Trypsin (EC 18.104.22.168) is a serine protease from the PA clan superfamily, found in the digestive system of many vertebrates, where it hydrolyses proteins. Trypsin is produced in the pancreas as the inactive proenzyme trypsinogen. Trypsin cleaves peptide chains mainly at the carboxyl side of the amino acids lysine or arginine, except when either is followed by proline. It is used for numerousbiotechnological processes. The process is commonly referred to as trypsin proteolysis or trypsinisation, and proteins that have been digested/treated with trypsin are said to have been trypsinized.
Activation of trypsin from proteolytic cleavage of trypsinogen in the pancreas can lead to a series of events that cause pancreatic self-digestion, resulting in pancreatitis. One consequence of the autosomal recessive disease cystic fibrosis is a deficiency in transport of trypsin and other digestive enzymes from the pancreas. This leads to the disorder termed meconium ileus. This disorder involves intestinal obstruction (ileus) due to overly thick meconium, which is normally broken down by trypsin and other proteases, then passed in feces.
Trypsin is available in high quantity in pancreases, and can be purified rather easily. Hence it has been used widely in various biotechnological processes.
In a tissue culture lab, trypsin is used to re-suspend cells adherent to the cell culture dish wall during the process of harvesting cells. Some cell types have a tendency to “stick” – or adhere – to the sides and bottom of a dish when cultivated in vitro. Trypsin is used to cleave proteins bonding the cultured cells to the dish, so that the cells can be suspended in fresh solution and transferred to fresh dishes.
Trypsin can also be used to dissociate dissected cells (for example, prior to cell fixing and sorting).
Trypsin can be used to break down casein in breast milk. If trypsin is added to a solution of milk powder, the breakdown of casein will cause the milk to become translucent. The rate of reaction can be measured by using the amount of time it takes for the milk to turn translucent.
Trypsin is commonly used in biological research during proteomics experiments to digest proteins into peptides for mass spectrometry analysis, e.g. in-gel digestion. Trypsin is particularly suited for this, since it has a very well defined specificity, as it hydrolyzes only the peptide bonds in which the carbonyl group is contributed either by an Arg or Lys residue.
Trypsin can also be used to dissolve blood clots in its microbial form and treat inflammation in its pancreatic form.
Commercial protease preparations usually consist of a mixture of various protease enzymes that often includes trypsin. These preparations are widely utilized in food processing:
- as a baking enzyme to improve the workability of dough;
- in the extraction of seasonings and flavourings from vegetable or animal proteins and in the manufacture of sauces;
- to control aroma formation in cheese and milk products;
- to improve the texture of fish products;
- to tenderize meat;
- during cold stabilization of beer;
- in the production of hypoallergenic food where proteases break down specific allergenic proteins into nonallergenic peptides. For example, proteases are used to produce hypoallergenic baby food from cow’s milk thereby diminishing the risk of babies developing milk allergies.
In order to prevent the action of active trypsin in the pancreas which can be highly damaging, inhibitors such as BPTI and SPINK1 in the pancreas and α1-antitrypsin in the serum are present as part of the defense against its inappropriate activation. Any trypsin prematurely formed from the inactive trypsinogen would be bound by the inhibitor. The protein-protein interaction between trypsin and its inhibitors is one of the tightest found, and trypsin is bound by some of its pancreatic inhibitors essentially irreversibly. In contrast with nearly all known protein assemblies, some complexes of trypsin bound by its inhibitors do not readily dissociate after treatment with 8M urea.