Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An electrophoretic technique for the determination of enterokinase activity is described. The natural substrate trypsinogen is hydrolysed in the presence of soybean trypsin inhibitor. Under the conditions of assay, neither the trypsin inhibitor nor the trypsin-trypsin inhibitor complex are detected. Enterokinase activity can be determined in biological materials such as duodenal aspirates without any interference from trypsin activity. A significant correlation exists between the present technique and a spectrophotometric technique by which the liberation of trypsin activity is used to determine enterokinase activity.
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PMID:Determination of enterokinase activity by measuring the disappearance of trypsinogen. 139 26

An endoproteolytic activity that specifically cleaves CCK 33, producing CCK 8, has been purified from a rat brain synaptosome preparation. The purification, which included anion exchange, chromatofocusing, hydroxyapatite, and gel filtration chromatography, resulted in a greater than 3000-fold increase in specific activity. This neutral endoprotease (pH optimum 8) exists as a 90-kDa species, which can be dissociated into active 40-kDa species. The enzyme is a non-trypsin serine protease, which is inhibited by diisopropyl-fluorophosphate and p-aminobenzamidine but not by soybean trypsin inhibitor, phenylmethylsulfonyl fluoride, aprotinin, or a number of thiol or metalloprotease inhibitors. It is highly substrate-specific and cleaves neither trypsin, enteropeptidase, kallikrein substrates, nor analogues of mono- or dibasic cleavage sites of prohormones other than pro-CCK. The endoprotease will not cleave CCK 12 desulfate or CCK (20-29), although these peptides contain common sequences with CCK-33. The protease does cleave [Glu27]CCK (20-29), a peptide in which the glutamate mimics the negative charge normally present on tyrosine sulfate. This suggests that the negative charge at position 27 is important in substrate recognition. The enzyme will also cleave CCK 33 and CCK (1-21) on the carboxyl-terminal side of a single lysine residue in position 11. The subcellular location and specificity of this endoprotease make it a good candidate for a CCK-processing protease.
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PMID:Characterization of a cholecystokinin 8-generating endoprotease purified from rat brain synaptosomes. 152 68

Proteins with trypsin-like immunoreactivity (first detected by a specific immunoenzymatic assay) were isolated from CAPAN-1 and CFPAC-1 cell culture-conditioned media by chromatography on an immunoadsorbent prepared with a polyclonal antibody directed against trypsin 1. The adsorbed proteins were devoid of free trypsin activity but trypsin activity was present after enterokinase activation demonstrating that the immunoreactive trypsin present in cell supernatants corresponds to trypsinogens. When characterised by Western blotting using a monoclonal antibody directed against human trypsin 1 two protein bands corresponding to trypsinogen 1 (23 kDa) and trypsinogen 2 (25 kDa) gave a positive reaction. These results demonstrate the presence of trypsinogens 1 and 2 in CAPAN-1 and CFPAC-1 cells and in their culture-conditioned media.
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PMID:Characterization of trypsinogens 1 and 2 in two human pancreatic adenocarcinoma cell lines; CFPAC-1 and CAPAN-1. 175 57

Bovine enterokinase (enteropeptidase) is a serine protease and functions as the physiological activator of trypsinogen. The enzyme has a heavy chain (115 kD) covalently linked to a light or catalytic subunit (35 kD). The amino acid composition showed that the light chain has nine half-cystine residues (four as intramolecular disulfides) and that one half-cystine was in a disulfide link between the light and heavy subunits. The amino-terminal 27 residues of the S-vinylpyridyl derivative of the light chain were determined by gas-phase Edman degradation. The sequence has homologies with other serine proteases containing one or two chains. The homologies suggest that the catalytic subunit has the same three-dimensional structure and, therefore, the same mechanism of enzymatic action as pancreatic chymotrypsin, trypsin, and elastase. The presence of the conserved amino-terminal activation peptide sequence (IVGG) shows that enterokinase must have a zymogen precursor and that the two-chain enzyme arises from limited proteolysis during posttranslational processing.
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PMID:The amino-terminal sequence of the catalytic subunit of bovine enterokinase. 179 6

