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)

Guinea pig alpha-macroglobulin was purified to apparent homogeneity by sequential chromatography on Sephacryl S-300, DEAE-cellulose, and hydroxyapatite. A molecular weight of 780,000 was obtained by equilibrium sedimentation. The preparation migrated as a single band of Mr = 180,000 in sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Rabbit antiserum raised against the final preparation partially cross-reacted with human and rat alpha-2-macroglobulins but not with rat alpha-1-macroglobulin. Guinea pig alpha-macroglobulin stimulated the amidolytic activity of trypsin towards a small substrate, but inhibited the proteolytic activity of trypsin towards remazol brilliant blue hide powder. When treated with trypsin or methylamine, four thiol groups per molecule were newly generated. The reaction with trypsin proceeded with at least at two different rates: half of the thiol groups were generated in a fast reaction and the remaining half in a slower reaction. On the other hand, such a two-step reaction was not detected in the reaction with methylamine. The methylamine-treated alpha-macroglobulin retained half the capacity to bind trypsin and its mobility in polyacrylamide gel under nondenaturing conditions remained virtually unchanged. These properties are in marked contrast to those reported for human alpha-2-macroglobulin, but resemble those of rat alpha-2- and mouse alpha-macroglobulins. The amidase activity of trypsin bound to guinea pig alpha-macroglobulin was impaired by soybean trypsin inhibitor to a much greater degree than that of trypsin bound to human or rat alpha-2-macroglobulin.
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PMID:Isolation and characterization of alpha-macroglobulin from guinea pig plasma. 242 4

A proton nuclear magnetic resonance (NMR) study is reported of human alpha-2-macroglobulin (alpha-2-M). It was observed that alpha-2-M, which consists of four identical subunits and has a molecular weight of 720,000, gives several sharp resonances. After cleavage of the "bait" region peptide with trypsin and subsequent removal of the peptide under a high salt condition, most of the sharp resonances disappeared, indicating that the sharp resonances observed in the native alpha-2-M originate from the amino acid residues in the bait region. Resonances due to the aromatic protons of the Tyr residue, which exists in the bait region, have been assigned on the basis of chemical shift. It was observed that the C3- and C5-H proton resonances for the Tyr residue are especially narrow, indicating that the side chain of the Tyr residue in the bait region is in a highly mobile state. Photochemically induced dynamic nuclear polarization experiments clearly show that the Tyr residue is actually exposed to the solvent. It was possible to identify resonances due to several His residues that are exposed to solvent. Other resonances, which probably originate from Arg residues in the bait region, were also observable in the conventional NMR spectra. On the basis of the present NMR data, we conclude that the bait region of the native alpha-2-M is highly flexible and exposed to solvent. On treatment of alpha-2-M with methylamine, no significant change has been detected in the NMR spectra observed in both the conventional and CIDNP mode.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Proton nuclear magnetic resonance study of human plasma alpha-2-macroglobulin. 243 25

Endocytotic internalization of alpha-galactosidase by cultured fibroblasts derived from a patient with Fabry's disease was achieved via receptor-mediated endocytosis of alpha-2-macroglobulin (alpha-2-M). alpha-galactosidase of coffee beans was conjugated to alpha-2-M when the latter was treated with trypsin. Internalization of the conjugate resulted in an increase of alpha-galactosidase activity in the crude cell extracts. The observed internalization was blocked by the presence of bacitracin, an inhibitor of binding between alpha-2-M and its receptor on the cell surface. When the cells were incubated at 4 degrees C with the conjugate, internalization was also inhibited. The alpha-galactosidase activity in the cells was saturated when the concentration of the conjugate in the medium was 40 micrograms/ml. Since non-conjugated alpha-galactosidase was not effectively internalized, the observed internalization of the conjugate was mediated by recognition of alpha-2-M by its receptor. The effective internalization of alpha-galactosidase described in this paper has a potential use in the enzyme replacement therapy of Fabry's disease.
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PMID:Endocytotic internalization of alpha-2-macroglobulin: alpha-galactosidase conjugate by cultured fibroblasts derived from Fabry hemizygote. 243 88

