PRODUCTS

TRYPSIN

Trypsin is a serine protease that plays an essential role in protein hydrolysis and absorption in mammals.
When converted from its zymogen trypsinogen, trypsin is available as an active peptide hydrolase form to cleave peptide chains, mainly at the carboxyl side of the amino acids lysine or arginine.
Trypsin contains a nucleophilic residue Ser in the enzyme active site which attacks the carbonyl moiety of the substrate peptide bond to form an acyl-enzyme intermediate.
This nucleophilic attack is facilitated by the catalytic triad consisting of histidine-57, aspartate-102, and serine-195.
Trypsin also contains an oxyanion hole that stabilizes the charge negative charge on the carbonyl oxygen atom formed from the cleavage of peptide bonds.
Therapeutic forms of trypsin is obtained from purified extracts of porcine or bovine pancreas and is intended to aid in digestion when administered orally.

Trypsin is a naturally occurring proteolytic enzyme found in the digestive tract of mammals.
Trypsin is used clinically in combination with bromelain and rutosides in the treatment of some inflammatory conditions.

CAS: 9002-07-7
European Community (EC) Number: 232-650-8

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Trypsin is an enzyme in the first section of the small intestine that starts the digestion of protein molecules by cutting long chains of amino acids into smaller pieces.
Trypsin is a serine protease from the PA clan superfamily, found in the digestive system of many vertebrates, where it hydrolyzes proteins.
Trypsin is formed in the small intestine when its proenzyme form, the trypsinogen produced by the pancreas, is activated.
Trypsin cuts peptide chains mainly at the carboxyl side of the amino acids lysine or arginine.
Trypsin is used for numerous biotechnological processes.
The process is commonly referred to as trypsinogen proteolysis or trypsinization, and proteins that have been digested/treated with trypsin are said to have been trypsinized.

Trypsin was discovered in 1876 by Wilhelm Kühne.
Although many sources say that Kühne named trypsin from the Ancient Greek word for rubbing, 'tripsis', because the enzyme was first isolated by rubbing the pancreas with glass powder and alcohol, in fact Kühne named trypsin from the Ancient Greek word 'thrýpto' which means 'I break' or 'I break apart'.

Trypsin is a serine protease enzyme, and hydrolyzes proteins at the carboxyl side of the Lysine or Arginine.
Trypsin activates PAR2 and PAR4.
Trypsin induces cell-to-cell membrane fusion in PDCoV infection by the interaction of S glycoprotein of PDCoV and pAPN.
Trypsin also promotes cell proliferation and differentiation.
Trypsin can be used in the research of wound healing and neurogenic inflammation.

Trypsin is a naturally occurring proteolytic enzyme found in the digestive tract of mammals.
Used clinically in combination with bromelain and rutosides in the treatment of some inflammatory conditions.

Trypsin or serine protease 1 is a medium size globular protein that functions as a pancreatic serine protease.
This enzyme hydrolyzes bonds by cleaving peptides on the C-terminal side of the amino acid residues lysine and arginine.
Trypsin has also been shown that cleavage will not occur if there is a proline residue on the carboxyl side of the cleavage site.
Trypsin was first discovered in 1876 by Kuhne, who investigated the proteolytic activity of the enzyme.
In 1931 the enzyme was purified by crystallization by Norothrop and Kunitz and later in 1974 the three dimensional structure of trypsin was determined.
Throughout the 1990's the role of trypsin in hereditary pancreatitis and the mutation that causes it was discovered.
Today trypsin is used in the development of cell and tissue protocols, as well as in the medical field to determine the role of trypsin in pancreatic diseases.

Trypsin (EC 3.4.21.4) 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 protease trypsinogen.
Trypsin cleaves peptide chains mainly at the carboxyl side of the amino acids lysine or arginine, except when either is followed by proline.
Trypsin is used for numerous biotechnological 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.

Cationic trypsin or Trypsin-1 is expressed in the pancreas.
Trypsin cleaves linkages involving lysine and arginine.

Anionic trypsin or Trypsin-2 is expressed in the pancreas.

Mesotrypsin or Trypsin-3 is expressed in brain and pancreas and is resistant to common trypsin inhibitors.