Pretreatment of the purified jack bean inhibitor with enterokinase activated human pancreatic preparation for 1 hr decreased its inhibitory capacity against crystalline bovine alpha-chymotrypsin by 30% but did not affect its trypsin inhibitory activity. Preincubation of the inhibitor with bovine chymotrypsin for 60 min resulted in partial loss of the inhibitory potency. Complex formation studies by gel chromatography on Sephadex G-100 indicated that the trypsin-inhibitor and chymotrypsin-inhibitor complexes dissociated to release inactivated inhibitor and active proteinases. Gel chromatography of the inhibitor in presence of 1.5 M ammonium sulphate indicated that the inhibitor showed a tendency to aggregate without loss of biological activity. However, in 4.2 M salt medium after 3 hr, antichymotryptic activity was lost completely without any effect on antitryptic activity. Treatment with methylamine, a nucleophile, caused a greater loss of antichymotryptic activity. Trinitrobenzene sulphonate and ethylacetamidate, the amino group modifiers, affected only the antichymotryptic activity. Treatment with ninhydrin, a specific arginine modifier, at pH 9.0 abolished the antitryptic activity whereas only 50% of the antichymotryptic activity was lost. Diethylpyrocarbonate, a histidine reagent, also decreased only the antitryptic activity. Modification of tryptophan and cysteine residues of the inhibitor had no effect on its inhibitory potency. Treatment with mercaptoethanol and sodium borohydride caused nearly 50% loss of antitryptic and antichymotryptic activities. Chloramine-T, a reagent that modifies methionine residues, inactivated the inhibitor.
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PMID:Chemical modification and complex formation studies with jack bean proteinase inhibitor. 181 77

We have recently demonstrated that many cancer cell lines produce a novel trypsinogen isoenzyme called tumor-associated trypsinogen 2 (TAT-2). It was found during a search of the target protease for tumor-associated trypsin inhibitor (TATI). We now show that degradation of subendothelial cell extracellular matrix (ECM) by four different cell lines (COLO 205 colon carcinoma, K-562 erythroleukemia, CAPAN-1 pancreatic carcinoma, and HT 1080 fibrosarcoma) can be partially inhibited by TATI or neutralizing trypsin antibodies. When cells were cultured in serum-free medium on ECM, TATI and trypsin antibodies inhibited the release of immunoreactive fibronectin fragments from ECM by 47-54 and 40%, respectively. Degradation of isotopically labeled ([3H]serine, [3H]proline, and [35S]sulfate) ECM was also significantly prevented by TATI. At its maximum, it exerted a 57% inhibition on the degradation of [3H]serine-labeled ECM. Plasminogen added exogenously to the culture medium further potentiated the proteolysis of ECM. Interestingly, addition of enteropeptidase, an activator of TAT-2, also enhanced cell-mediated proteolysis as assessed by degradation of purified fibronectin coated onto the surface of wells. Immunoblot analysis showed that enteropeptidase-mediated proteolysis generated a pattern of fibronectin fragments similar to that obtained by digestion of purified fibronectin by TAT-2. These results demonstrate the existence of a proteolytic system in tumor cells which is dependent on the activation of TAT-2. We suggest that TAT-2 is involved in a protease cascade-stimulating tumor cell invasion and degradation of extracellular matrix.
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PMID:Tumor-associated trypsin participates in cancer cell-mediated degradation of extracellular matrix. 200 30

