Chymotrypsin  (Synonyms: EC 3.4.21.1; Chymotrypsin A)

Chymotrypsin (Chymotrypsin A) is a serine protease produced by the pancreas. Chymotrypsin cleaves protein chains at the carboxyl side of aromatic amino acids^[1][2].

Recommended to reconstitute in 1mM HCl at 10 mg/mL and add 2 mM calcium chloride as stabilizer. Resuspended Chymotrypsin can be stored for up to one week at 4 Celsius.

Preparation of Protein
1. Solubilization/Denaturation: Dissolve protein in 100 mM Tris-HCl, 10mM CaCl₂ (pH 8.0). Proteins that are difficult to dissolve or require denaturation for efficient digestion can be solubilized in a minimum volume in a denaturant such as 6-8 M urea or 6 M guanidine HCl at room temperature to 37 Celsius for up to one hour. For some proteins, it may be beneficial to heat the sample to 60 Celsius over this time period (95 °C for 15-20 minutes for extreme cases).
2. Disulphide Reduction: To the dissolved protein add DTT (or β-mercaptoethanol) to a final concentration of 5 mM; heat this sample at 50-60 Celsius for 20 minutes.
3. Alkylation: Allow the reduced protein mixture to cool to room temperature, and add iodoacetamide to a final concentration of 15 mM. Incubate in the dark for 15 minutes at room temperature.
4. Finally adjust the reaction volume with 100mM Tris-HCl, 10mM CaCl₂ (pH 8.0) such that the urea or guanidine concentration is 1M or less.
Enzyme Reconstitution
Dissolve Chymotrypsin in 1mM HCl. We recommend a final concentration 0.5-1 μg/μL.
Digestion
Add Chymotrypsin to a final protease:protein ratio of 1:200 to 1:20 (w/w), and incubate sample for 2-18 hours at 25 Celsius. The reaction may be stopped, if desired, by adding 0.5% trifluoroacetic acid.

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.

9004-07-3

Solid

White to off-white

[Chymotrypsin]

Room temperature in continental US; may vary elsewhere.

Please store the product under the recommended conditions in the Certificate of Analysis.

• [1]. W.R. Terra, et al. 4.5 - Biochemistry of Digestion, Editor(s): Lawrence I. Gilbert, Comprehensive Molecular Insect Science, Elsevier, 2005, Pages 171-224.

  [2]. Steven W. Cotton. Chapter 33 - Evaluation of exocrine pancreatic function, Editor(s): William Clarke, Mark A. Marzinke, Contemporary Practice in Clinical Chemistry (Fourth Edition), Academic Press, 2020, Pages 573-585.