Synovial fluids of patients suffering from rheumatoid arthritis and osteoarthritis with effusions of the knees were examined. Different parameters were evaluated out of the synovial fluid (immunglobulins, Complement-1Q,-3,-4, haptoglobins, alpha-1-anti-trypsin, alpha-2-macroglobulin, transferrin, ceruloplasmin, rheumatoid factors, total count of cells, and ragocytes) and out of the plasma (blood sedimentation rate). The proteins were analysed by a nephelometricturbidimetric automatic centrifugal analyser. All parameters have been tested by valuable statistical methods and correlated to each other. The results worked out proved the reliability of the used test kits and apparative systems. Correlations within groups of parameters according to their formations (intra-and/or extraarticular) could not have been worked out in a way as it may be supposed. In contrast some parameters themselves are statistically different comparing rheumatoid arthritis and osteoarthritis. In general the results are on a higher level in the rheumatoid arthritis group. Using all parameters mentioned above the statistical differential diagnostic level is based on about 94%. If only blood sedimentation rate, total cell count and ragocytes are evaluated the level is based on 68%.
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PMID:[Differential diagnostic value of nephelometrically determined protein fractions in the synovial fluid]. 244 12

Alpha-2-macroglobulin (a2M) is a wide spectrum plasma inhibitor which functions by a unique mechanism and is a secondary inhibitor of coagulation and fibrinolytic enzymes. Human activated protein C (APC) is the central enzyme of a major regulatory system of coagulation and fibrinolysis. APC is primarily regulated (inhibited) by a specific plasma inhibitor. We undertook this study to investigate the role of a2M as a secondary inhibitor of APC. APC did not interact with a2M by any of the known mechanisms of interaction. APC failed to bind and form the classic proteinase-a2M complex as seen with similar serine proteases, thrombin and trypsin. APC also failed to cleave the a2M molecule. Experiments, using purified APC and either purified a2M or plasma, failed to demonstrate APC binding to a2M in gel filtration chromatography. No enzymatic activity of APC or radiolabeled APC was demonstrated in the a2M peak. Using an immuno-enzymatic assay (Harpel, J Biol Chem 260:4257, 1985) for an a2M and enzyme complex, the amount of APC bound to a2M was less than 3% of the added APC (non-specific binding only); whereas in similar experiments with thrombin, 75-86% of the added trypsin or thrombin bound. These data demonstrate that APC is one of a small number of unique serine proteases that do not interact with a2M. The absence of sequence homology between the a2M 'bait region' and the APC substrate cleavage sequences appear to be the reason APC is not inhibited by a2M.
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PMID:Activated protein C is not regulated by alpha-2-macroglobulin in plasma. 247 41

Reactions of rabbit alpha-2-macroglobulin with methylamine and trypsin were studied and the results were compared with those obtained for previously described 2-macroglobulins from other species. Rabbit alpha-2-macroglobulin was cleaved by trypsin at a number of sites, whereas the human homologue was split essentially only in the "bait" region into two fragments of similar sizes. Reaction of native or methylamine-treated rabbit alpha-2-macroglobulin with trypsin resulted in a substantial decrease in the intensity of fluorescence induced by binding of 6-(p-toluidino)-2-naphthalenesulfonate or bis(8-anilino-1-naphthalenesulfonate). Under the same conditions, the fluorescence of the human protein increased. The time course of the reaction of rabbit alpha-2-macroglobulin with methylamine was studied by measuring (i) the generation of thiol groups, (ii) the decrease in trypsin-inhibiting activity with remazol brilliant blue hide powder as the substrate, and (iii) the decrease in trypsin-protein amidase activity. The thiol appearance reaction exhibited a multiphasic time course. The initial phase was found to follow second-order kinetics with an apparent rate constant of 1.2 M-1.s-1. Under the same conditions, the human protein showed monophasic kinetics with a rate constant of 12 M-1.s-1. Both the trypsin-inhibiting activity and the trypsin-protein amidase activity concurrently decreased at a slower rate than the thiol appearance. These results indicate that rabbit alpha-2-macroglobulin is more stable to nucleophilic attack by methylamine but less resistant to proteolysis by trypsin than the human homologue, and that the final conformation induced by methylamine differs considerably from that induced by trypsin.
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PMID:Changes in trypsin-binding properties and conformation of rabbit alpha-2-macroglobulin on reaction with methylamine. 247 83