It has been proposed that modulation of cholecystokinin (CCK) release by proteinases, proteinase inhibitors and protein is mediated by a pancreatic secretory trypsin inhibitor (PSTI), also called monitor peptide, in the rat. When human [125I]-PSTI was incubated with fasting small bowel juice or activated pancreatic juice greater than 88% of tracer eluted from gel chromatography in the characteristic position of hydrolysed PSTI. However, when the small bowel juice had been pre-incubated with soybean trypsin inhibitor 3 g/l, casein 5 g/l or lactalbumin 30 g/l, the hydrolysis of PSTI diminished so that 95%, 32%, and 33% respectively, now eluted in the characteristic position of free (i.e. intact and not bound to an enzyme) PSTI. When [125I]-PSTI was incubated with pure trypsin, chymotrypsin, elastase or enterokinase greater than 95% of tracer eluted in the position of PSTI-enzyme complex. Incubation of PSTI with trypsin plus one other enzyme was required to produce hydrolysis. The degree of protection of PSTI from hydrolysis in duodenal juice produced by these substances correlates with their affects on CCK release. Our findings support the hypothesis that PSTIs mediate the modulation of CCK release by intraluminal proteinases, proteinase inhibitors and proteins.
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PMID:Interactions of pancreatic secretory trypsin inhibitor in small intestinal juice: its hydrolysis and protection by intraluminal factors. 209 78

A global kinetic analysis of the mechanisms of the trypsinogen activation by enterokinase and trypsin is presented. The kinetic equations of both the transient-phase and the steady-state of these mechanisms are presented. In addition, we here derive the corresponding kinetic equations for the case in which the condition of rapid equilibrium prevails and we propose a kinetic data analysis. The significance of this approach to the treatment of other zymogen activation processes is discussed.
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PMID:Kinetics of the trypsinogen activation by enterokinase and trypsin. 221 4

The effects of aging upon pancreatic digestive enzymes were studied in 27- and 3-month-old Fischer 344 rats. Mean pancreatic weight, protein and DNA concentration and content, and protein-DNA ratios did not differ in the two groups of animals. Pancreatic amylase concentration was reduced by 41% and lipase concentration was increased by 29% in the aging animals, whereas, trypsinogen concentrations did not differ. Young and aging rats were fed diets enriched with fat (72%) or sucrose (75%) for seven days to define whether the different enzyme contents were intrinsic to the aging process or adaptable. In young, but not in aging rats, lipase concentration increased 25% during high fat compared to high sucrose diet feeding. High starch diet feeding induced a 26% increase in amylase in young rats but not in the old. Trypsinogen concentration was unaffected by dietary manipulation. Jejunal enteropeptidase concentration was modestly reduced in the aging rat. Postprandial luminal concentrations of trypsin and amylase did not differ in the two groups. Thus, aging may induce modest changes in pancreatic digestive enzymes and in jejunal enteropeptidase which are unlikely to be physiologically important. However, the pancreas of aging rats does not adapt to changes in dietary intake as well as young rats.
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PMID:Influence of aging upon pancreatic digestive enzymes. 242 64

Effects of FUT-175 on the pancreatic enzymes in vitro and in vivo in the enterokinase-induced experimental acute pancreatitis were investigated, and they were compared with those of gabexate and aprotinin. In in vitro experiments, FUT-175 inhibited the pancreatic protease activities 10 to 100 times more potently than gabexate. Furthermore, FUT-175 inhibited the enterokinase activity. Unlike aprotinin, FUT-175 inhibited alpha 2-macroglobulin bound trypsin activity as well as free trypsin. In in vivo experiments, at doses of 0.5-50 micrograms/kg/min, FUT-175 suppressed the elevated protease activities in the experimental acute pancreatitis more potently than gabexate. Differently from the action of aprotinin, FUT-175 suppressed trypsin activities both in the pancreas and in the plasma to the same extent. Furthermore, FUT-175 reduced the mortality of rats in the experimental acute pancreatitis in a dose-dependent manner. These data strongly support that FUT-175 is clinically useful in the therapy of acute pancreatitis.
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PMID:Pharmacological studies of FUT-175, nafamostat mesilate. V. Effects on the pancreatic enzymes and experimental acute pancreatitis in rats. 242 60


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