Product Class: Other

Trypsin-ultra™, Mass Spectrometry Grade
NEBU 37

P8101_cutsite

Product Introduction

Trypsin-ultra, Mass Spectrometry Grade is a serine endopeptidase, which selectively cleaves peptide bonds C-terminal to lysine and arginine residues.  Trypsin-ultra cleaves at Lys-Pro and Arg-Pro bonds at a much slower rate than when Lys and Arg are N-terminal to other residues.

  • Analyze complex proteomes with minimal autolysis
  • Compatible for simultaneous co-digestion with PNGase F (NEB# P0709)
  • Compatible for simultaneous co-digestion with Endoproteinase LysC (NEB# P8109)
  • Suitable for both in-gel and solution digests
  • Supplied in dry format
  • Optimal activity and stability for up to 12 months
Catalog # Size Concentration
P8101S 100.0 µg

Product Information

Description

Trypsin-ultra™, Mass Spectrometry Grade is a serine endopeptidase. It selectively cleaves peptide bonds C-terminal to lysine and arginine residues (1). Trypsin-ultra is treated with L-(tosylamido-2-phenyl) ethyl chloromethyl ketone (TPCK) to inactivate any remaining chymotryptic activity. It is modified by acetylation of the ε-amino groups of lysine residues to prevent autolysis. Trypsin-ultra (TPCKtreated) cleaves at Lys-Pro and Arg-Pro bonds at a much slower rate than other amino acid residues (2).

Digestion with Trypsin-ultra results in a high number of proteins identified, and sequence coverages as high as 90%

3-hour Trypsin-ultra digest of Pyrococcus furiosus
A 3-hour Trypsin-ultra digest of Pyrococcus furiosus using a 1:50 enzyme:substrate ratio, followed by ESI-MS, resulted in 418 proteins identified with individual protein sequence coverage as high as 90%.

Product Source

Isolated from bovine (Bos taurus) pancreas.

Reconstitution

Trypsin-ultra, Mass Spectrometry Grade should be reconstituted by the addition of 20–200 μl of high purity water. Rapid autolysis is a function of enzyme concentration.
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
  • P8101S     -20    
      Trypsin-ultra™, Mass Spectrometry Grade P8101SVIAL -20 5 x 20 µg Not Applicable
      Trypsin-ultra, Reaction Buffer B8101SVIAL -20 1 x 1.5 ml 2 X

Properties & Usage

Reaction Conditions

1X Trypsin-ultra, Reaction Buffer
Incubate at 37°C

1X Trypsin-ultra, Reaction Buffer
50 mM Tris-HCl
20 mM CaCl2
(pH 8 @ 25°C)

Usage Concentration

100 ng/μl

Molecular Weight

Theoretical: 23675 daltons

Application Features

  • Digestion of proteins for proteomic analysis by Mass Spectrometry
  • Protein and peptide identification

Product Notes

  1. Substrate must be in phosphate-free buffer to prevent calcium precipitation with both reconstituted enzyme and enzyme buffer.
  2. Trypsin-ultra, Mass Spectrometry Grade is acetylated on multiple lysine residues. This protein appears as a single band on SDS-PAGE. This sequence is also available at www.neb.com.
  3. Storage Conditions: Supplied in dry format from a sodium acetate and calcium chloride buffer. Store at -20°C.
  4. 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.

References

  1. Northrop, J.H. and Kunitz, M. (1931). Isolation of protein crystals possessing tryptic activity. Science. 73,
  2. Perona, J.J. and Craik, C.S. (1995). Structural basis of substrate specificity in the serine proteases. Protein Sci.. 4, 337-360.

Protocols, Manuals & Usage

Protocols

  1. Protocol using Trypsin-ultra™, Mass Spectrometry Grade (P8101)
  2. Trypsin Digestion Protocol using NEB Trypsin-ultra™ and the FASP Kit

Application Notes

Tools & Resources

Selection Charts

FAQs & Troubleshooting

FAQs

  1. Is there a compatible buffer for digesting with Trypsin (P8101S) and Endoproteinase GluC (P8100S) together?
  2. I have a very low concentration of protein and would prefer not to denature as a separate step with buffer exchange before digestion. What denaturants can he use in the Trypsin reaction itself?
  3. I would like to use Trypsin to digest some proteins which are present in a cell culture supernatant. The supernatant is composed of DMEM supplemented with 0.2% BSA and antibiotics. Will the enzyme work in the cellular supernatant? The final reaction volume will be 100-300 ul.
  4. I am using Trypsin and am wondering about specificity. Does it cut at additional sites when in high concentration?
  5. I want to know if calcium can be left out of the buffer since it is causing my DNA-protein complex to precipitate.
  6. I want to use Trypsin on 20 ug of protein. How much enzyme do I need to use?
  7. How does one do a Trypsin in-gel digest?
  8. Is there a simple way to remove Trypsin after protein cleavage?

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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