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:3.2.1.36 (hyaluronidase)
4,606 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Polyacrylamide beads containing entrapped 35S-labelled proteoglycan molecules have been prepared. 2. The measurement of release of radioactivity provides an extremely sensitive assay for proteoglycan-degrading enzymes, including proteinases and hyaluronidase. 3. The amount of label released is a logarithmic function of enzyme concentration or time of incubation. Experiments were made in an attempt to explain this. 4. Assays were made by the new method at several pH values, and with the inclusion of inhibitors to identify the proteoglycan-degrading enzymes of rabbit ear cartilage. 5. A previously undescribed proteinase active against proteoglycan at pH4.5 but unaffected by pepstatin, was discovered. The enzyme was named cathepsin F, and was partially purified and characterized; it was detected in human articular cartilage.
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PMID:Proteoglycan-degrading enzymes. A radiochemical assay method and the detection of a new enzyme cathepsin F. 2 63

Studies were undertaken to define more fully the antigenic properties of human articular cartilage proteoglycans, in anticipation of its potential contribution to alterations arising in diseased states and following cartilage transplantation. Proteoglycans, extracted from normal, adult articular cartilage by dissociative measures, were subjected to purification by cesium density gradient ultracentrifugation, under conditions facilitating both molecular aggregation and dissociation. A polydisperse population of reactive determinants was observed in immunodiffusion and hemagglutination inhibition systems, employing proteoglycan specific antisera on gradient fractions. Highly aggregated proteoglycan species appeared to contain potentially masked antigenic determinants, which were revealed after guanidine dissociation but not hyaluronidase digestion. Polyacrilamide disc gel electrophoresis in sodium dodecyl sulfate, in conjunction with disc elution experiments, confirmed proteoglycan antigenic polydispersity.
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PMID:Polydispersity of human articular cartilage proteoglycan antigens. 6 14

The proteoglycans of cartilage are complex molecules in which chondroitin sulphate and keratan sulphate chains are covalently linked to a protein core, forming a polydisperse population of proteoglycan monomers. By interaction with hyaluronic acid and link proteins, the monomers form large macromolecular complexes. In vivo the proteoglycans mainly occur in such aggregates. In the electron microsope, the cartilaginous matrix can be seen to be made up of thin collagen fibrils and polygonal granules about 10-50 nm in diameter Addition of the polyvalent cationic dye Ruthenium Red to glutaraldehyde and osmium tetroxide fixatives yields a dense selective staining of the matrix granules. Following a short digestion of cartilage slices with either of the chondroitin sulphate-degrading enzymes hyaluronidase and chondroitinase or with the proteolytic enzyme papain, the matrix granules were few in number or completely absent and the proteoglycan content, measured as hexosamine, decreased by up to 90%. Similarly, extraction of the cartilage with 4 M guanidine-HCl removed all matrix granules and most of the proteoglycans. From these findings, it can be concluded that the matrix granules represent proteoglycans, most probably in aggregate form, and that Ruthenium Red staining may be used to study the distribution of these macromolecules in thin sections. As a complement to chemical studies on proteoglycan structure, it is also possible to observe and measure individual molecules in the electron microscope after spreading them into a monomolecular layer with cytochrome c. This technique has been applied in investigations on proteoglycans isolated from bovine nasal cartilage and other hyaline cartilages. The molecules in the monomer fractions appeared as an extended central core filament to which about 25--30 side-chain filaments were attached at various intervals. The core filament, averaging about 300 nm in length, was interpreted as representing the polysaccharide binding part of the protein core and the side-chain filaments, averaging about 45 nm in length, as representing the clusters of chondroitin sulphate chains. Statistical treatment of the collected data indicated that no distinct subpopulations existed within the monomer fractions. The electron microscopic results correlated well with chemical data for the corresponding fractions and together with recent observations on various aggregate fractions strongly support present concepts of proteoglycan structure.
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PMID:Electron microscopy of cartilage proteoglycans. 6 24

