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:3.4.21.1 (chymotrypsin)
10,938 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We recently demonstrated that chymotrypsin substrate analogues inhibit receptor-mediated insulin internalization in isolated rat adipocytes. In this study, the effect on glucose transport of inhibiting insulin internalization with these agents was examined. Glucose transport was assayed by measuring [3H]-2-deoxyglucose uptake, and internalized insulin was measured after rapidly dissociating surface-bound insulin with an acidic buffer. The chymotrypsin substrate analogue N-acetyl-Tyr ethyl ester inhibited insulin internalization by 85% while increasing surface-bound insulin by 80-110%. Under these conditions, ATP levels were minimally altered, and basal glucose transport was unchanged; however, insulin-stimulated glucose transport was decreased by 86%. The inhibition of insulin-stimulated glucose transport was not overcome by supramaximal concentrations (400 ng/ml) of insulin. When insulin internalization and insulin-stimulated glucose transport were measured in the presence of increasing concentrations of N-acetyl-Tyr ethyl ester (0.1-1 mM), a strong and highly significant correlation (r = .97, P less than .001) was found between inhibition of insulin internalization and inhibition of insulin-stimulated glucose uptake. Fragments of N-acetyl-Tyr ethyl ester that do not inhibit insulin internalization were also without effect on insulin-stimulated glucose transport. In addition to N-acetyl-Tyr ethyl ester, four other chymotrypsin substrate analogues that are effective inhibitors of insulin internalization also markedly inhibited insulin-stimulated glucose transport. These results indicate that insulin internalization and insulin-stimulated glucose transport share a common postbinding step in adipocytes and that this step is inhibitable by chymotrypsin substrate analogues.
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PMID:Insulin-stimulated glucose transport and insulin internalization share a common postbinding step in adipocytes. 354 51

In this study, we purified insulin-like substance (ILS) in the human pancreatic juice by the combined use of affinity chromatography and radioimmunoassay (RIA). The amino acid sequence of ILS in the N-terminal region is the same as that of human insulin. The influence of the enzymes present in the pancreatic juice on the RIA procedure, was examined. Trypsin, chymotrypsin and amylase showed steep influences on radioactivity. The addition of enzyme inhibitors could not reduce pseudo-activity, but the elimination of enzymes in the pancreatic juice by ultrafiltration with the Mole-Cut (Millipore, Japan) resulted in a lowering of the pseudo-insulin activity. Affinity chromatography on Sepharose 4B coupled with anti-porcine insulin was used to capture ILS. ILS was eluted by 1 M acetic acid from the affinity column monitoring pH and the insulin activity by RIA. The amino acid sequences of two components of ILS in amino terminal region were Phe-Val and Gly-Ile-Val. This indicates that ILS obtained from human pancreatic juice was the insulin derived from endocrine secretion of pancreas.
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PMID:Identification of insulin in the human pancreatic juice. 354 48

Fibrinogen-platelet interaction was studied in suspensions of platelets obtained from patients with uncontrolled diabetes mellitus of long duration and from control individuals. Fibrinogen binding sites were exposed by stimulating platelets with ADP or with chymotrypsin. There was no significant difference in fibrinogen mediated aggregation between ADP-stimulated platelets of 80 control and 47 diabetic subjects. The Km values for fibrinogen mediated aggregation of ADP-stimulated platelets obtained from control and diabetic donors were 1.39 +/- 0.13 X 10(-7)M and 1.44 +/- 0.13 X 10(-7)M; the Vmax values (expressed in arbitrary light transmission units) were 87.8 +/- 3.14 and 92.8 +/- 4.5 (mean +/- S.E.M.). The analysis of variance showed no significant relationship between Km, Vmax, age and sex in control group; in patient group there was no significant relationship between Km, Vmax, age, sex, type of diabetes, presence of vascular complications and type of treatment (insulin and/or oral hypoglycemic agents). Fibrinogen mediated aggregation of chymotrypsin-treated platelets showed similar pattern in 25 control and in 25 diabetic donors. In 24 normal individuals and in 24 diabetic patients Scatchard analysis revealed 48,820 +/- 5350 fibrinogen binding sites per one normal platelet (Kd = 4.7 X 10(-7)M) and 45,350 +/- 4663 sites per one diabetic platelet (Kd = 3.5 X 10(-7)M). Our data suggest a normal pattern of interaction between fibrinogen and fully activated platelets of diabetic subjects.
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PMID:Fibrinogen interaction with platelets of diabetic subjects. 360 36

