For trypsin digestion of peptides, use a ratio of about 1:100 to 1:20 for trypsin:peptide. The typical use for this product is in removing adherent cells from a culture surface. The concentration of trypsin necessary to dislodge cells from their substrate is dependent primarily on the cell type and the age of the culture. Trypsins have also been used for the re-suspension of cells during cell culture, in proteomics research for digestion of proteins and in various in-gel digestionsns†. Additional applications include assessing crystallization by membrane-based techniques and in a study to determine that protein folding rates and yields can be limited by the presence of kinetic traps.
Biochem/physiol Actions
Trypsin cleaves peptides on the C-terminal side of lysine and arginine residues. The rate of hydrolysis of this reaction is slowed if an acidic residue is on either side of the cleavage site and hydrolysis is stopped if a proline residue is on the carboxyl side of the cleavage site. The optimal pH for trypsin activity is 7-9. Trypsin can also act to cleave ester and amide linkages of synthetic derivatives of amino acids. EDTA is added to trypsin solutions as a chelating agent that neutralizes calcium and magnesium ions that obscure the peptide bonds on which trypsin acts. Removing these ions increases the enzymatic activity.
Serine protease inhibitors, including DFP, TLCK, APMSF, AEBSEF, and aprotinin, amongst others, will inhibit Trypsin.
Unit Definition
1 U corresponds to the amount of enzyme which increases the absorbance at 253 nm by 0.001 per minute at pH 7.6 and 25°C (N-benzoyl-L-arginine ethyl ester, Cat. No. 12880, as substrate)
One BAEE unit will produce a ΔA253 of 0.001 per min at pH 7.6 at 25° C using BAEE as substrate. One BTEE unit = 320 ATEE units. Reaction volume = 3.2 mL (1 cm light path).
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A simple and efficient method based on magnetic-bead technology has been developed for the separation of phosphorylated and nonphosphorylated low-molecular-weight biomolecules, such as nucleotides, phosphorylated amino acids, or phosphopeptides. The phosphate-binding site on the bead is an alkoxide-bridged dinuclear zinc(II)
Rapid communications in mass spectrometry : RCM, 27(6), 655-670 (2013-02-19)
Matrix-assisted laser desorption/ionization (MALDI) imaging mass spectrometry provides the means to map the in situ distribution of tryptic peptides in formalin-fixed clinical tissue samples. The ability to analyze clinical samples is of great importance to further developments in the imaging
A vast amount of enzyme activity fails to be absorbed within the human pancreas: implications for cost-effective islet isolation procedures.
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