Trypsin protein digestion

The term catalysis was applied to the biological phenomena listed above and to several other reactions discovered in the next several years. The active component of bitter almonds was named emulsin in 1837. Other carbohydrate-splitting activities (ptyalin in saliva, amylase in malt) had already been described. Pepsin and trypsin, protein-digesting agents from the stomach and pancreas, were also discovered during this period. The activities of these materials were contrasted with the materials responsible for fermentation. Early theories of Willis and of Stahl to explain fermentation as a disruption caused by violent motion of... 

FIGURE 2.16 pH versus enzymatic activity. The activity of enzymes is very sensitive to pH. The pH optimum of an enzyme is one of its most important characteristics. Pepsin is a protein-digesting enzyme active in the gastric fluid. Trypsin is also a proteolytic enzyme, but it acts in the more alkaline milieu of the small intestine. Lysozyme digests the cell walls of bacteria it is found in tears. 

Dilute the solution 4-fold with lOOmM 25mM NH4HCO3, and then add 20 pg trypsin to digest the proteins. Incubate at 37°C for 4 hours or overnight at 30°C with shaking. 

The positive effect of convection of the substrate solution on mass transfer can be observed even better with macromolecular substrates that undergo processes such as protein digestion. For example, Fig. 9 compares reversed-phase chromatograms of cytochrome c digests obtained by cleavage with trypsin immobilized in both packed and molded column reactors, and clearly demonstrates the much higher activity of the monolithic device under otherwise similar circumstances [90]. 

Wang C., Oleschuk R., Ouchen F., Li J., Thibault R, and Harrison D.J. (2000), Integration of immobilized trypsin bead beds for protein digestion within a micro-fluidic chip incorporating capillary electrophoresis separations and an electrospray mass spectrometry interface, Rapid Commun. Mass Spectrom. 14(15), 1377-1383. 

Protein digestion occurs in two stages endopeptidases catalyse the hydrolysis of peptide bonds within the protein molecule to form peptides, and the peptides are hydrolysed to form the amino acids by exopeptidases and dipeptidases. Enteropeptidase initiates pro-enzyme activation in the small intestine by catalysing the conversion of trypsinogen into trypsin. Trypsin is able to achieve further activation of trypsinogen, i.e. an autocatalytic process, and also activates chymotrypsinogen and pro-elastase, by the selective hydro-... 

Trypsin is the most frequently used serine protease. Generally, porcine or bovine pancrease is used as the source of pure trypsin. Trypsin usually digests proteins at their lysine and arginine residues. The superiority of trypsin is that it displays good activity both in solution and in-gel digestion protocols. 

Modified trypsin is preferred for protein digestion, as it is less susceptible to autodigestion (7). The trypsin 10X stock solution is prepared by dissolving the trypsin (20 gig) in the solvent supplied with the enzyme (50 mMacetic acid 200 pL). This is stored at -70°C in aliquots (10 pL) and is stable for at least 1 yr. 

On-line protein digestion has been achieved by trypsin adsorbed in a porous PVDF (poly [vinylidene fluoride]) membrane (0.45-pm pore) coupled in a PDMS chip. In this way, on-chip protein digestion and subsequent MS analysis have been carried out for horse heart cytochrome c [817]. 

For trypsinogen activation, enteropeptidase (enterokinase, EC 3.4.21.9) [23] is a key enzyme for mammalian protein digestion. The selective cleavage site of trypsinogen by enteropeptidase initiates Lys6-Ile7 bond. Then trypsin (EC 3.4.21.4) activates other zymogens. Thus, the formation of trypsin by enteropeptidase is the master activation step. 

Peptide mapping studies, generated by the cleavage of a protein into peptide fragments, must be highly reproducible and quantitative. Several electropherograms of protein digests have been obtained when chicken ovalbumin was cleaved by trypsin... 

Trypsin, chymotrypsin, and elastase are three of the most important protein-digesting enzymes secreted by the pancreas. Despite their similarities they have different substrate specificity, that is, they cleave different peptide bonds during protein digestion. 

Rapid protein digestion, Trypsin Poly(vinylidene fluoride) [354]... 

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