Endoproteinase AspN

Description

Endoproteinase AspN (flavastacin) is a zinc metalloendopeptidase which selectively cleaves peptide bonds N-terminal to aspartic acid residues(1).

This enzyme is recommended to digest peptides with molecular weights of 5 kDa or less.

Product Source

Purified from Flavobacterium menigosepticum.

Reconstitution


Endoproteinase AspN should be reconstituted by the addition of 50-500 µl of high purity water. Rapid autolysis is a function of enzyme concentration.

Reagents Supplied

The following reagents are supplied with this product:

Store at (°C)Concentration
Endoproteinase AspN Reaction Buffer-202X

Properties and Usage

Reaction Conditions

1X Endoproteinase AspN Reaction Buffer
Incubate at 37°C

1X Endoproteinase AspN Reaction Buffer:
50 mM Tris-HCl
2.5 mM ZnSO4
pH 8 @ 37°C

Usage Concentration

100 ng/μl

Storage Temperature

-20°C

Molecular Weight

Theoretical: 40089.9 daltons

Notes

  1. Storage Conditions: Supplied in lyophilized form. Can be stored frozen in solution at -20°C for up to 2 weeks. A decrease in activity will occur if stored in solution. Use only freshly reconstituted protease for best results.
  2. Endoproteinase AspN contains a O-linked carbohydrate on the protein. The protein appears as a single band by SDS-PAGE analysis.
  3. Aspartic acid residues are strongly favored by Endoproteinase AspN in all buffer conditions we have examined (Tris-HCl, ammonium bicarbonate and potassium phosphate) (2).
  4. Endoproteinase AspN is recommended for cleavage of proteins of 5 KDa or smaller.

References

  1. Tarentino A.L. Flavastacin (2004). In Handbook of Proteolytic Enzymes. 2nd Ed. , 631-632, Elsevier, London.
  2. Tarentino, A.L. et al. (1995). Arch biochem biophys . 319, 281-285.
  3. Grimwood, B.G., Plummer, T.H. and Tarentino, A.L. (1994). Arch Biochem Biophys . 311, 127-132.

FAQs

  1. Is your Endoproteinase AspN the same as the Endoproteinase AspN sold by other companies?
  2. The Endoproteinase AspN is not fully dissolving when I reconstitute it in water. How do I get it completely into solution?
  3. Which residues does Endoproteinase AspN cut?
  4. I can’t get Endoproteinase AspN to digest my protein.
  5. Do you have a protocol or pointers for digesting proteins in-gel with endo- proteinase Asp-N followed by Endoproteinase Glu-C? I'm following the protocol for each but would like to streamline the combined digest.
  6. How pure is Endoproteinase AspN? Has it been checked for nuclease activity?
  7. Tris will interfere with my reactions downstream. Does your Endoproteinase AspN, when reconstituted in H2O, contain Tris-HCl?
  8. What is the Endoproteinase AspN sequence in text format?
  9. Can Endoproteinase AspN be used with a volatile buffer?
  10. How stable is this enzyme? We dissolved our enzyme in water approx 2 weeks ago and the literature indicates that activity will start to drop after this length of storage time(and longer).
  11. I digested BSA with Endoproteinase AspN following standard conditions and examined the released fragments with Mass-spec. I did not see many peaks that matched the predicted fragment but saw a lot of unpredicted bands, perhaps indicating non-specific enzyme activity.
  12. Can endoproteinase AspN be heat inactivated?

Datacards

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.

Selection Charts

Interactive Tools

Quality Control

Quality Assurance Statement

  • Endoproteinase AspN is free of glycerol and detergents which may interfere with Matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) or Electrospray Ionization (ESI) Mass Spectrometry (MS), or liquid chromatography (LC) methods.

Quality Control Assays

The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page. Further information regarding NEB product quality can be found here.
  • Specific Activity (Fluorometric):
    The endoproteinase is tested in a reaction with labeled peptide and the initial rate of digestion is determined by measurement of the increase in fluorescence

Safety Data Sheet

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Datacards

The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.