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)

1. Rat skeletal muscle was homogenized in 0.05M-Tris/HCl, pH 8.5, containing 1M-KCl. Myofibrillar proteins were precipitated by addition of (NH4)2SO4 (33% saturation). 2. The alkaline proteolytic activity that was precipitated with the myofibrillar proteins was solubilized with trypsin (conjugated to Sepharose) and further purified by affinity chromatography, ion-exchange chromatography and gel filtration. 3. The purified enzyme migrates as a single band in polyacrylamide-disc electrophoresis, and has optimum hydrolytic activity with azocasein and [14C]haemoglobin as substrates at pH 9.4 and 9.6 respectively. Its apparent molecular weight, as determined by gel filtration on Sephadex G-75, is 30800. 4. The purified alkaline proteinase is strongly inhibited by equimolar amounts of soya-bean trypsin inhibitor and ovomucoid, whereas di-isopropyl phosphorofluoidate and alpha-toluenesulphonyl fluoride have no effect. On the other hand N-ethylmaleimide and p-chloromercuribenzoate have inhibitory effects on the enzyme activity. 5. Bivalent metal ions (Fe2+, Co2+, Zn2+, Mg2+, Mn2+) diminish the proteolytic activity, at 1mM concentrations. Ca2+ ions and the metal-ion-chelating agent EDTA are without effect on enzyme activity. 6. The enzyme is part of the alkaline proteolytic activity that appears to be associated with myofibrillar proteins.
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PMID:Purification and some properties of an alkaline proteinase from rat skeletal muscle. 2 72

A trypsin inhibitor was isolated from beans of Phaseolus vulgaris, cultivar. Kintoki, and the specific activity increased 200 times as high as that of the crude extract. It was homogeneous on several electrophoreses and the molecular weight was about 13,000. The amino acid composition was characterized by high ratios of cystine, aspartic acid, and serine. It inhibited trypsin in a molar ratio of 1 : 1 and alpha-chymotrypsin in a molar ratio of 2 : 1. It, however, inhibited neither pepsin nor pronase. It was relatively stable to heat treatment in the acidic medium, but not in the alkaline medium. Neither pepsin nor pronase destroyed the inhibitory function.
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PMID:The isolation and characterization of a trypsin inhibitor from Kintoki bean (Phaseolus vulgaris). 2 95

1. The solubilization and partial purification of a proteinase from the intestinal smooth muscle of rats fed on protein-free diets are described. 2. It has a mol.wt. of about 33000 and it is stable over a narrow pH range. 3. From its susceptibility to known modifers of proteolytic enzymes, it appears to be a serine proteinase of a trypsin-like nature. Active-site titration with soya-bean trypsin inhibitor shows that the concentration of proteinase was about 3 microgram/g wet wt. of intestinal smooth muscle. However, the muscle proteinase demonstrates a marked ability for inactivating enzymes in their native conformation at neutral pH. It is about 100 times more efficient than pancreatic trypsin when the inactivating activities are compared on an approximately equimolar basis. 4. Inactivation of the substrate enzymes is accompanied by limited proteolysis, as demonstrated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. 5. An endogenous inhibitor was separated from the proteinase by fractionation with (NH4)2SO4. 6. Contamination of the muscle tissue by lumen, mucosal or blood proteinases and inhibitors is shown to be unlikely. 7. A role for the neutral trypsin-like proteinase in initiating the degradation of intracellular enzymes is considered.
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PMID:The inactivation of native enzymes by a neutral proteinase from rat intestinal muscle. 2 34

1. Amylase (EC 3.2.1.1) activity found in the intestinal tract of chicks posterior to the stomach is of endogenous origin, as amylase in the food is inactivated by the low pH in the stomachs. 2. Ingestion of raw soya-bean diet (RSD) or of heated soya-bean diet (HSD) supplemented with trypsin inhibitors induced higher amylase activites in the lower part of the small intestine and caecum as compared with HSD. 3. Ingestion of RSD after ligation at the end of the duodenum, end of the ileum or one of the cacea, or injection of soya-bean trypsin inhibitor into a aligated caecum, indicated that there is no amylase synthesis by the intestinal tract cells or microflora as a response to the presence of RSD or trypsin inhibitors. 4. It seems that amylase found in the digestive tract of the chick is of pancreatic origin and the RSD or trypsin inhibitors induce higher pancreatic amylase secretion than HSD which (the additiona amylase) accumulates mainly in the caeca.
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PMID:The level and origin of amylase (EC 3.2.1.1) in the digestive tract of chicks receiving trypsin inhibitors in their diet. 2 57

