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)

Tissue plasminogen activator was treated with Sepharose-bound trypsin or chymotrypsin. Trypsin rapidly converted the one-chain activator to the two-chain form. This caused a marked increase in the amidolytic activity, while plasminogen activation initially increased but then decreased again. SDS/polyacrylamide gel electrophoresis in combination with [3H]diisopropylfluorophosphate active-site labeling revealed that after the conversion to the two-chain activator a minor cleavage occurred in the B chain, while the A chain was substantially degraded. Chymotrypsin caused a marked decrease in both amidolytic activity and plasminogen activation. SDS/polyacrylamide gel electrophoresis under reducing conditions revealed that two pairs of new bands had appeared, with Mr or about 50,000/52,000 and 17,000/20,000 respectively. N-terminal sequence analysis identified cleavage sites at peptide bonds 420-421 and 423-424. These bonds are located in a region of the activator which is homologues to the segments of trypsin and chymotrypsin, where autocatalytic cleavages occur during their activations. However, treatment of two-chain activator with chymotrypsin had markedly less effect on plasminogen activation and amidolytic activity. By treatment of samples of chymotrypsin-digested one-chain activator with plasmin, amidolytic activity could be largely restored. Thus, chymotrypsin may, by cleaving bonds 420-421 and 423-424, convert the active one-chain activator into an 'inactive' zymogen, which is again 'activated' by plasmin cleavage.
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PMID:Proteolytically induced variations in the enzymatic properties of tissue plasminogen activator. Activations, inactivations and reactivations. 294 92

The turnover of basement membrane macromolecules in injured skeletal muscle has not been studied in contrast to other biologic systems undergoing remodeling. Plasminogen activators and other neutral proteases that are able to degrade these basement membrane macromolecules are secreted by cultured muscle cells. We sought to determine if locally released plasminogen activators could act on basement membrane components. Such degradation might be implicated in the disadhesion of nerve from muscle after motor nerve denervation. To test this hypothesis, we first undertook a study of the sensitivity of muscle extracellular matrix antigens following in vitro exposure to various proteases on frozen muscle sections. Fibronectin was found to be most sensitive, followed by type IV collagen and laminin. Of serine proteases, trypsin was the most active but was not selective, digesting matrix and sarcoplasmic components alike in less than 30 min. Purified urokinase was inactive unless plasminogen (also inactive alone) was previously added to tissue sections, at which time only matrix antigens were digested. Little if any observable degradation of sarcoplasmic proteins took place under these conditions. Using a highly sensitive and selective assay, we found that plasminogen activators were present in muscle tissue and increased 8- to 10-fold after 10 days of denervation. Using an extract of denervated muscle in the presence of plasminogen, we observed degradation of matrix antigens. No degradation was observed with control muscle extract. We next evaluated the degradation of these antigens in denervated muscle during a temporal study. The results, analyzed by quantitative image analysis, indicates that with increasing time after denervation a marked decrease of fibronectin and type IV collagen, followed by laminin occurred but, again, only in the present of plasminogen. These results indicate a selective sensitivity of basement membrane antigens of muscle and a role for plasminogen activators in the degradation of these adhesive basement membranes macromolecules after denervation.
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PMID:Degradation of muscle basement membrane zone by locally generated plasmin. 294 9

Combined assessment of some parameters related to hemostasis and blood plasminogen-plasmin and kinin systems and their inhibitors was carried out in 150 patients with stable angina and postinfarction cardiosclerosis during rationed exercise (bicycle ergometry). Patients with low stress tolerance showed considerably activated kallikrein-kinin system, depressed total fibrinolytic activity, elevated trypsin and enhanced platelet adhesive properties, while their inhibitor activities were reduced. Postmyocardial infarction patients showed a more dramatic increase in free kinins, coupled with depressed inhibitor proteinases and kininase activities, and a slight increment in fibrinolytic activity, as compared to patients having to history of myocardial infarction. The magnitude of free kinins and trypsin rise and kininase fall increases with age in these patients.
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PMID:[Various indicators of hemostasis, plasminogen-plasmin and kinin systems of the blood and their inhibitors in patients with stable angina pectoris and post-infarction cardiosclerosis after graded physical load]. 295 50

