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 acidic glycosaminoglycans (AGAG) in normal human kidneys were fractionated on Dowex 1-X2 columns and analysed by electrophoretic separation in three buffers on cellulose acetate membranes and gel filtration on Sephadex G-100 columns, before and after digestion with chondroitinases and streptomyces hyaluronidase. Thin-layer chromatography was also performed to separate glucosamine from galactosamine moieties. Enzymatic digestion combined with electrophoretic characterization indicated that heparan sulfates exist as the main AGAG which accounted for two-fifths of the total AGAG. Hyaluronic acid and dermatan sulfates accounted for one-fourth and one-sixth of the total kidney AGAG, respectively. Chondroitin sulfate isomers (4-sulfate and 6-sulfate) consisted of the residual one-sixth of the total AGAG. An oversulfated chondroitin sulfate was detected in a small amount by demonstration of the unsaturated disulfated disaccharide after digestion with chondroitinase-ABC but not with chondroitinase-AC.
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PMID:Acidic glycosaminoglycans in human kidney tissue. 12 23

1. Human embryonic lung and skin fibroblasts were allowed to incorporate 32SO42- or 35SO42- and D-[1-3H]glucosamine. After removal of the medium the monolayer was subjected to sequential extractions by using EDTA, brief trypsin digestion, extraction with dithiothreitol ofllowed by freeze--thawing and extraction with trichloroacetic acid. The heparan sulphate and galactosaminoglycan contents of the various extracts were estimated after deaminative cleavage of the former component. Heparan sulphate was the major component of the trypsin digest, whereas galactosaminoglycans were the dominant component of other fractions. 2. Galactosaminoglycans of the various fractions were subjected to chemical (periodate oxidation/alkaline elimination) and enzymic (chondroitinase-AC and -ABC, as well as testicular hyaluronidase) degradations. Galactosaminoglycans from the insoluble cell fraction and the dithiothreitol extract contained larger amounts of L-iduronic acid than did those of other fractions. 3. Pulse-chase experiments were performed with and without replating of the cells at the start of the chase period. Radioactive glycans were isolated from the various extracts during the chase period. The half-lives of glycans of the insoluble cell fraction and the dithioreitol extract were shorter (5--8h) than were those of the trypsin digest and the EDTA extract (22h and 11h respectively). After replating of the cells in chase medium, radioactive cell-associated glycans were secreted from the cells and could be recovered in the trypsin digest, the EDTA extract and the medium. Furthermore, 35S/3H ratios of glycans from all these fractions decreased during the chase period. The following conclusions were reached. The insoluble cell fraction contains the synthesis pool and some structural material, whereas the soluble cell fraction is the storage and degradation pool. The dithiothreitol extract appears to contain the immediate precursors of secreted material. The trypsin-released glycans comprise structural components as well as material destined for pinocytosis or secretion into the medium. The EDTA extract is considered to consist of glycans en route to the medium. 4. The two presumptive precursor pools were preferentially depleted of L-iduronic acid-rich galactosaminoglycans during the chase. Glycans recovered from the trypsin digest, the EDTA extract and the medium during the chase contained larger amounts of periodate-resistant uronic acid residues (D-glucuronic acid and/or L-iduronic acid O-sulphate) than did their precursors. It is proposed that polymer-level modifications of secreted glycans are partly responsible for the results.
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PMID:Structure and metabolism of sulphated glycosaminoglycans in cultures of human fibroblasts. Structural characteristics of co-polymeric galactosaminoglycans in sequential extracts of fibroblasts during pulse-chase experiments. 22 Sep 58

Transfer of radioactive materials to fixed cells from an overlying layer of living cells has been examined to determine whether fixed cells can act as acceptors of glycosyltransferases of living cells. After the incubation of living cells were removed by EDTA treatment, and the radioactivity associated with the fixed cells was determined. Lipids, proteins and carbohydrates were found to be transfered from the living cells to the fixed cells. The amount of radioactivity transferred to the fixed cells was dependent on the number of both fixed and living cells and increased with the time of incubation. When fixed cells were treated with chloroform-methanol before the addition of living cells, the transfer of both lipids and proteins to the fixed cells decreased drastically, but only a slight decrease incarbohydrate transfer was observed. Most of the radioactive materials transferred from living cells labeled with glucosamine or fucose to chloroform-methanol-treated fixed cells were solubilized by trypsin but not by the detergents tested. Approximately 55% of the materials transferred from the cells labeled with glucosamine could be solubilized by hyaluronidase and chondroitinase, and the rest was solubilized by neuraminidase and a glycosidase mixture. The treatment of chloroform-methanol-extracted fixed cells with trypsin caused a significant decrease in the transfer from cells labeled with glucosamine. When nucleotide sugars were used as the radioactive precursor, no significant amount of radioactivity was transferred to the fixed cells.
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PMID:Cellular interaction between fixed and living cells. Transfer of radioactive materials from living cells to fixed cells. 37 19