Three cases of clinically benign pancreatic papillary cystic tumors in young female patients were studied by immunohistochemistry and electron microscopy in order to define the cellular nature of this type of neoplasm. Two of the tumors showed focal cytokeratin- and desmoplakin-positivity as evidence of focal epithelial differentiation, while the tumor cells were in all cases positive for vimentin--the intermediate filament protein typical of (but not specific for) mesenchymal cells. Electron microscopy showed some cell-cell junctions, but there was no evidence of acinar or islet cell differentiation. The tumors were at least focally positive for neuron-specific enolase, and small clusters of polypeptide hormone immunoreactive cells were present in all cases (glucagon 3/3, somatostatin 2/3, insulin 2/3). However, the tumors were negative for synaptophysin and neurofilament proteins, unlike most islet cell tumors. Trypsin and chymotrypsin immunoreactivity was found in all tumors, but because many nonpancreatic carcinomas were also positive, we doubt whether these two enzyme proteins can act as specific markers for pancreatic acinar cell differentiation. Two of the tumors that were studied immunohistochemically for the presence of nuclear estrogen receptors, were negative. Therefore no proof of the suggested hormone dependence of this tumor could be obtained. We conclude that papillary cystic tumor is a neoplasm of primitive pancreatic epithelial cells, that may exhibit focal endocrine cell differentiation.
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PMID:Papillary cystic tumor of the pancreas. An analysis of cellular differentiation by electron microscopy and immunohistochemistry. 367 83

Human bone cells isolated from femoral heads were cultured in BGJb medium containing bovine serum albumin (100 micrograms/ml), insulin (1 microgram/ml) and epidermal growth factor (10 ng/ml), and the conditioned medium collected. The medium was concentrated, chromatographed using HPLC gel filtration (TSK 2000 SW), and assayed for mitogenic activity using [3H]thymidine incorporation into embryonic chick calvarial cells. The conditioned medium contained mitogenic activity which eluted with a different elution time than insulin or epidermal growth factor. Characterization of this activity suggests that it was due to human skeletal growth factor (SGF), a mitogen which had been previously isolated from human bone matrix. Common properties include: stimulation of DNA synthesis in cultured embryonic chick calvarial cells, competition with human SGF for binding to anti-SGF antibodies, elution from HPLC gel filtration as a large factor (Mr 100,000) under native conditions but as a small factor (Mr 10,000) under dissociative conditions (4 M guanidine HCl), elution time on HPLC reverse-phase chromatography (small SGF), inactivation by dithiothreitol, stability to heat, acidic or alkaline conditions and inactivation by trypsin and chymotrypsin. These observations provide evidence that human bone cells produce SGF. Conditioned medium from human skin cell cultures also contained mitogenic activity. However, the activity was less than that from bone cells and did not cross-react with the rat anti-SGF antibodies.
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PMID:Skeletal growth factor is produced by human osteoblast-like cells in culture. 377 45

Solid and papillary epithelial neoplasms of the pancreas from six female patients were studied using immunohistochemistry and electron microscopy to define better their histogenesis. The tumors ranged in diameter from 5 to 15 cm (average: 9 cm), and, on cross section, most had areas of hemorrhage and necrosis, sometimes extensive. Microscopically, there was a solid and pseudopapillary pattern, with tumor cells typically having ovoid nuclei with delicate folding and indistinct nucleoli. Of note were the following: a relatively low mitotic rate (range: 0-6/20 hpf), the presence of hyaline globules (four of six cases), and collections of foam cells (three of six cases). Staining for cytoplasmic argyrophil granules was negative in each case. Ultrastructurally, the solid and papillary epithelial neoplasms of the pancreas showed evidence of acinar or ductular differentiation. Two contained zymogen granules, one had intermediate filaments (probably keratin), and three had abundant rough endoplasmic reticulum and mitochondria. Immunostaining was positive for chymotrypsin (six of six cases), trypsin (four of six), and amylase (three of six). None was positive for alpha-1-antitrypsin, neuron-specific enolase, pancreatic polypeptide, gastrin, glucagon, somatostatin, or insulin. The findings support an origin from exocrine pancreas, and follow-up indicates a low rate of malignancy, with local recurrence in two of the six patients.
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PMID:Solid and papillary epithelial neoplasm of the pancreas. An ultrastructural and immunocytochemical study of six cases. 381 76

