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:3.1.6.4 (chondroitinase)
2,039 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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

1. Proteoglycan aggregates from bovine nasal cartilage were studied by using electron microscopy of proteoglycan/cytochrome c monolayers. 2. The aggregates contained a variably long central filament of hyaluronic acid with an average length of 1037nm. The proteoglycan monomers attached to the hyaluronic acid appeared as side chain filaments varying in length (averaging 249nm). They were distributed along the central filament at an average distance of about 36nm. 3. Chondroitin sulphate side chains were removed from the proteoglycan monomers of the aggregates by partial chondroitinase digestion. The molecules obtained had the same general appearance as intact aggregates. 4. Proteoglycan aggregates were treated with trypsin and the largest fragment, which contains the hyaluronic acid, link protein and hyaluronic acid-binding region, was recovered and studied with electron microscopy. Filaments that lacked the side chain extensions and had the same length as the central filament in the intact aggregate were observed. 5. Hyaluronic acid isolated after papain digestion of cartilage extracts gave filaments with similar length and size distribution as observed for the central filament both in the intact aggregate and in the trypsin digests. 6. Umbilical-cord hyaluronic acid was also studied and gave electron micrographs similar to those described for hyaluronic acid from cartilage. However, the length of the filament was somewhat shorter. 7. The electron micrographs of both intact and selectively degraded proteoglycans corroborate the current model of cartilage proteoglycan structure.
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PMID:Cartilage proteoglycan aggregates. Electron-microscopic studies of native and fragmented molecules. 21 57

Fibronectin is a major surface protein of normal animal cells but is absent from many transformed cells. Addition of fibronectin to transformed cells causes increased cell substrate adhesion and changes in the morphology and cytoskeleton of the cells. We have coupled fibronectin to photoactivable chemical cross-linkers and have added it to cells to identify those molecules to which it binds. In this way, fibronectin can be cross-linked to sulfated proteoglycans at the cell surface. The cross-linking is specific for fibronectin. The fibronectin-proteoglycan complex is sensitive to chondroitinase ABC and AC and to trypsin. Addition of fibronectin also affects binding of hyaluronic acid to the cells. These results suggest that fibronectin interacts with proteoglycans at the cell surface. The existence of such interactions may have implications for the role of fibronectin and proteoglycans in cell adhesion.
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PMID:Cross-linking of fibronectin to sulfated proteoglycans at the cell surface. 22 72

The chondrocyte is a specialized cell that synthesizes proteoglycans of a type found only in cartilage and nucleus pulposus. These proteoglycans are distinct in forming multiple aggregates of unique structure in which hyaluronic acid provides a central chain to which many proteoglycan molecules are bound at one end only. Chondrocytes were isolated from adult cartilage and used in suspension culture to test the effect of compounds in the medium on the synthesis of proteoglycans. Hyaluronic acid alone, among a number of compounds extracted from or analogous to those in cartilage, reduced the incorporation of [35S] sulphate into macromolecular material. Oligosaccharides of hyaluronic acid of the size of decasaccharides and above also had this effect but hyaluronic acid already bound to proteoglycan did not. The proportion of total labelled material associated with the cells increased at the expense of that in the medium. Treatment of the cells with trypsin abolished the effect of hyaluronic acid but treatment with chondroitinase did not. It is suggested that hyaluronic acid interacts with proteoglycans at the cell surface by a specific mechanism similar to that involved in proteoglycan aggregation, as a result of which the secretion and synthesis of proteoglycans is reduced.
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PMID:Influence of the cells on the pericellular environment. The effect of hyaluronic acid on proteoglycan synthesis and secretion by chondrocytes of adult cartilage. 23 23