Reversible unfolding of bovine chymotrypsinogen A in 2H2O either by heating at low pH or by exposure to 6 M guanidinium chloride results in the exchange of virtually all the nitrogen-bound hydrogens that give rise to low-field 1H NMR peaks, without significant exchange of the histidyl ring Cepsilon1 hydrogens. These preexchange procedures have enabled the resolution of two peaks, using 250-MHz correlation 1H NMR spectroscopy, that are attributed to the two histidyl residues of chymotrypsinogen A. Assignments of the Cepsilon1 hydrogen peaks to histidine-40 and -57 were based on comparison of the NMR titration curves of the native zymogen with those of the diisopropylphosphoryl derivative. Two histidyl Cepsilon1 H peaks were also resolved with solutions of preexchanged chymotrypsin Aalpha. The histidyl peaks of chymotrypsin Aalpha were assigned by comparison of NMR titration curves of the free enzyme with those of its complex with bovine pancreatic trypsin inhibitor (Kunitz). The NMR titration curves of histidine-57 in the zymogen and enzyme and histidine-40 in the zymogen exhibit two inflections; the additional inflections were assigned to interactions with neighboring carboxyl groups: aspartate-102 in the case of histidine-57 and aspartate-194 in the case of histidine-40 of the zymogen. In bovine chymotrypsinogen A in 2H2O at 31 degrees C, histidine-57 has a pK' of 7.3 and aspartate-102 a pK' of 1.4, and the histidine-40-aspartate-194 system exhibits inflections at pH 4.6 and 2.3. In bovine chymotrypsin Aalpha under the same conditions, the histidine-57-aspartate-102 system has pK' values of 6.1 and 2.8, and histidine-40 has a pK' of 7.2. The results suggest that the pK' of histidine-57 is higher than the pK' of aspartate-102 in both zymogen and enzyme. A significant difference exists in the structure and properties of the catalytic center between the zymogen and activated enzyme. In addition to the difference in pK' values, the chemical shift of histidine-57, which is highly abnormal in the zymogen (deshielded by 0.6 ppm), becomes normalized upon activation. These changes may explain part of the increase in the catalytic activity upon activation. The 1H NMR chemical shift of the Cepsilon1 H of histidine-57 in the chymotrypsin Aalpha-pancreatic trypsin inhibitor (Kunitz) complex is constant between pH 3 and 9 at a value similar to that of histidine-57 in the porcine trypsin-pancreatic trypsin inhibitor complex [Markley, J.L., and Porubcan, M. A. (1976), J. Mol. Biol. 102, 487--509], suggesting that the mechanisms of interaction are similar in the two complexes.
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PMID:Zymogen activation in serine proteinases. Proton magnetic resonance pH titration studies of the two histidines of bovine chymotrypsinogen A and chymotrypsin Aalpha. 3 98

A trypsin inhibitor was purified from the tubers of Colocasia antiquorum. The inhibitor acted on bovine trypsin, human trypsin and weakly on bovine chymotrypsin. The inhibitor, which had a molecular weight of 40 000, contained trace amounts of carbohydrates. The purified inhibitor was stable over a pH range of 2.0--12.0 and was more thermostable than the crude preparations. Trinitrobenzene sulphonate treatment resulted in the inactivation of the inhibitor. Chymotrypsin, pepsin and pronase digested the inhibitor. Pretreatment with trypsin at neutral pH resulted in the partial loss of antitryptic activity, whereas treatment at pH 3.7 led to complete inactivation. Evidence for the formation of a trypsin-inhibitor complex at pH 7.6 is provided. During the plant growth, in the early phase (0--40 days) there was a gradual increase in protein content and in antitryptic activity. The middle phase (40--55 days) was characterized by a rapid fall and abolition of the antitryptic activity and a diminution in protein content in the tubers. The immature tubers had low antitryptic activity compared to the mature ones. Mild heat treatment caused a sharp rise in antitryptic activity in the extracts of immature tubers but not with the mature tuber preparations.
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PMID:Natural plant enzyme inhibitors. VI. Studies on trypsin inhibitors of Colocasia antiquorum tubers. 3 37