Proteoglycans extracted with 4M-guanidinium chloride from pig laryngeal cartilage and bovine nasal septum were purified by density-gradient centrifugation in CsCl under 'associative' followed by 'dissociative' conditions [Hascall & Sajdera (1969) J. Biol. Chem. 244, 2384-2396]. Proteoglycans were then digested exhaustively with testicular hyaluronidase, which removed about 80% of the chondroitin sulphate. The hyaluronidase was purified until no proteolytic activity was detectable under the conditions used for digestion. The resulting 'core' proteins of both species were fractionated by a sequence of gel-chromatographic procedures which gave four major fractions of decreasing hydrodynamic size. Those that on electrophoresis penetrated 5.6% (w/v) polyacrylamide gels migrated as discrete bands whose mobility increased with decreasing hydrodynamic size. The unfractionated 'core' proteins had the same N-terminal amino acids as the intact proteoglycan, suggesting that no peptide bonds had been cleaved during hyaluronidase digestion. Alanine predominated as the N-terminal residue in all the fractions of both species. Fractions were analysed for amino acid, amino sugar, uronic acid and neutral sugar compositions. In pig 'core' proteins, the glutamic acid content increased significantly with hydrodynamic size, but in bovine 'core' proteins this trend was less marked. Significant differences in amino acid composition between fractions suggested that in each species there was more than one variety of proteoglycan. The molar proportions of xylose to serine destroyed on alkaline beta-elimination were equivalent in most fractions, indicating that the serine residues destroyed were attached to the terminal xylose of chondroitin sulphate chains. The ratio of serine residues to threonine residues destroyed on beta-elimination, was similar in all fractions of both species. Since the fractions of smallest hydrodynamic size contained less keratan sulphate than those of larger size, it implies that in the former the keratan sulphate chains were shorter than in the latter.
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PMID:The nature of the protein moieties of cartilage proteoglycans of pig and ox. 12 55

Extracts of human peripheral blood polymorphonuclear leukocyte granules, and two purified proteases derived from such extracts, an elastase and a chymotrypsin-like enzyme, degrade isolated bovine nasal cartilage proteoglycan at neutral pH. Viscosity studies indicate that the leukocyte granule extracts lack hyaluronidase activity and that their degradative effect on proteoglycan at physiological pH is due entirely to proteolytic action. Sepharose 4B gel chromatography and SDS-polyacrylamide gel electrophoresis of proteoglycan fractions treated with leukocyte granule enzymes at pH 7.0 indicate that they degrade one of the proteoglycan link proteins, release a fragment from the hyaluronic acid-binding portion of the proteoglycan subunit core protein, and break down the remainder of the proteoglycan subunit molecule into peptide fragments with varying numbers of chondroitin sulfate chains. Immunodiffusion studies indicate that the antigenic determinants of the proteoglycan subunit core protein and the link proteins survive treatment with granule proteases. Similar degradation of human articular cartilage proteoglycan by granule neutral proteases can be presumed to occur, in view of the similarity of structure of human articular and bovine nasal cartilage proteoglycans. The release of granule enzymes in the course of neutrophil-mediated inflammation can thus result in the degradation of cartilage matrix proteoglycan, leading to cartilage destruction and joint injury.
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PMID:Degradation of cartilage proteoglycan by human leukocyte granule neutral proteases--a model of joint injury. II. Degradation of isolated bovine nasal cartilage proteoglycan. 12 83

Rat liver cells grown in primary cultures in the presence of [(35)S]sulphate synthesize a labelled heparan sulphate-like glycosaminoglycan. The characterization of the polysaccharide as heparan sulphate is based on its resistance to digestion with chondroitinase ABC or hyaluronidase and its susceptibility to HNO(2) treatment. The sulphate groups (including sulphamino and ester sulphate groups) are distributed along the polymer in the characteristic block fashion. In (3)H-labelled heparan sulphate, isolated after incubation of the cells with [(3)H]galactose, 40% of the radioactive uronic acid units are l-iduronic acid, the remainder being d-glucuronic acid. The location of heparan sulphate at the rat liver cell surface is demonstrated; part of the labelled polysaccharide can be removed from the cells by mild treatment with trypsin or heparitinase. Further, a purified plasma-membrane fraction isolated from rats previously injected with [(35)S]sulphate contains radioactively labelled heparan sulphate. A proteoglycan macromolecule composed of heparan sulphate chains attached to a protein core can be solubilized from the membrane fraction by extraction with 6m-guanidinium chloride. The proteoglycan structure is degraded by treatment with papain, Pronase or alkali. The production of heparan [(35)S]sulphate by rat liver cells incubated in the presence of [(35)S]sulphate was followed. Initially the amount of labelled polysaccharide increased with increasing incubation time. However, after 10h of incubation a steady state was reached where biosynthetic and degradative processes were in balance.
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PMID:Structure and metabolism of rat liver heparan sulphate. 14 28