Previous studies of alkali burns have provided evidence for an important role of the plasminogen activator (PA)/plasmin system in corneal ulceration. Current studies have utilized a sensitive, plasminogen-dependent fluorescent assay to demonstrate that PA is present mostly in a latent (trypsin- or plasmin-activatable) form (proactivator) in cultures of rabbit corneal epithelial cells or normal corneas. Cultures of ulcerating corneas demonstrate only active PA early in organ culture, whereas, latent PA levels increase later in culture. Thus, ulceration is correlated with the apparent conversion of latent to active PA. Moreover, profiles of proactivator and latent collagenase and of active PA and active collagenase in vitro, respectively, are similar, suggesting that activator and collagenase are under coordinate control. Cultures of normal epithelial cells and nonulcerating corneas contain PA molecular weight species of 72,000 and 46,000 MW, and ulcer corneas, species of 72,000, 46,000, and 35,000 MW. Double-diffusion analysis indicates that rabbit epithelial cells, fibroblasts, and ulcer corneas produce urokinase (UK)-like PA; and human cornea extracts and tears also contain PA immunoreactive with anti-UK antibodies. The existence of PA in a latent form identifies another level of regulation in the cascades that lead to stromal ulceration.
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PMID:Latent and active plasminogen activator in corneal ulceration. 298 39

Certain group A streptococci demonstrate surface receptors that bind selectively to the key fibrinolytic enzyme, plasmin. These bacteria show no reactivity with the zymogen protein plasminogen or with other serine class proteases, such as trypsin or urokinase. Bacterium-bound plasmin retains its ability to cleave synthetic substrates and its ability to hydrolyze a fibrin clot. The bacterium-bound plasmin is not effectively regulated by its physiological regulator, alpha 2-plasmin inhibitor. This study is the first report of a bacterium-associated receptor for plasmin.
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PMID:Identification of a specific receptor for plasmin on a group A streptococcus. 303 53

An immunosorption procedure is developed for quantitative estimation of plasminogen activator of the urokinase type in human blood serum. The procedure involved sorption of urokinase from blood serum using polyclonal antibodies to the enzyme as a sorbent at the first step of the assay. Concentration of the immunoreactive urokinase in a preparation correlated with the absorbed fraction of standard urokinase added into the sample at the second step after washing of the sorbent. The residual fibrinolytic activity of the standard dose was detected in fibrin-agar gel. Results of the assay were not altered in presence of high molecular (bovine blood serum, soy bean inhibitor of trypsin) and low molecular inhibitors of urokinase (benzamidine, arginine, epsilon-aminocapronic acid). Sensitivity of the assay constituted 0.05 IU/sample (2.5 IU/ml, 0.25 ng/sample) and might be increased by a decimal order after addition of plasminogen into fibrin-agar gel. Content of urokinase in blood serum of healthy men (21-48 years old) was equal to 166 +/- 8.9 IU/ml with individual variations from 107 to 260 IU/ml.
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PMID:[Immunosorption method of urokinase determination]. 307 Sep 32