When the jelly-less eggs removed from the most cephalic region of the oviduct (pars recta) of the toad Bufo arenarum were inseminated at a high sperm concentration, high frequencies of fertilization were obtained. On the other hand, control eggs removed from the pleuroperitoneal cavity (coelomic eggs)) were neither fertilized upon insemination under identical conditions, nor with the water extract of the jelly. Under these inseminating conditions, however, a high frequency of fertilization was obtained when coelomic eggs were preincubated in the presence of the fluid secreted by the epithelium of the pars recta or of an extract prepared from pars recta homogenate. Experimental evidence is presented showing that the component responsible for this effect acts on the vitelline envelope of the egg, increasing its susceptibility to sperm lysin. It is probable, therefore, that it induces successful fertilization of coelomic eggs by making the vitelline envelope more easily penetrable by sperm. The active factor was partially purified by Sephadex chromatography. The product obtained was of high activity and, as judged by its inhibition with soybean trypsin inhibitor and lima-bean trypsin inhibitor, it is likely to be a trypsin-like enzyme. The molecular weight of the factor was estimated to be 47000 by Sephadex chromatography. Secretion of the pars recta factor is hypophysis-dependent and its activity is not influenced by pH within the range testes (6.0--8.4).
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PMID:A trypsin-like oviducal proteinase involved in Bufo arenarum fertilization. 3 7

After exposure to extracts from blood fed A. aegupti cultured ookinetes of P. gallinaceum were damaged to various, defined extents. Immature ookinetes were found to be more sensitive to damage than mature ones. The damage was dependent on the digestion time after which the Aedes extracts had been prepared and could be correlated with the proteolytic activity in the extracts. Control experiments demonstrated that the factors responsible for damage were neither present in unfed mosquitoes nor in blood alone and that the damage was not a result of osmotic stress. After the treatment of the Aedes extracts with lima bean trypsin inhibitor the ookinete damage was much less, indicating that the Aedes trypsin was the major agent of damage. These results were supported by experiments in which the tryptic activity of the extracts was eliminated by thermal denaturation. It is concluded that in the mosquito midgut most of the ookinetes are damaged by digestive enzymes and that this is one factor leading to the discrepancy between the number of ingested macrogametocytes and the number of oocysts which is usually found in nature. It seems that the only ookinetes which have a chance of surviving are those which develop in the centre of the blood clot, away from the site of enzyme action.
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PMID:In vitro damage of cultured ookinetes of Plasmodium gallinaceum by digestive proteinases from susceptible Aedes aegypti. 4 87

The role of trypsin in the elicitation of G-banding on human chromosomes was studied in two separate laboratories. Enzyme activity and ability of trypsin to chelate calcium were manipulated by dilution of the treatment solution, and by inhibition with diisopropylphosphofluoridate, diphenylcarbamyl chloride, or soybean trypsin inhibitor. In all cases, chromosomes were affected in proportion to the enzyme activity of the treatment solution rather than the ability of the solution to bind calcium. It is concluded that calcium chelation is not sufficient to explain G-banding trypsin, but that proteolytic activity is required.
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PMID:The role of trypsin in the pre-treatment of chromosomes for Giemsa banding. 5 76

The present study using immunologic methodology confirms previous observations from this laboratory of an absence of a protease component with arginine esterase activity in plasma of patients with cystic fibrosis. In this study, the pooled plasma from control individuals was activated and partially purified after adsorption on columns of soybean trypsin inhibitor conjugated to Sepharose 4B followed by elution with benzamidine. The fraction was further purified by isoelectrofocusing on polyacrylamide gels. Proteins around the pI range of 5.5 were eluted and utilized to prepare an antiserum. Immunoelectrophoresis of activated plasma samples from control subjects and patients with cystic fibrosis was performed utilizing the antiserum. In controls, four precipitin arcs with residual esterase activity were observed, whereas only three were seen in plasma from patients with cystic fibrosis. Double gel diffusion experiments using specific antisera ruled out the presence of trypsin, chymotrypsin, plasminogen, prothrombin, C1 esterase, alpha one-trypsin inhibitor, and inter-alpha-trypsin inhibitor in the concentrated benzamidine eluate. The antisera to alpha two-macroglobulin gave an immunoprecipitate which was readily stained for proteolytic activity. On immunoelectrophoresis, the alpha two-macroglobulin precipitin band corresponded to the band absent in plasma of patients with cystic fibrosis. In contrast, the alpha two-macroglobulin levels were similar in plasma of control subjects and patients with cystic fibrosis. Using the antiserum to the protein fractith proteolytic activity could be demonstrated in control plasma. One specific enzyme-active "rocket" was absent in plasma of patients with cystic fibrosis. In a double blind study of 15 control samples and 15 samples from patients with cystic fibrosis, a specific "rocket" was shown to be present in 13 control samples and absent in 14 cystic fibrosis samples. alpha two-Macroglobulin was determined by both an immunologic procedure and by its trypsin binding (trypsin protein esterase concentration). The ratio of the immunologic assay to the biologic activity assay was 90 for the normal plasma samples and only 65 for cystic fibrosis samples.
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PMID:Absence of an alpha two-macroglobulin-protease complex in cystic fibrosis. 6 Jul 35


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