The amount of glycosaminoglycan (GAG) in dry costal cartilage tissue of rats decreased with aging, while the GAG content in mg DNA (unit cartilage cell) remained the same with aging. These results can be explained by the finding that the total number of cartilage cells decreased with aging. Electrophoretic analysis showed that chondroitin 4-sulfate was the major GAG in rat costal cartilage of various ages. Rat costal cartilage of different ages was incubated with radioactive precursors, and newly synthesized GAG was prepared and the radioactivity analyzed to determine the biosynthetic activity. As to changes in the radioactivity uptake with aging per mg dry cartilage tissue, aging influenced [35S]sulfate incorporation into GAG more significantly than [3H]glucosamine incorporation into GAG. There was a significant decrease in the specific radioactivity of [35S]sulfate per mg DNA (unit cartilage cell), whereas the specific radioactivity of [3H]glucosamine per mg DNA did not change significantly with aging. Both the total sulfotransferase activity and the specific activity per mg DNA decreased significantly with aging. Analysis of disaccharide units formed after chondroitinase ABC digestion of labeled GAG isolated from young and old cartilage showed that the percentage of incorporation of [3H]glucosamine into deltaDi-OS increased significantly with aging. These results suggested that the appearance of nonsulfated positions in the structure of the chondroitin sulfate chain increased with aging. On the basis of gel chromatography on Bio-Gel A-1.5 m no significant difference in the approximate molecular size of chondroitin sulfate was observed between the young and old GAG samples. The present study indicated that the sulfation of chondroitin sulfate chains from rat costal cartilage decreased with the process of aging.
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PMID:The effect of aging on the synthesis of hexosamine-containing substances from rat costal cartilage. A decrease in sulfation of chondroitin sulfate with aging. 42 44

Glycosaminoglycans have been characterized from a normal human breast cell line (HBL-100) and two different cell lines from human breast carcinoma (MDA-MB-231 and MCF-7). The glycosaminoglycans were labeled by exposure of cell cultures to [3H]glucosamine and [35S]sulfate and then isolated from both spent media and cells by pronase digestion and cetylpyridinium chloride fractionation. They were further characterized by (a) hexosamine composition, (b) controlled-pore glass exclusion chromatography, (c) reactivity with specific enzymes (hyaluronidase chondroitinase, heparitinase, and heparinase), (d) nitrous acid degradation, and (e) DEAD-Sephadex chromatography. The results indicate that the HBL-100 line synthesizes mainly hyaluronic acid, most of which is secreted into the medium. Chondroitin sulfate and heparan sulfate are the predominant glycosaminoglycans synthesized by the cancer lines; both are found mainly in the spent medium, but the hyaluronic acid synthesized by the MDA-MB-231 line remains cell associated. The cell-associated heparan sulfate had a molecular weight in excess of 13,000 and may contain linkages susceptible to testicular hyaluronidase. The MCF-7 cells produce significantly lower amounts of glycosaminoglycans than do the other two lines.
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PMID:Glycosaminoglycans of normal and malignant cultured human mammary cells. 42 76

This study demonstrates that the dorsal ectoderm of the stage 14 chick embryo synthesizes hyaluronic acid. About 49 to 52% of the H3 glucosamine-labeled glycosaminoglycan that is synthesized by explanted ectoderm can be identified as hyaluronic acid on the basis of its susceptibility to Streptomyces hyaluronidase or isolation of chondroitinase ABC digestion products. In addition, autoradiographic evidence shows that the ectoderm, unlike adjacent tissues like epithelial somites or neural tube, incorporates glucosamine into hyaluronidase-sensitive material which becomes largely extracellular and localized in the subectodermal cell-free space. Ultrastructural evidence shows that there is a fine fibrillar matrix between the ectodermal cells and in the subectodermal spaces when tannic acid is included in the primary fixative. This material resembles authentic hyaluronate, similarly fixed, and is absent when tannic acid is omitted from the fixative or when embryos have been previously treated in ovo with Streptomyces hyaluronidase. The concomitant reduction in the intercellular and subectodermal cell-free spaces after in ovo treatment with Streptomyces hyaluronidase supports the hypothesis that the dorsal ectoderm plays a morphogenetic role by contributing hyaluronate to the forming extracellular spaces. It is proposed that ectodermally derived hyaluronate might influence the morphogenesis of subjacent tissues such as the dermatome and neural crest.
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PMID:The synthesis of hyaluronic acid by ectoderm during early organogenesis in the chick embryo. 47 12