Native one-chain tissue plasminogen activator (t-PA) was rapidly converted to the two-chain form by trypsin-Sepharose cleavage. This caused an increase in the amidolytic activity on low molecular weight peptide substrates, while plasminogen activation in the presence of fibrin markedly decreased. Cleavage sites were identified by N-terminal sequence analysis of reduced and carboxymethylated peptides. In the B-chain, the expected cleavage at Arg278-Ile279 was identified. Furthermore, a specific cleavage site was found at Arg302-Ser303, 24 amino acids from the N-terminus of the B-chain. The peptide released by this cleavage (designated B1-24) remained associated with the activator molecule by strong noncovalent interactions but could be dissociated under denaturing conditions (4 mol/L of guanidine hydrochloride), leading to a 20-fold decrease in amidolytic activity. Addition of purified B1-24 peptide to t-PA treated in this manner restored the activity in a concentration-dependent way. In contrast to trypsin, cleavage of the single-chain t-PA molecule with endoproteinase Lys-C generated a two-chain form of the activator, without simultaneous increase in the amidolytic activity. By sequence analysis, a major cleavage was identified at Lys280-Gly281, two residues into the B-chain. Together, the results presented provide additional information on the one-chain to two-chain conversion of t-PA and the role of the free N-terminus of the B-chain.
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PMID:Proteolytic modification of tissue plasminogen activator: importance of the N-terminal part of the catalytically active B-chain for enzymatic activity. 314 3

A plasminogen activator inhibitor was purified to apparent homogeneity from conditioned media of U138 cells. The inhibitor is a glycoprotein with a pI of 5.4 and an apparent molecular weight of 45,000. The inhibitor forms sodium dodecyl sulfate-stable complexes with plasminogen activators and trypsin but not with plasmin, thrombin, or pancreatic kallikrein. Some biochemical and immunochemical characteristics of the U138 inhibitor distinguish it from other known plasminogen activator inhibitors. The expression of this inhibitor by U138 cells could be modulated by incubation in phorbol myristate acetate, interleukin-1, tumor necrosis factor, and gamma interferon, but not in beta interferon. Thus, the expression of the plasminogen activator inhibitor can be influenced by biological response modifiers known to be active in the brain and in the neural response to inflammatory stimuli. Therefore, this inhibitor, along with protease nexin, may be involved in brain development and regulation.
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PMID:Purification and partial characterization of a plasminogen activator inhibitor from the human glioblastoma, U138. 314 98

Plasminogen Activator (PA) and its response to glucocorticoids and androgens was studied in viable rat thymocytes in suspension. PA was measured by its ability to convert plasminogen to plasmin, and the formed plasmin determined by cleavage of 14C-labeled globin. Using this functional assay, PA was found to be associated with the outer surface of thymic cells, and only negligible activity recovered from the incubation medium. Rat thymocytes also contain cytoplasmic and nuclear inhibitor(s) of the serine proteases plasmin, trypsin, chymotrypsin and thymic PA. Release of these inhibitors prevented determination of thymic PA activity in presence of lysed cells. The specific activity of PA in thymocytes isolated from adrenalectomized-castrated rats did not differ significantly from the specific activity associated with cells from intact animals. Furthermore, treatment of adrenalectomized-castrated rats with 0.1 mg of dexamethasone/kg for 2 days induced thymic involution without affecting thymic PA activity. These observations suggest that PA activity of thymocytes is not involved in glucocorticoid-mediated thymic involution.
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PMID:Plasminogen activator and protease inhibitor activities in isolated rat thymocytes. 315 48

Human and rabbit kidney and urine contain an inactive form of kallikrein. Studies on the mRNA sequence suggested that the active form of the enzyme and the propeptide are linked by a peptide bond between a basic and hydrophobic amino acid. We studied the activation of prokallikrein by serine proteases and a neutral metalloproteinase, thermolysin, because serine proteases cleave the peptide chain after a basic amino acid and thermolysin before a hydrophobic amino acid. The activity of kallikrein was measured by RIA and with a fluorogenic peptide substrate. Trypsin was used as a standard reference activator. We found that human plasmin and plasminogen, activated by urokinase, activate prokallikrein. Pronase coupled to Sepharose also enhanced the activity of the renal kallikrein zymogen. On a molar basis, thermolysin was a more effective activator of prokallikrein than trypsin. The activation by thermolysin was blocked by the inhibitor phosphoramidon, but not by DFP or SBTI. These experiments indicate that, in addition to serine proteases, neutral metalloproteases of tissues may activate prokallikrein.
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PMID:Activation of human and rabbit prokallikrein by serine and metalloproteases. 315 29


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