Trypsins, pancreatic

Using this approach, Bizzozero and Zweifel (9) and Bizzozero and Dutler (10) have constructed molecular models of two intermediates (an enzyme-substrate complex and a tetrahedral intermediate) by appropriate modification of the models of stable enzyme-species. The stable enzyme-species used (15, 16) are trypsin-benzamidine complex (TR-B) (17), trypsin-pancreatic trypsin inhibitor complex (TR-PTI) (18, 19) and tosyl-chymotrypsin (Tos-CHT) (20) which are related to enzyme substrate complex, tetrahedral intermediate and acyl-enzyme respectively. 

Dietary triglycerides are hydrolyzed in the small intestine by pancreatic lipase. This enzyme action is associated with a cofactor, colipase, also a pancreatic protein, molecular weight (MW) 12,000, which helps to anchor lipase to the fat droplets. Without colipase, lipase is rapidly denatured. Colipase is apparently secreted by the pancreas as a zymogen and is activated to its active form in the small intestine by trypsin. Pancreatic lipase appears in the circulation in large amounts during acute pancreatitis. 

The hydrophobic free energy contribution to the stability of protein-protein complexes can also be estimated making similar assumptions to the ones described above (9). In this case as well (Table V) it appears that hydrophobic interactions contribute greatly to the overall stability of the complexes. This is particularly interesting in the case of the trypsin-pancreatic trypsin inhibitor complex since very few residues are involved in the interaction of the two molecules. 

The exocrine pancreas secretes phospholipase A2 in an inactive zymogen form, prophospholipase A2. The enzyme is activated in the intestinal lumen by proteolytic cleavage by trypsin. Pancreatic lipase, however, is secreted in its active form, and only needs to bind colipase and substrate to be active. 

Vincent, J. P., and Lazdunski, M. (1972). Trypsin-pancreatic trypsin inhibitor association. Dynamics of the interaction and role of disulfide bridges. Biochemistry 11(16), 2967-2977. 

This phenomenon has been observed by Abderhalden anpancreatic juice, and the extract of yeast, by Schmidt-Nielsen with rennet, by Horlow with ptyalin, by Shaklle and Meltzer with pepsin. The results, however, depend on experimental conditions imder which this shakin takes place. Duration and rapidity, the content of the liquid in enzyme, and the temperature play an important r61e in this% phenomenon. An active solution, introduced into a reaction tube and agitated for 2 minutes, can lose as much as 75 per cen.t of its activity. After 5 minutes the disappearance is almost ... 

A, MeloandM. J. Ramos,/. Peptide Res., SO, 382 (1997). The Nature of Trypsin-Pancreatic Trypsin Inhibitor Binding Free Energy Calculation of Tyr39 -> Phe39 Mutation in Trypsin,... 

As examples of applications, we present the overall accuracy of predicted ionization constants for about 50 groups in 4 proteins, changes in the average charge of bovine pancreatic trypsin inhibitor at pH 7 along a molecular dynamics trajectory, and finally, we discuss some preliminary results obtained for protein kinases and protein phosphatases. 

The presented algorithm was applied to 4 proteins (lysozyme, ribonuclease A, ovomucid and bovine pancreatic trypsin inhibitor) containing 51 titratable residues with experimentally known pKaS [32, 33]. Fig. 2 shows the correlation between the experimental and calculated pKaS. The linear correlation coefficient is r = 0.952 the slope of the line is A = 1.028 and the intercept is B = -0.104. This shows that the overall agreement between the experimental and predicted pKaS is good. 

M. H. Hao, M. R. Pincus, S. Rackovsky, and H. A. Scheraga. Unfolding and refolding of the native structure of bovine pancreatic trypsin inhibitor studied by computer simulations. Biochemistry, 32 9614-9631, 1993. 

In periodic boimdary conditions, one possible way to avoid truncation of electrostatic interaction is to apply the so-called Particle Mesh Ewald (PME) method, which follows the Ewald summation method of calculating the electrostatic energy for a number of charges [27]. It was first devised by Ewald in 1921 to study the energetics of ionic crystals [28]. PME has been widely used for highly polar or charged systems. York and Darden applied the PME method already in 1994 to simulate a crystal of the bovine pancreatic trypsin inhibitor (BPTI) by molecular dynamics [29]. 

Brooks B and M Karplus 1983. Harmonic Dynamics of Proteins Normal Modes and Fluctuations in Bovine Pancreatic Trypsin Inhibitor. Proceedings of the National Academy of Sciences USA 80 6571-6575. 

The so-called "trypsin," obtainable from pancreatic juice and from fresh extracts of the pancreas, is not a simple enzyme but a mixture of trypsin proper (which hydrolyses proteins to proteoses and peptones) and a series of enzymes which hydrolyse these breakdown products to their constituent amino-acids. The term trypsin," when used below, refers to this mixture. 

Prior to the bating process, the hides are delimed with ammonium sulfate and/or ammonium chloride. Proteases are then appUed. The early preparation proposed by Rn hm was pancreatic trypsin. The use of a bating enzyme makes the hides soft and supple to prepare them for tanning. A new microbial protease, Pyrase 250 MP (82) (Novo Nordisk A/S) has been found to be a promising substitute for pancreatic trypsin [9002-07-7] which is more expensive because it must be extracted from pancreatic glands. 

Mucolytics reduce the viscosity of tenacious and purulent mucus, thus faciUtating removal. The distinction between mucolytics and other classes of expectorants is frequently blurred. Steam, sometimes in conjunction with surfactants or volatile oils, has long been used to decrease viscosity by physical hydration. However, agents that chemically depolymerize certain components of mucus are available. Trypsin and other proteolytic enzymes have shown good clinical activity because of their abiUty to cleave glycoproteins. Pancreatic domase, which depolymerizes DNA found in purulent mucus, also has shown clinical utihty. 

To date, a number of simulation studies have been performed on nucleic acids and proteins using both AMBER and CHARMM. A direct comparison of crystal simulations of bovine pancreatic trypsin inliibitor show that the two force fields behave similarly, although differences in solvent-protein interactions are evident [24]. Side-by-side tests have also been performed on a DNA duplex, showing both force fields to be in reasonable agreement with experiment although significant, and different, problems were evident in both cases [25]. It should be noted that as of the writing of this chapter revised versions of both the AMBER and CHARMM nucleic acid force fields had become available. Several simulations of membranes have been performed with the CHARMM force field for both saturated [26] and unsaturated [27] lipids. The availability of both protein and nucleic acid parameters in AMBER and CHARMM allows for protein-nucleic acid complexes to be studied with both force fields (see Chapter 20), whereas protein-lipid (see Chapter 21) and DNA-lipid simulations can also be performed with CHARMM. 

M Vasquez, ElA Scheraga. Calculation of protein conformation by the build-up procedure. Application to bovine pancreatic trypsin inhibitor using limited simulated nuclear magnetic resonance data. J Biomol Struct Dyn 5 705-755, 1988. 

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