The distribution of glycosaminoglycans and glycoproteins has been studied in cytoplasmic and particulate fractions of neurons isolated in bulk from rat cerebrum. Lysis of the neurons in 25 mM sodium phosphate buffer at pH 7.5 released 20% of the protein and over 90% of the lactate dehydrogenase in a soluble form. Eighty-two percent of the chondroitin sulfate was also released, together with 55% of the heparan sulfate and 24-25% of the hyaluronic acid and glycoproteins. The chondroitin sulfate remaining in the membranes was completely depolymerized to disaccharides after treatment with chondroitinase ABC, and treatment of the neuronal membranes with 0.1% trypsin removed 55-63% of the chondroitin sulfate and heparan sulfate but only 25% of the sulfated glycoproteins. The results reported here support our previous conclusion that the soluble chondroitin sulfate proteoglycan of brain is largely a cytoplasmic constitutent of neurons (and astrocytes) and is not primarily present in nervous tissue as an extracellular ground substance.
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PMID:Presence of chondroitin sulfate in the neuronal cytoplasm. 28 11

Proteoglycans isolated under associative conditions in the presence of protease inhibitors from human nucleus pulposus contained 17% aggregate and 83% non-aggregating monomer (Kav = 0.5 on Sepharose CL-2B). Isolated aggregate after reduction and alkylation was resolved into two components (Kav = 0.15 and 0.43) on Sepharose CL-2B. Labeled proteoglycans isolated from parallel samples pulsed with [35S]sulfate and chased for up to 18 h were present largely as aggregated material (up to 78%). Reduction and alkylation of the labeled samples gave a labeled proteoglycan monomer with Kav = 0.15. Both the labeled and unlabeled chondroitin sulfate chains had the same distribution on Sepharose CL-6B and equivalent molecular weights (Mr = 2.0 x 10(3)). After chondroitinase ABC digestion, the unlabeled keratan sulfate-protein core was polydisperse with a Kav = 0.38 on Sepharose CL-4B while the labeled keratan sulfate-protein core had a Kav = 0.05. This indicates that the newly synthesized proteoglycan had a large core protein and suggests that the proteoglycans present in nucleus pulposus are originally synthesized as large molecular weight, aggregating proteoglycans.
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PMID:Aggregated proteoglycan synthesis in organ cultures of human nucleus pulposus. 50 May 96

It was shown that a proteoglycan is synthesised by embryos of a Japanese sea urchin, Hemicentrotus pulcherrimus. This proteoglycan appears as a single peak on sucrose density gradient ultracentrifugation throughout the development. About half of the mucopolysaccharide moiety in this proteoglycan was found to be dermatan sulphate and the rest to be chondroitinase-resistant mucopolysaccharides. Evidence is presented to show that both types of mucopolysaccharide do not exist in a free form but reside as an integral part of the proteoglycan. The linkage between mucopolysaccharide and protein moieties of the proteoglycan appeared not to be an O-glycosidic bond, which is common among other proteoglycans such as proteochondroitin sulphate and proteodermatan sulphate.
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PMID:Appearance of a proteoglycan in developing sea urchin embryos. 66 28

Monomer proteoglycan was isolated from porcine ovarian follicular fluid by isopycnic CsCl centrifugation in the presence of 4 M guanidine HCl and protease inhibitors. The elution profile of the D1 preparation on Sepharose 2B was similar to that of monomer proteoglycan from bovine nasal cartilage, indicating a similar molecular size. Follicular fluid proteoglycans consist of about 20% protein, 50% dermatan sulfate, and 20% oligosaccharides rich in sialic acid, galactose, mannose, glucosamine, and galactosamine. The amino acid composition of this proteoglycan is significantly different from that of cartilage proteoglycans, with a higher proportion of aspartic acid, threonine, and lysine, and lower amounts of proline and glycine. Alkali-released dermatan sulfate chains are larger on Sepharose 6B (average Mr = 56,000) than chondroitin sulfate chains from cartilage proteoglycans (average Mr = 25,000), and iduronic acid accounts for 9% of total hexuronic acid. Disaccharide units released by chondroitinase ABC consists of 67% 4-sulfated, 22% 6-sulfated, 5% non-sulfated, and 5% disulfated disaccharides. After treatment with 0.05 M NaOH, 1 M NaBH4 at 45 degrees C for 24 h, two major sialic acid-containing oligosaccharides were observed on Sephadex G-25, corresponding to penta- and hexasaccharides. The pentasaccharide contained sialic acid, galactose, glucosamine, and galactosamine in the proportions 1:2:1:1. The galactosamine is O-glycosidically linked to the protein core. This oligosaccharide accounts for approximately 77% of all the sialic acid in the follicular fluid proteoglycans. The hexasaccharide fraction contained sialic acid, galactose, mannose, and glucosamine in the proportions 1:2:1:2. It also contained a small amount of fucose and galactosamine. The linkage of these oligosaccharides to the protein core remains to be determined. The follicular fluid proteoglycans, unlike those from cartilage, do not interact with hyaluronic acid. Digestion with trypsin, chymotrypsin, or plasmin released dermatan sulfate-peptides nearly as small as those released by papain or alkali; in contrast, cartilage proteoglycans were resistant to plasmin and released peptides containing an average of more than four chondroitin sulfate chains after trypsin or chymotrypsin digestion.
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PMID:Isolation and characterization of proteoglycans from porcine ovarian follicular fluid. 76