The parietal pleura of rabbits was incubated with 14C-glucosamine. It was found that 14C-glucosamine was incorporated into the fraction of crude glycosaminoglycans. Then the crude glycosaminoglycans were fractionated by using specific mucopolysaccharide-lyases (hyaluronidase from streptomyces hyalurolytics, chondroitinase AC and chondroitinase ABC). As a result, evidence was obtained that hyaluronic acid was synthesized in parietal pleura and was released into the surroundings.
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PMID:[Biosynthesis of hyaluronic acid in parietal pleura of the rabbit (author's transl)]. 49 96

Rat ovarian granulosa cells were isolated from immature female rats after stimulation with pregnant mare's serum gonadotropin and then maintained in culture. Proteoglycans were labeled using [35S]sulfate, D-[3h]glucosamine, or L-[3H]serine as precursors. 35S-labeled proteoglycans in the medium increased linearly up to 72 h after a 6- to 8-h lag period, and those in a 4 M guanidine HCl extract of the cell layer increased for about 16 h and then reached a plateau and stayed fairly constant up to 72 h. Two distinct sizes of proteoglycans were observed in the medium. The smaller (Kav = 0.60 on Sepharose CL-2B) had lower buoyant densities in dissociative gradients (rho less than 1.4 g/ml). The larger (Kav = 0.26 on Sepharose CL-2B) had high buoyant densities (recovered mainly in the bottom (D1) fraction of the dissociative gradient). More than 90% of the D1 proteoglycans contained dermatan sulfate chains (average Mr = 38,000) which yielded 84% 4-sulfated and 15% disulfated disaccharides after digestion with chondroitinase ABC. About 8% of the 35S-label in D1 was present as a heparan sulfate proteoglycan. When [3H]-glucosamine was used as a precursor, 28% of the 3H activity in the D1 proteoglycans was located in three major oligosaccharide components, two of which were similar or identical with those observed previously in D1 proteoglycans isolated from porcine follicular fluid. These results plus similar susceptibility of the labeled proteoglycans to proteolytic enzymes, especially plasmin, suggest that the granulosa cells synthesize the predominant follicular fluid proteoglycans.
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PMID:Biosynthesis of proteoglycans by rat granulosa cells cultured in vitro. 50 Jul 20

The development of the sclerotome is considered as a model for the formation of mesenchyme from an epithelium. In early epithelial somites, transmission and scanning electron microscopy indicate considerable ultrastructural similarity between the future sclerotome and dermamyotomal regions. Subsequently, these two regions diverge in their development. In the forming dermamyotome, junctional complexes become more extensive and the cells become elongated, closely applied to each other, and have angular surface contours. In the forming sclerotome, there is an early reduction in apical junctions. The cells elongate, keeping their original polarity, and acquire numerous filopodia which contain punctate junctions at sites of cell-to-cell contact. Associated with cellular extension is an expansion of the intercellular spaces which do not contain any ultrastructurally recognizable material. Evidence for a role of hyaluronic acid in the expansion of the intercellular spaces is presented. As identified by the susceptibility of cetylpyridinium chloride precipitates to Streptomyces hyaluronidase and chromatographic separation of chondroitinase ABC digestion products, as much as 64--68% of the [3H]glucosamine-labeled glycosaminoglycans synthesized by explanted somites is hyaluronic acid. In addition, hyaluronidase-sensitive label is localized in the intercellular spaces of the sclerotome, as demonstrated by autoradiography. When Streptomyces hyaluronidase is injected in ovo into living embryos, the sclerotomal mesenchyme differentiates morphologically, but intercellular spaces are drastically reduced. It is hypothesized that the sclerotomal cells produce a hyaluronate-enriched extracellular matrix which is inflated by hydration to mediate the expansion of the sclerotomal mass towards the notochord.
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PMID:The role of extracellular matrix in the formation of the sclerotome. 52 73

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


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