Transfection of genes into tissue culture cell lines has demonstrated that relatively short DNA sequences can allow expression of immunoglobulin, insulin and chymotrypsin genes in their appropriate cell types. A definitive test of cell-specific gene expression, however, requires testing genes in every possible cell type, an experiment performed easily by introducing the gene in question into the germ line of an animal. Transfer of intact genes into mice has demonstrated that a mouse immunoglobulin kappa gene is expressed specifically in B lymphocytes, a rat elastase I gene is expressed specifically in pancreas and a chicken transferrin gene is expressed preferentially in liver. Mouse metallothionein-growth hormone fusion genes introduced into mice are preferentially expressed in the liver, consistent with the expression of endogenous metallothionein genes, but initial experiments with beta-globin genes have not revealed proper regulation. To identify the DNA elements required for pancreas-specific expression of the rat elastase I gene, we joined the 5'-flanking region of this gene to the human growth hormone (hGH) structural gene and introduced the fusion gene into mice. Here we demonstrate that a fusion gene containing only 213 base pairs (bp) of elastase I gene sequence directs expression of hGH in pancreatic acinar cells.
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PMID:Specific expression of an elastase-human growth hormone fusion gene in pancreatic acinar cells of transgenic mice. 384 51

The indirect estimation of chymotrypsin by the tubeless ALTAB-test was performed in 17 patients with well-defined exocrine pancreatic insufficiency, 10 of them after operation for chronic pancreatitis. In comparison with 12 healthy subjects the test proved to be a non-expensive, certain and specific method for detection of moderate and severe exocrine insufficiency. It exists positive correlations with the secretin-pancreocymine-test and with the maximal stimulable secretion of insulin. Therefore the ALTAB-test after operations with modified anatomy of the gastro-intestinal tract especially is able to substitute extensive testing of secretion in screening and controlling of progression. The value of the 3-hour serum level of PABA corresponds to the urine output within 6 hours. There are such advantages like independence from the kidney-function, avoidance of incomplete urine-collection and a considerable reduced test-time too.
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PMID:[Modified ALTAB test in the diagnosis of exocrine pancreatic insufficiency]. 387 16

Thioltransferase was purified 650-fold from rabbit liver by procedures including acid treatment, heat treatment, gel filtration on Sephadex G-50, column chromatography on DEAE-cellulose, isoelectric focusing (pH 3.5-10) and gel filtration on Sephadex G-75. The final enzyme preparation was almost homogeneous in polyacrylamide gel electrophoretic analysis. Only one active peak with an apparent molecular weight (Mr) of 13,000 was detected by gel filtration on Sephadex G-50 and only a single protein band with a molecular weight of 12,400 was detected by sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Isoelectric focusing revealed only one enzyme species, having an isoelectric point (pI) of 5.3. The enzyme has an optimum pH about 3.0 with S-sulfocysteine and GSH as substrates. The purified enzyme utilized some disulfides including S-sulfocysteine, alpha-chymotrypsin, trypsin, bovine serum albumin, and insulin as substrates in the presence of GSH. The enzyme does not act as a protein : disulfide isomerase (the activity of which can be measured in terms of reactivation of randomly reoxidized soybean Kunitz trypsin inhibitor). The enzyme activity was inhibited by chloramphenicol, but not by bacitracin. The inhibition by chloramphenicol was non-competitive (apparent K1 of 0.5 mM). Thioltransferase activity was found in the cytosol of various rabbit tissues.
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PMID:Purification and some properties of rabbit liver cytosol thioltransferase. 401 39

1. With the aid of a coupled system involving glutathione reductase, the reaction of glutathione with the disulphide bonds of purified proteins has been studied. 2. Bovine serum albumin, conalbumin, lysozyme, trypsin inhibitors from egg white, lima bean and soya bean either did not react with glutathione or reacted only slightly. With these proteins reactivity was markedly increased by limited proteolysis. 3. Bovine and human gamma-globulins, fibrinogen and beta-lactoglobulin exhibited some reactivity (less than 15%) with glutathione and again this was increased by limited proteolysis. Pepsin, trypsin and chymotrypsin exhibited greater reactivity than the proteins previously mentioned. Di-isopropylphosphoryl-chymotrypsin exhibited less reactivity than chymotrypsin, suggesting that autolysis under the experimental conditions used contributed towards the reactivity of this protein. Proteolysis also increased the reactivity of these proteins. The three disulphide bonds of insulin were reduced by glutathione. 4. Above 35 degrees the disulphide bonds of serum albumin show a progressive increase in reactivity and at 55 degrees half of the bonds become accessible to glutathione. 5. From the results obtained with the proteins investigated, the conclusion reached is that the disulphide bonds of native proteins are structurally protected and do not react with glutathione under physiological conditions.
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PMID:The reactivity of the disulphide bonds of purified proteins in relationship to primary structure. 486 Apr 70


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