a 1:20 enzyme to substrate ratio, followed by LC-MS with a Thermo™ LTQ Orbitrap XL, identified 97 peptides with 77% sequence coverage.

Product Source

Staphylococcus aureus Protease V8 gene cloned and expressed with histidine-tag in Bacillus subtilis

Reconstitution

Endoproteinase GluC should be reconstituted by the addition of 50-500 µl of high purity water. Finger flick the volume of water in the tube to fully resuspend the enzyme. Rapid autolysis is a function of enzyme concentration; any sample reconstituted in a small volume should be used immediately. To get the most use out the enzyme, resuspend the enzyme in 500 μl H2O and aliquot 50 ul each in 10 tubes. Freeze the tubes that are not being used immediately at -20°C for up to two weeks or less. Storage at - 80°C will prolong enzyme stability approximately 2-4 additional weeks.
This product is related to the following categories:
Proteases Products,
Proteome Analysis Products,
This product can be used in the following applications:
Glycomics and glycoproteomics,
Protein Digestion

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • P8100S     -20    
      Endoproteinase GluC P8100SVIAL -20 1 x 50 µg Not Applicable
      GluC Reaction Buffer B8100SVIAL -20 1 x 1.5 ml 2 X

Properties & Usage

Reaction Conditions

1X GluC Reaction Buffer
Incubate at 37°C

2X GluC Reaction Buffer
50 mM Tris-HCl
0.5 mM Glu-Glu
(pH 8 @ 25°C)

Usage Concentration

100 ng/μl

Molecular Weight

Theoretical: 29849 daltons

Product Notes

  1. Storage conditions: Supplied in dry format from Tris-HCl and sodium chloride buffer.
  2. Repeated freeze/thawing is not recommended.
  3. GluC may also be used in 20-50mM phosphate or ammonium bicarbonate buffer. For optimal activity, supplement reactions with Glu-Glu dipeptide. Glutamic acid residues are highly favored in Tris, phosphate and ammonium bicarbonate buffers.
  4. GluC is purified with a His-tag at its C-terminal of the protein. The predominant species has four histidine residues at the C-terminus with the remaining species showing a distribution from one to six His tags.

References

  1. Drapeau, G.R., Boily, Y. and Houmard, J. (1972). Purification and properties of an extracellular protease of Staphylococcus aureus. J. Biol. Chem.. 247, 6720-26.
  2. Benner, J.S., Martin, D. and Schampel, A.K. unpublished results.
  3. Birktoft, J.J. and Breddam, K. (1994). Proteolytic Enzymes: Serine and Cysteine Peptidases. A.J. Barrett(Ed.), Glutamyl endopeptidases. Methods Enzymol.. 244, 114-126. San Diego: Academic Press.
  4. Breddam, K. and Meldal, M. (1992). Substrate preference of glutamic-acid-specific endopeptidases assessed by synthetic peptide substrates based on intramolecular fluorescence quenching. Eur. J. Biochem.. 206, 103-7.

Protocols, Manuals & Usage

Protocols

  1. Typical GluC Digest Reaction Conditions (P8100)
  2. Typical Trypsin and GluC Co-Digest Reaction Conditions (P8100)
  3. Typical GluC In-Gel Digest Reaction Conditions (P8100)

Tools & Resources

Selection Charts

FAQs & Troubleshooting

FAQs

  1. What is the stability/compatibility of Endoproteinase GluC with regards to presence of DTT, urea, concentrations of methanol, acetonitrile, nitrosoguanidine, thiourea and glycerol?
  2. How does one carry out limited proteolysis experiments with Trypsin and Endoproteinase Glu-C? Which chemicals can be used to quench the reactions at given time points to monitor proteolysis progress via SDS- PAGE?

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 [email protected].

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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