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)

Botulinum A neurotoxin (BoNtx) produced a partial inhibition of carbachol induced 3H-noradrenaline (3H-NA) release from bovine adrenal chromaffin cells in monolayer culture. Each of the polysialogangliosides GD1a, GT1b and GD1b enhanced the block of exocytosis when they were applied prior to the toxin exposure. The monosialoganglioside GM1 was not effective. Chromaffin cells treated with neuraminidase lost their sensitivity to BoNtx. Application of gangliosides to these cells, however, restored their susceptibility to the toxin. Treatment of the cells with trypsin did not affect the BoNtx-blockade of 3H-NA-release. The potency of botulinum A toxin was increased in a solution of low ionic strength in which sodium chloride was replaced by sucrose. In agreement with the potency of botulinum A neurotoxin in blocking exocytosis under the various conditions, binding of 125I-botulinum A neurotoxin to chromaffin cells was enhanced in low ionic strength solution and by pretreatment of the cells with gangliosides. The binding was decreased by digestion of gangliosides with neuraminidase. It is concluded that botulinum A neurotoxin binds exclusively to polysialogangliosides which subsequently serve as carriers for the toxin. The low ionic strength may increase some physico-chemical interaction of the toxin with the polysialogangliosides.
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PMID:The translocation of botulinum A neurotoxin by chromaffin cells is promoted in low ionic strength solution and is insensitive to trypsin. 204 36

Human mast cell tryptase was purified from lung tissue by high salt extraction, ammonium sulphate precipitation, octyl Sepharose and heparin-agarose chromatography. The tryptase isolated was a tetramer with a molecular weight of 132 kD on gel filtration, and on SDS-polyacrylamide gel electrophoresis was reduced to a single diffuse band with a mean molecular weight of 32.5 kD. Purified tryptase catalysed the cleavage of the tryptic substrates tosyl L-arginine methyl ester and benzoyl DL-arginine p-nitroanilide; enzymatic activity was enhanced in the presence of heparin but markedly decreased in the presence of 2 M sodium chloride. Rabbit antisera and three new monoclonal antibodies (AA1, AA3 and AA5) were produced which were specific for tryptase in indirect ELISAs, immunoenzymatic overlay in crossed immunoelectrophoresis and by Western blotting. Additive and competitive ELISA experiments suggested that the three monoclonal antibodies all recognized epitopes within a single highly immunogenic area of the tryptase molecule, and enzyme assays indicated that this site was distant from the active site. Binding of monoclonal antibodies to tryptase was not affected by the presence of heparin, or by periodate treatment of the antigen suggesting that carbohydrate epitopes were not recognized. Western blotting indicated that some heterogeneity in molecular weight for monomeric tryptase was not reflected in antigenic differences. An immunofluorescence procedure with cytocentrifuge preparations of enzymatically dispersed lung, colon and skin revealed highly specific localization of tryptase to the granules of all mast cells, but there was no binding to other cells in these preparations, to cultured keratinocytes, to basophils or to any other blood leucocyte.
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PMID:Production and characterization of monoclonal antibodies specific for human mast cell tryptase. 225 91

A monoclonal antibody, FAC2, was isolated by immunization of mice with a Photosystem II core preparation followed by splenic fusion and standard monoclonal antibody screening and production techniques. This antibody recognizes the 49-kDa polypeptide of Photosystem II which is the apoprotein of CPal. The antigenic determinant recognized by this antibody lies on a cyanogen bromide fragment which appears as a doublet with an apparent molecular mass of 14.5 kDa. FAC2 was used to follow the effects of trypsin on the 49-kDa polypeptide in a membrane environment. Our results indicate that the extrinsic polypeptides of Photosystem II which are known to be involved in oxygen evolution protect the 49-kDa polypeptide from tryptic attack. Additionally, Photosystem II membranes which are treated with alkaline Tris exhibit a large increase in the ability to bind FAC2. This increase is not observed with membranes treated with calcium chloride or sodium chloride. These results indicate that the 49-kDa polypeptide may be at least structurally associated with the component(s) responsible for oxygen evolution.
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PMID:Use of a monoclonal antibody in structural investigations of the 49-kDa polypeptide of photosystem II. 244 Mar 84