In vitro, cartilage proteoglycans (PGs) are effective inhibitors of hydroxyapatite formation and growth. Their inhibitory ability decreases with decreasing PG size and charge density. It has been suggested that the enzyme-mediated alteration in the size and conformation of PGs in the growth plate may similarly facilitate the calcification process. In this study, a gelatin gel system was used to monitor hydroxyapatite formation and growth in the presence of proteoglycan aggregates, before and after enzyme treatment. To reproduce the physeal degradation cascade, an enzyme preparation was used that contained all of the growth plate enzymes. At a concentration of 500 micrograms/ml, the untreated proteoglycan aggregates reduced the amount of mineral formed by 30%. When the aggregates were treated with the heat-inactivated enzyme, the same extent of inhibition was found. In contrast, treating the aggregates with the crude growth plate enzyme preparation removed all the inhibitory ability, such that 500 micrograms/ml of proteoglycan preparation yielded 10% more mineral than the controls. Treatment of the aggregates with chondroitinase ABC and trypsin, similarly removed all the inhibitory ability. These data, suggest that enzymatic degradation of proteoglycans may contribute to the regulation of growth plate calcification.
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PMID:Treatment of proteoglycan aggregates with physeal enzymes reduces their ability to inhibit hydroxyapatite proliferation in a gelatin gel. 131 95

Cultured chick embryo skin fibroblasts release a major component with a native molecular mass of about 1 MDa, which resolves into three polypeptide bands of about 300, 350 and 600 kDa upon reduction. We report here the purification of this oligomeric protein and show, by means of polyclonal and monoclonal antibodies, that its three polypeptide constituents are closely related. The 600-kDa polypeptide is likely to be a dimer of two smaller subunits which are cross-linked by non-reducible bonds. By electron microscopy, isolated oligomeric molecules exhibit a novel cruciform structure with a large central globular domain. One arm has the shape of a thin rod about 70 nm in length. The three other arms are thicker, longer (90 nm) and flexible, and carry a prominent double globule at their distal ends. Collagenase treatment of the oligomeric fibroblast protein yields two resistant fragments of about 270 kDa and 320 kDa. The intact 350-kDa and 600-kDa (but not the 300-kDa) polypeptides are chondroitinase sensitive and labeled by metabolic incorporation of [35S]sulfate; collagenase treatment does not remove any [35S] sulfate. Hence, the intact fibroblast protein has glycosaminoglycan chains attached to its non-collagenous domain. Three amino acid sequences obtained from chymotryptic fragments of the fibroblast protein correspond to sequences predicted for chick type-XII collagen from its full-length cDNA [Yamagata, M., Yamada, K. M., Yamada, S. S., Shinomura, T., Tanaka, H., Nishida, Y., Obara, M. & Kimata, K. (1991) J. Cell Biol. 115, 209-221]. However, the novel fibroblast protein described here differs significantly from previously isolated forms of type-XII collagen: its subunits are larger by one third, and it is a proteoglycan.
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PMID:A major oligomeric fibroblast proteoglycan identified as a novel large form of type-XII collagen. 132 60


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