Pancreatic pseudocyst fluid from eight patients was examined biochemically. The fluid was found to be a mixture of plasma proteins and pancreatic juice, possessing a high proteolytic activity against high- as well as low-molecular-weight proteins. The proteolytic activity was found to be trypsin-, kallikrein- and plasmin-like. Gel filtration studies showed proteolytic activity to be present corresponding to alpha-2-macroglobulin-bound proteases and also to free proteases. Quantitative immunochemical levels were about 30-100% of normal plasma levels for alpha-2-macroglobulin, C1 inhibitor, antithrombin III and alpha-2-antiplasmin. However, there was practically no functional inhibitory capacity left in the pseudocyst fluid, except for alpha-1-protease inhibitor, which retained its inhibitory capacity. Neither native kininogen nor complement factor C3 was found: this was probably a result of the proteolytic activity. It is concluded, that a continuing proteolytic activity within the pseudocyst, although decreasing with aging of the cyst, could explain symptoms and complications caused by the pseudocyst.
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PMID:Pancreatic pseudocyst fluid--a mixture of plasma proteins and pancreatic juice possessing a high proteolytic activity. 253 13

Two glycoproteins having trypsin-protein esterase activity were purified to apparent homogeneity from murine plasma. One was alpha-macroglobulin, a homologue of human alpha-2-macroglobulin, while the other, tentatively named murinoglobulin, did not correspond to any of the known plasma protease inhibitors that have been well characterized in men or other mammals. Murinoglobulin contained about 7.6% carbohydrate and was composed of a single-polypeptide chain of Mr = 180,000 as judged by the equilibrium sedimentation analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis under reducing conditions. Murinoglobulin did not cross-react immunologically with mouse alpha-macroglobulin nor with human alpha-2-macroglobulin. Protease-inhibiting properties of murinoglobulin were compared with those of mouse alpha-macroglobulin and human alpha-2-macroglobulin. All the three proteins inhibited trypsin, papain, and thermolysin, although they differed considerably in both the degree of inhibition and the binding stoichiometry of protease-inhibitor complexes. The two macroglobulins inhibited pepsin at pH 5.5, whereas murinoglobulin was inactivated at this pH. Murinoglobulin was more sensitive to methylamine than the two macroglobulins. No protein corresponding to murinoglobulin was detected in human plasma.
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PMID:Murinoglobulin, a novel protease inhibitor from murine plasma. Isolation, characterization, and comparison with murine alpha-macroglobulin and human alpha-2-macroglobulin. 257 55

Thirty-four cases of eosinophilic granulomas, 18 cases of diffuse histiocytosis-X, 2 cases of Letterer-Siwe-like syndrome with immunodeficiency, 4 cases of malignant histiocytosis and virus associated hemophagocytic syndrome were studied. On paraffin section, S100 protein, lysozyme, alpha-1-anti-trypsin, alpha-1-antichymotrypsin, alpha-2-macroglobulin, Transferrin, Ferritin, peanuts agglutinin, Concanavalin-A, and dolichos biflorus associated antigen were stained by the immunoperoxidase method. In a few fresh materials, T-cell subpopulation by use of monoclonal antibodies (OKT-3, 4, 6, and OK-M1) was examined by the immunoperoxidase method. Two types of Langerhans' cells were found, one is positive for Ferritin and alpha-2-macroglobulin in diffuse histiocytosis-X cells, and another is negative for them in both eosinophilic granulomas. Diffuse histiocytosis-X cell resembled the transformed type of Langerhans cell more than eosinophilic granuloma cells in cellular differentiation. It seemed that the term prolangerhans' cell proliferation disorder might be responsible for it.
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PMID:Immunohistochemical and ultrastructural studies on histiocytosis in children. 330 93

Endogenous gram-negative peritonitis leading to septic shock was induced in rats by a defined perforation of the coecum. Cationic trypsin-like immunoreactivity (CTLI) was measured in peritoneal fluid and serum by a radioimmunoassay method. Five, 10, and 15 h after the coecal perforation, CTLI in peritoneal fluid was significantly higher than before the coecal perforation and also higher than in the corresponding control rats. Moreover, CTLI in serum was under the same conditions significantly higher 10 and 15 h after the induction of peritonitis. Gel chromatography of peritoneal fluid and serum during peritonitis showed free CTLI and CTLI bound to both alpha-1-antitrypsin and alpha-2-macroglobulin, whereas only free CTLI could be detected in serum from control rats. These findings were accompanied by local ultrastructural changes in the acinar cells as evaluated by electron microscopy. The pathophysiologic implications of the findings are discussed.
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PMID:Impact of experimental endogenous gram-negative peritonitis on the pancreas of the rat as evaluated by cationic trypsin-like immunoreactivity in peritoneal fluid and serum and by electron microscopy of pancreatic tissue. 358


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