The murine nerve growth factor, when injected i.v. or, combined in vitro with plasma, was found largely associated with the mouse alpha-macroglobulin (a homologue of human alpha 2-macroglobulin). The nerve growth factor-alpha-macroglobulin complex produced is sufficiently stable to resist separation by gel filtration in 1.0 M sodium chloride, polyacrylamide gel electrophoresis, and immunoprecipitation by antibodies against alpha-macroglobulin. As determined by equilibrium binding studies and computer generated Scatchard analysis, alpha-macroglobulin apparently possesses two types of binding sites with the apparent dissociation constants of 1.2 x 10(-6) and 2.9 x 10(-9) M, respectively, saturable by 3.7 and 0.03 moles of nerve growth factor. Hence, about one mole of nerve growth factor is bound to each of the four subunits of alpha-macroglobulin. Nerve growth factor can be readily dissociated from alpha-macroglobulin in sodium dodecyl sulfate gel electrophoresis in the absence of a reductant. Procedures that affect the proteinase-binding or methylamine- activities of alpha-macroglobulin do not affect the binding of nerve growth factor, and the binding is unaffected by the presence of zinc ions or EDTA. Hence, nerve growth factor is noncovalently associated with alpha-macroglobulin at a site separate from that of the proteinase-, methylamine-, and zinc-binding sites of alpha-macroglobulin. Mouse alpha-macroglobulin can protect the nerve growth factor from inactivation by trypsin. Even in the presence of trypsin, alpha-macroglobulin-nerve growth factor complexes still can stimulate the neurite outgrowth by dorsal root ganglia of 9-day-old chicken embryos. Since alpha-macroglobulin can specifically and noncovalently carry nerve growth factor, one important role of this alpha-macroglobulin in the circulation and extracellular spaces may be to protect the nerve growth factor from proteinase inactivation.
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PMID:Interaction of nerve growth factor with murine alpha-macroglobulin. 246 89

Octadecyl-bonded silica, commonly used for reverse-phase high-pressure liquid chromatography, was modified using surfactants bearing ionizable groups and the modified packing used in ion-exchange chromatography of proteins. The surfactants 2-(n-hexadecylheptaethoxy)acetic acid, 1-(n-hexadecyloctaethoxy)ethylene-diamine, and N-(n-hexadecyloctaethoxy)pyridinium were adsorbed onto test columns packed with octadecyl-bonded silica particles. The proteins lysozyme, bovine serum albumin, trypsin, horse serum cholinesterase, and bovine liver carboxylesterase were used to study the ion-exchange characteristics of the modified packings. The retention order of the proteins on the surfactant-modified stationary phases were as predicted by the isoelectric point of each protein. In addition, the interaction of enzymes with the packings did not result in significant loss of enzymatic activity. Surfactant removal was possible with the use of organic solvents and this allowed the octadecyl-bonded surface to be used again in the reverse-phase mode. During the course of the experiments, no degradation in the packing's performance was observed due to loss of adsorbed surfactant, even after over 85,000 column volumes of sodium chloride and Tris-HCl buffers were circulated through the column.
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PMID:Reversible conversion of octadecyl-bonded silica to ion-exchange surfaces for protein separations. 254 Jun 75