Trypsinogen is the precursor form or zymogen of Trypsin.
Zymogens are enzyme precursors that are made and secreted in the lysosome of the cell.
Zymogens are not active until they go through a chemical process such as hydrolysis, cleavage, or other cleavages that reveal the active site.
The zymogen precursor is necessary in order to prevent the destruction of cellular proteins and to allow the enzyme to be in it's active state only when in appropriate conditions.
Trypsinogen is specifically produced in the exocrine cells of the pancreas.
There are three isoforms of Trypsinogen that are secreted by the pancreas.
The precursor is only activated when it reaches the lumen of the small intestine.
This activation occurs through the help of an enteropeptidase and once activated trypsin stimulates the formation of more trypsinogen.
The structure of Bovine Trypsinogen is shown in the figure to the right.
Trypsin has many applications due to fact that it is easily purified in high quantities.
The trypsin enzyme is often used in the research setting to digest proteins and then identify the resulting peptides using mass spectrometry.
Trypsin has many uses in the medical field such as dissolving blood clots and treating inflammation.
Other applications include its use in pre-digesting of baby food, fingerprinting and sequencing work, and environmental monitoring.

Trypsin is a serine protease in the digestive system of human and animals.
The main function of this enzyme is to hydrolyze proteins into smaller peptides or even amino acids.
Trypsin and other digestive proteases such as chymotrypsin are responsible for the digestion of food protein in the small intestine.
This proteolytic function of trypsin has been widely used in the protein chemistry, proteomics, and nutrition research.
This function is influenced by the sources of enzyme, and environmental factors such as pH, temperature, and the presence of trypsin inhibitors in the enzymatic reaction medium.
Trypsin is used in the food processing to improve the functional properties such as solubility, emulsification, foaming and gelling properties of food proteins, to improve the digestibility of vegetable and seed proteins.
Trypsin is used to reduce the concentration of allergens in some foods and to produce protein hydrolysates and bioactive peptides that are used in infant formulas and for people with special health problems such as hypertension.
In food science research, trypsin is used for the food protein sequencing, in-vitro determination of food protein digestibility.
In combination with bromelain and rutin, trypsin is used for osteoarthritis.
Trypsin is used to remove necrotic tissue and debris during wound and ulcer cleaning.
Trypsin supplements may be used to remove dead tissue cells that remain after trauma, infection or surgical procedures, allowing new skin or tissue cells to grow.

Properties
Melting point: 115°C
Density: 1.37[at 20℃]
vapor pressure: 0Pa at 25℃
storage temp.: -20°C
solubility: Reconstitute in aqueous buffer
form: lyophilized powder
pka: pK1:6.25 (25°C,μ=0.1)
color: White powder
Odor: Odorless
Water Solubility: Soluble in water (10 mg/ml), phosphate buffers (10 mg/ml), and balanced salt solutions (1 mg/ml).
Merck: 13,9865
Stability: Stable. Incompatible with strong oxidizing agents.
LogP: -1.3 at 20℃

Function
In the duodenum, trypsin catalyzes the hydrolysis of peptide bonds, breaking down proteins into smaller peptides.
The peptide products are then further hydrolyzed into amino acids via other proteases, rendering them available for absorption into the blood stream.
Tryptic digestion is a necessary step in protein absorption, as proteins are generally too large to be absorbed through the lining of the small intestine.

Trypsin is produced as the inactive zymogen trypsinogen in the pancreas.
When the pancreas is stimulated by cholecystokinin, it is then secreted into the first part of the small intestine (the duodenum) via the pancreatic duct.
Once in the small intestine, the enzyme enterokinase (also called enteropeptidase) activates trypsinogen into trypsin by proteolytic cleavage.
The trypsin then activates additional trypsin, chymotrypsin and carboxypeptidase.

Application
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 digestions.
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.

Components
Trypsin consists of a single chain polypeptide of 223 amino acid residues, produced by the removal of the N-terminal hexapeptide from trypsinogen which is cleaved at the Lys-lle peptide bond.
The sequence of amino acids is cross-linked by 6 disulfide bridges.
This is the native form of trypsin, beta-trypsin.
BETA-trypsin can be autolyzed, cleaving at the Lys-Ser residue, to produce alpha-trypsin.
Trypsin is a member of the serine protease family.