Proteoglycans were extracted from bovine articular cartilage with guanidine-HCl and fractionated in cesium chloride density gradients by equilibrium ultracentrifugation. The acidic glycosaminoglycan (AGAG) components were then determined enzymatically with chondroitinase-ABC and streptomyces hyaluronidase. Under associative and dissociative conditions, the distribution of the AGAG components was as follows: the ratio of 4-sulfated disaccharide units to total AGAG increased with decreasing density gradients whereas that of 6-sulfated disaccharide units to total AGAG increased with increasing density gradients. The ratio of disulfated disaccharide units to total AGAG increased somewhat with decreasing density gradients whereas that of non-sulfated disaccharide units tended to decrease. Although the cartilage proteoglycan macromolecules were heterogeneous, a certain regularity was observed with respect to the distribution of sulfate and the degree of sulfation in the chondroitin sulfate chains of the proteoglycans.
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PMID:Constitutional heterogeneity of the glycosaminoglycans in articular cartilage proteoglycans. 14 4

The basic subunit of cartilage proteoglycan consists of multiple glycosaminoglycan chains covalently attached to a core protein. It is unclear as to whether there is a single core protein or multiple different core proteins, since previous studies using either chondroitinase or testicular hyaluronidase to enzymatically remove chondroitin sulfate side chains from the proteoglycan subunit have yielded conflicting results. In the present study, a chondroitinase-produced core protein preparation isolated as a single peak on Sepharose gel chromatography was found to contain at least two immunologically distinct components. Hyaluronidase-produced core protein from the same proteoglycan subunit fraction was found to contain multiple components nearly all of which were smaller than the components in the chondroitinase digest. A possible explanation of these findings is that they resulted from proteolytic degradation of the core protein in the course of the enzymatic removal of its chondroitin sulfate. The presence of small amounts of protease contaminants in several commercial chondroitinase and hyaluronidase preparations was detected by an extremely sensitive radioassay. Until proteases can be rigorously excluded from enzyme preparations used to degrade the proteoglycan subunit, it will not be possible to determine whether it consists of a single or several different core proteins.
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PMID:A comparison of bovine nasal cartilage proteoglycan core protein produced by chondroitinase and hyaluronidase: the possible role of protease contaminants. 14 80

From studies of isolated cartilage proteoglycans in solution it has been inferred that they occur in the tissue as aggregates of high molecular weight which consist of proteoglycan monomers, hyaluronic acid and specific link proteins. The present investigation provides direct evidence for the existence of hyaluronic acid-containing aggregates in vivo, as indicated by the following observations: Treatment of sections of coastal cartilage from newborn rabbits with Streptomyces hyaluronidase led to complete disappearance of the electron dense granules, which have been previously identified as chondroitin sulfate proteglycans, from the extracellular matrix. Similar results were obtained on digestion with leech hyaluronidase which, like the Streptomyces enzyme, specifically degrades hyaluronic acid. Proteoglycan aggregation occurs not only in the extracellular compartment but intracellularly as well, since a portion of the hyaluronidase-senstive, electron dense proteoglycan granules are found in intracellular vesicles. It is concluded that the ability of proteoglycan monomers to form aggregates is a true reflection of the in vivo organization of these molecules and that aggregate formation is an important factor in the maintenance of the normal physiological function of cartilage tissue.
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PMID:Role of hyaluronic acid in the in vivo aggregation of cartilage proteoglycans. 15 Sep 64

Commercial testicular hyaluronidase preparations are contaminated by a small amount of protease activity which is partially inhibited by serine-protease inhibitors or pepstatin. These protease inhibitors can be shown by Sepharose gel column chromatography to abolish or reduce hyaluronidase-induced degradation of bovine nasal cartilage proteoglycan subunit without affecting the ability of the enzyme to degrade chondroitin sulfate. In addition, immunodiffusion studies indicate that pretreatment of hyaluronidase with these protease inhibitors reduces or abolishes the ability of the enzyme to produce a second "link-related" immunoprecipitin line upon digestion of link protein-containing proteoglycan fractions. Thus, the enhancement of immune reactivity and the unmasking of an additional antigen noted after digestion of cartilage proteoglycan with testicular hyaluronidase are most likely due to the exposure of additional antigenic sites or the the release of more highly immunoreactive fragments by the contaminant proteases rather than to the action of hyaluronidase itself.
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PMID:The effect of contaminant proteases in testicular hyaluronidase preparations on the immunological properties of bovine nasal cartilage proteoglycan. 15 47


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