Corncobs, which are distinct morphological units formed by the ordered coaggregation of a filamentous microorganism and streptococci, can be made in vitro by using oral strains of Fusobacterium nucleatum and Streptococcus sanguis. Previous studies have shown that strains of F. nucleatum contain one of at least two different types of corncob receptor. The objective of this study was to isolate the receptor from F. nucleatum ATCC 10953 as the first step in the elucidation of the molecular basis of corncob formation. The cell envelope fraction from this bacterium was treated with trypsin, delipidated with chloroform-methanol, and subjected to ion-exchange chromatography. A single polypeptide (apparent Mr, 39,500), which was eluted from the column with 0.5 M sodium chloride and extracted with dodecyltrimethylammonium bromide to remove contaminating lipopolysaccharide, inhibited corncob formation between strain ATCC 10953 and S. sanguis CC5A. Similarly derived cell fractions from either F. nucleatum FDC 364 or Fusobacterium necrophorum failed to effect coaggregation in the inhibition assay. Amino acid analysis of the polypeptide showed a moderately hydrophobic character (polarity index, 41) and 11% basic residues. Antiserum made against the purified polypeptide agglutinated F. nucleatum ATCC 10953, neutralized the ability of this bacterium to form corncobs, and agglutinated whole cells of S. sanguis CC5A that were precoated with the receptor polypeptide. The identification and isolation of this receptor should greatly enhance our ability to define some of the complex intergeneric coaggregation mechanisms that are thought to occur in the human oral cavity.
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PMID:Isolation of a corncob (coaggregation) receptor polypeptide from Fusobacterium nucleatum. 291 93

In chickens inoculated into the heart with a sodium chloride extract of Escherichia coli strain (serotype O2) isolated from a chicken with colibacillosis, characteristic hemorrhages into the anterior chamber of the eyes (hyphema) were found. Significant lesions were limited to the eyes. Cyclophosphamide-treated chickens were more sensitive to the extract than untreated chickens and hyphema was usually seen in association with hemorrhages of the iris. These activities were not reduced by heating the extract at 60 degrees C for one hour or by trypsin digestion. Chickens inoculated into the heart with commercial lipopolysaccharides of E. coli (serotypes O111:B4 and O55:B5) and Salmonella typhimurium showed similar lesions in the eyes as the chickens inoculated with the sodium chloride extract. These findings suggest that the endotoxin may induce hyphema in chickens.
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PMID:The effect of sodium chloride extract and commercial lipopolysaccharides of Escherichia coli and Salmonella typhimurium on chickens. 328 82

Trypsin and extracellular proteinases produced by Bacillus sp. were purified by column chromatography on coffee grain particles. The ballast proteins were eluted with water, while the adsorbed proteinases were eluted with 1 M sodium chloride solution. The capacity is approximately 2 mg of trypsin per ml of the packed sorbent.
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PMID:Purification of trypsin and bacterial proteinases by column chromatography on coffee grain particles. 330 67

The combined effect of water activity (aw) and pH on growth and toxin production by Clostridium botulinum type G strain 89 was investigated. The minimum aw at which growth and toxin formation occurred was 0.965, for media in which the pH was adjusted with either sodium chloride or sucrose. The minimum pH (at the optimum aw) for growth and toxin production of C. botulinum type G was found to be 5.6. Optimum conditions for toxin activation were a trypsin concentration of 0.1%, a pH of the medium of 6.5, and an incubation for 45 min at 37 degrees C. These data did not show evidence of heat-labile spores, since a heat shock of 75 degrees C for 10 min did not significantly decrease the spore count of strain 89G in media at pH 7.0 or 5.6. It was frequently observed that cells grown at reduced aw or pH experienced severe morphological changes.
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PMID:Effect of water activity and pH on growth and toxin production by Clostridium botulinum type G. 351 31

The binding ability of Cl. botulinum neurotoxin to synaptosomes upon treatment with various enzymes (neuraminidase, trypsin, and beta-bungarotoxin containing phospholipase A2 activity) was studied. When synaptosomes were treated with neuraminidase, their ability to bind toxin decreased; trypsin and beta-bungarotoxin had slightly week or no effect. The decrease in toxin-binding ability of synaptosomes was paralleled by a release of sialic acid from the synaptosomes by the neuraminidase treatment. The toxin-binding ability of synaptosomes treated with neuraminidase was lower than untreated ones at a high concentration of sodium chloride. The binding of the toxin to synaptosomes occurred at least at the two types of structural sites, one site which contained sialic acid, and other site which was sensitive to high ionic strength. It may be possible that another binding state except these is present at the synapse.
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PMID:Binding ability of Clostridium botulinum neurotoxin to the synaptosome upon treatment of various kinds of the enzymes. 356 63


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