Mechanism
The enzymatic mechanism is similar to that of other serine proteases.
These enzymes contain a catalytic triad consisting of histidine-57, aspartate-102, and serine-195.
This catalytic triad was formerly called a charge relay system, implying the abstraction of protons from serine to histidine and from histidine to aspartate, but owing to evidence provided by NMR that the resultant alkoxide form of serine would have a much stronger pull on the proton than does the imidazole ring of histidine, current thinking holds instead that serine and histidine each have effectively equal share of the proton, forming short low-barrier hydrogen bonds therewith.
By these means, the nucleophilicity of the active site serine is increased, facilitating its attack on the amide carbon during proteolysis.
The enzymatic reaction that trypsin catalyzes is thermodynamically favorable, but requires significant activation energy (it is "kinetically unfavorable").
In addition, trypsin contains an "oxyanion hole" formed by the backbone amide hydrogen atoms of Gly-193 and Ser-195, which through hydrogen bonding stabilize the negative charge which accumulates on the amide oxygen after nucleophilic attack on the planar amide carbon by the serine oxygen causes that carbon to assume a tetrahedral geometry.
Such stabilization of this tetrahedral intermediate helps to reduce the energy barrier of its formation and is concomitant with a lowering of the free energy of the transition state.
Preferential binding of the transition state is a key feature of enzyme chemistry.

The negative aspartate residue (Asp 189) located in the catalytic pocket (S1) of trypsin is responsible for attracting and stabilizing positively charged lysine and/or arginine, and is, thus, responsible for the specificity of the enzyme. This means that trypsin predominantly cleaves proteins at the carboxyl side (or "C-terminal side") of the amino acids lysine and arginine except when either is bound to a C-terminal proline, although large-scale mass spectrometry data suggest cleavage occurs even with proline.
Trypsin is considered an endopeptidase, i.e., the cleavage occurs within the polypeptide chain rather than at the terminal amino acids located at the ends of polypeptides.

Properties
Human trypsin has an optimal operating temperature of about 37 °C.
In contrast, the Atlantic cod has several types of trypsins for the poikilotherm fish to survive at different body temperatures.
Cod trypsins include trypsin I with an activity range of 4 to 65 °C (40 to 150 °F) and maximal activity at 55 °C (130 °F), as well as trypsin Y with a range of 2 to 30 °C (36 to 86 °F) and a maximal activity at 21 °C (70 °F).

As a protein, trypsin has various molecular weights depending on the source.
For example, a molecular weight of 23.3 kDa is reported for trypsin from bovine and porcine sources.

The activity of trypsin is not affected by the enzyme inhibitor tosyl phenylalanyl chloromethyl ketone, TPCK, which deactivates chymotrypsin.

Trypsin should be stored at very cold temperatures (between −20 and −80 °C) to prevent autolysis, which may also be impeded by storage of trypsin at pH 3 or by using trypsin modified by reductive methylation.
When the pH is adjusted back to pH 8, activity returns.

Clinical significance
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, which involves intestinal obstruction (ileus) due to overly thick meconium, which is normally broken down by trypsin and other proteases, then passed in feces.

Applications
Trypsin is available in high quantity in pancreases, and can be purified rather easily.
Hence, trypsin has been used widely in various biotechnological processes.

In a tissue culture lab, trypsin is used to resuspend cells adherent to the cell culture dish wall during the process of harvesting cells.
Some cell types adhere to the sides and bottom of a dish when cultivated in vitro.
Trypsin is used to cleave proteins holding the cultured cells to the dish, so that the cells can be removed from the plates.

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 causes the milk to become translucent.
The rate of reaction can be measured by using the amount of time needed 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 arginine or lysine residue.

Trypsin can also be used to dissolve blood clots in its microbial form and treat inflammation in its pancreatic form.

In food

Commercial protease preparations usually consist of a mixture of various protease enzymes that often includes trypsin.
These preparations are widely used in food processing:

- as a baking enzyme to improve the workability of dough
- in the extraction of seasonings and flavorings 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.

Trypsin is a serine protease that specifically cleaves at the carboxylic side of lysine and arginine residues.
The stringent specificity of trypsin is essential for protein identification, and it has become the gold standard for protein digestion to peptides for shotgun proteomics.

Trypsin is an enzyme that aids with digestion.
An enzyme is a protein that speeds up a certain biochemical reaction.
Trypsin is found in the small intestine. It can also be made from fungus, plants, and bacteria.
But it is usually made for commercial purposes from the pancreas of livestock.

Trypsin is given to people who lack enzymes needed for digestion.

Trypsin is also given in combination with bromelain and rutin for treatment of osteoarthritis and many other conditions, but there is no good scientific evidence to support these other uses.

Some people apply trypsin directly to wounds and ulcers to remove dead tissue and improve healing.

Trypsin removes dead skin cells (tissue) and allows healthy tissue to grow.
Trypsin in combination with other enzymes seems to reduce inflammation and swelling.