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)

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

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

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

Bovine enterokinase (enteropeptidase) activates trypsinogen to trypsin at pH 8.0. In the presence of chicken ovomucoid, a stable complex of ovomucoid-trypsin is produced, inactivating trypsin and eliminating autoactivation of trypsinogen. The molecular size of trypsin (24,000 Da) is increased twofold on forming the ovomucoid-trypsin complex (52,000 Da). Size-exclusion chromatography on a Toya Soda TSK G2000SW column in an HPLC system and with computer-assisted analyses gives a direct quantitative determination of the amount of substrate (trypsinogen) and product (ovomucoid-trypsin). The rate of disappearance of substrate is equal to the rate of formation of product in agreement with kinetic theory. The simultaneous determination of both rates increases the reliability of the assay. The HPLC assay has an extended linear range for the velocity of the activation process as a function of enzyme concentration. The assay is reliable and accurate for highly purified preparations, samples at different steps in the purification scheme, and for a direct assay of the intestinal contents. The assay should be useful in clinical analyses.
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PMID:A direct high-performance liquid chromatography assay of the enzymatic activity of enterokinase (enteropeptidase). 271 84

An enterokinase (Enteropeptidase, EC. 3.4.21.9) has been described in the pharate adult of Glossina mositans morsitans. The enzyme is present in pharate adults, 21 days after pupation. It activated commercial crystalline bovine trypsinogen to trypsin. It showed affinity for concanavalin A bound to sepharose and was reversibly sensitive to boiling at pH 6.0. The apparent molecular weight, as determined by gel permeation on sepharose 6B-CL, suggests self-aggregation or an association with a large molecule (M.Wt. approximately equal to 2.5 X 10(6)).
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PMID:An enterokinase in the gut of pharate adult of Glossina morsitans morsitans Westwood (Diptera: Glossinidae). 285 54

In the 1950's, the specific cleavages of the peptide bonds occurring in bovine cationic chymotrypsinogen and trypsinogen were among the first examples of limited proteolyses. According to the split bond(s), the precursor is transformed into enzyme or different forms of zymogen or again into inert protein. The conversion of trypsinogen into trypsin triggers the activations of all the other enzyme precursors of pancreatic juice. In the pancreas, several factors oppose trypsinogen autoactivation, whereas, in the duodenum, all the conditions are favorable for trypsinogen activation by enteropeptidase.
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PMID:Limited proteolyses in pancreatic chymotrypsinogens and trypsinogens. 314 4

A method--enzymoblotting--was developed for localizing various enzymes after electrophoretic separation, transfer to nitrocellulose, and incubation with specific substrates. As an application, the proteinases porcine trypsin (EC 3.4.21.4), bovine chymotrypsin (EC 3.4.21.1), porcine elastase (EC 3.4.22.11), and their zymogen forms from porcine pancreas homogenate were analyzed utilizing specific p-nitroanilide substrates. After agarose gel electrophoresis, transfer of the separated proteinases to a nitrocellulose membrane was performed by capillary diffusion for 30 min. After air-drying of the nitrocellulose membrane, it was incubated in the appropriate substrate solution for 60 min. N-alpha-Benzoyl-DL-arginine-para-nitroanilide HCl was used as a substrate for trypsin, N-benzoyl-L-tyrosine-para-nitroanilide and succinyl-L-phenylalanine-para-nitroanilide for chymotrypsin, and N-succinyl-L-alanyl-L-alanyl-L-alanine-para-nitroanilide for elastase. p-Nitroaniline, the product thus obtained, was diazotized with N-(1-naphthyl)ethylenediamine to a red azo dye, visible at the site of the proteinases on the nitrocellulose membrane. The results could be preserved at -18 degrees C. Zymogen forms of the pancreas proteinases were detected in a similar manner. They were converted to active proteinases in situ on the nitrocellulose membrane after preincubating the nitrocellulose membrane in the activation enzymes enteropeptidase or trypsin.
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PMID:Enzymoblotting: a method for localizing proteinases and their zymogens using para-nitroanilide substrates after agarose gel electrophoresis and transfer to nitrocellulose. 351 6

Zymogen activation is an important biochemical control process and has important physiological and pathological implications. We have simultaneously measured both procarboxypeptidase A, the enzyme precursor, and carboxypeptidase A, its active product, in serum by using an affinity resin and the synthetic peptide substrate N-(2-furanacryloyl)-L-phenylalanyl-L-phenylalanine. Serum procarboxypeptidase A is activated by trypsin, chymotrypsin, plasmin, subtilisin, or urokinase but not by thrombin or enteropeptidase. The molecular weight of the precursor is approximately 5000-10 000 greater than that of the active product. Both enzyme and precursor increase in serum in the course of pancreatic inflammation, but the degree of activation can vary up to 2000-fold, independent of the amount of precursor present. The existence of this pancreatic proteolytic precursor in serum opens new avenues for the investigation of zymogen activation and its regulation.
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PMID:Human serum procarboxypeptidase A. 634 78

Mouse pancreatic proteases were analyzed by one- and two-dimensional electrophoresis. Active proteases that existed in the luminal fluid were separated into at least eight bands in 8% polyacrylamide gel. Pancreatic proteases activated by intestinal extract were separated into at least seven bands. The mobilities of these bands were exactly the same as those of proteases in the luminal fluid except for those of the most cathodal band. Two kinds of trypsin (Try-I group and Try-II) and one kind of chymotrypsin (Chy-I) were determined by specific and nonspecific protease staining. Try-I group and Try-II were derived from different trypsinogens (Try G-I group and Try G-II), whereas Chy-I was derived from a single chymotrypsinogen (Chy G). Although Try G-II was activated by both intestinal extract and by bovine trypsin, Try G-I group activated only by intestinal extract. Intestinal-activating factors were analyzed by two-dimensional electrophoresis. Mouse enterokinase (enteropeptidase EC 3.4.4.8), which can activate bovine trypsinogen, had a slow mobility. In the intestine of the mouse there are several activating factors in addition to enterokinase. Although it is unclear what intestinal-activating factors can activate Chy G, there is a factor that can convert chymotrypsinogen into chymotrypsin directly. These data suggest that intestinal-activating factors play an important role in the activating mechanisms of mouse pancreatic zymogens.
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PMID:Electrophoretic analysis of pancreatic proteases and zymogen-activating factors in the mouse. 637 96

Two trypsin assay methods for the estimation of this enzyme in duodenal fluid from children have been compared. Assay results for a fluorometric method based on the use of N-carbobenzoxy-diglycyl-L-arginyl-2-naphthylamide hydrochloride (GANA) as the trypsin substrate were found to correlate well (r = 0.91, P less than 0.001) with those obtained with a much less sensitive titrimetric assay which used benzoylarginine ethylester hydrochloride (BAEE) as substrate. The higher sensitivity of the fluorometric assay has allowed accurate determination of trypsin activity in 10 microliter aliquots of duodenal fluid. This low volume requirement makes the assay suitable for studies on infants of all ages and conserves duodenal fluid for use in other investigations often warranted during the assessment of childhood malabsorption. The fluorometric assay has also been used to monitor the separation of enteropeptidase from trypsin(ogen) by chromatography on Sephacryl S-200 in samples of duodenal fluid from two children. Different proteolytic pathway deficiencies were confirmed in these children.
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PMID:Fluorometric microassay of trypsin and enteropeptidase in children--comparison with a titrimetic assay. 639 83


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