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)

To study the molecular structure and function of bovine skin proteodermatan sulfate, on a determinant by determinant basis, several monoclonal antibodies to this molecule have been produced and characterized. Based on the results of a preliminary immunogenetic analysis of 4 inbred mouse strains, SJL/J (H-2s) mice were immunized for the fusions. Ten hybridomas were produced and the monoclonal antibodies from four of these were selected for further investigation. Employing an ELISA inhibition assay, none showed any detectable affinity for bovine collagen types I, II, III, or IV, bovine fibronectin or chondroitin or dermatan sulfate glycosaminoglycans. Each monoclonal antibody bound the chondroitinase ABC-derived protein core and none was significantly inhibited by proteinase digests of the intact molecule suggesting that the epitope of each contains a protein component. The results of competitive binding ELISA assays and immunoblots of the cyanogen bromide cleavage products of proteodermatan sulfate indicate that the 4 antibodies recognize at least 3 distinct antigenic determinants on this molecule. Immunohistochemical methods located the antigen in the dermis of bovine skin and revealed that a change in proteodermatan sulfate distribution occurs during skin development.
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PMID:Production and characterization of monoclonal antibodies to bovine skin proteodermatan sulfate. 257 62

The low molecular weight proteoglycan fraction extracted from articular discs with 4 M guanidinium chloride was found to consist predominantly of an iduronate-rich dermatan sulphate proteoglycan, together with chondroitin sulphate-containing material. The dermatan sulphate proteoglycan was purified by ion-exchange and gel-filtration chromatography and its core protein isolated after digestion with chondroitinase ABC. The amino acid composition and pattern of cyanogen bromide peptides obtained from this core were closely similar to those of the protein core of bovine skin proteodermatan sulphate. Four monoclonal antibodies raised against bovine skin proteodermatan sulphate also reacted with the disc protein core and its cyanogen bromide peptides. Results of digestion with glycopeptidase F demonstrated the presence of three N-linked oligosaccharides. The combined size of these oligosaccharides appeared to be somewhat less than the size of those on skin proteodermatan sulphate. The glycosaminoglycan chain released by digestion with cathepsin C had a higher molecular weight than that from skin. These differences in glycosylated structures may be responsible for the different effects on collagen fibrillogenesis in vitro; whereas skin proteodermatan sulphate only reduced the rate of fibril growth, disc dermatan sulphate proteoglycan also increased the length of the lag-phase and the final opacity.
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PMID:Proteoglycans of the articular disc of the bovine temporomandibular joint. II. Low molecular weight dermatan sulphate proteoglycan. 279 47

Cyanogen bromide treatment of bovine nasal cartilage proteoglycan monomer gave rise to three major fractions (CN-1 to CN-3), isolated by Sepharose CL-6B chromatography. The uronate-rich fraction in the void volume (CN-1) digested with chondroitinase ABC (C treatment) yielded a fragment (CN-1 C/6B) with a unique N-terminal sequence. The same fraction, when digested sequentially by chondroitinase ABC and trypsin (CT treatment), was resolved into two distinct fractions, CN-1 CT/6B-1 and CN-1 CT/6B-2. CN-1 CT/6B-1 consisted in a keratan sulfate-rich region, representing the N-terminal moiety of the CN-1 fraction; these data suggested, according to the model of the proteoglycan monomer structure described by Heinegard, D. and Axelsson, I. (1977) J. Biol. Chem. 252, 1971-1979, that its C-terminal moiety is localized at the end of the core bearing the chondroitin sulfate chains. CN-1 CT/6B-2 contained two fragments from the chondroitin sulfate-bearing region: one of them has been submitted to Edman degradation. The CN-2 fraction upon chondroitinase and trypsin treatments gave rise to a keratan-bearing region (CN-2 CT/6B-1) and a mannose-rich region (CN-2 CT/6B-2). After reduction and alkylation of CN-2, the N-terminal sequence of the isolated major fragment (CN-2 RA/6B-1) was determined. The CN-3 fraction revealed a pattern upon electrophoresis similar to that of the cyanogen bromide-treated hyaluronic acid-binding region.
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PMID:Identification of cyanogen bromide-fragments of the protein core of bovine nasal cartilage proteoglycan monomer. 682

Capillary electrophoresis based on cetyltrimethylammonium bromide micellar electrokinetic capillary chromatography (MECC) was developed for the separation and determination of glycosaminoglycan (GAG) disaccharide units without derivatization. The influence of changes in several separation conditions was studied, and the separation mechanisms are discussed. Tests of repeatability and linearity were performed for qualitative and quantitative evaluation of the method. The described procedure gives a rapid and efficient determination of GAG disaccharides. Samples of chondroitin sulphates and mink skin were treated with proteases, and the extent of protein cleavage was followed by free zone capillary electrophoresis. The result of the chondroitinase ABC treatment following the protease treatment was evaluated by the MECC method.
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PMID:Separation and determination of glycosaminoglycan disaccharides by micellar electrokinetic capillary chromatography for studies of pelt glycosaminoglycans. 828 40

The extracellular matrix of the mineralizing eggshell contains molecules hypothesized to be regulators of biomineralization. To study eggshell matrix molecules, a bank of monoclonal antibodies was generated that bound demineralized eggshell matrix or localized to oviduct epithelium. Immunofluorescence staining revealed several staining patterns for antibodies that recognized secretory cells: staining for a majority of columnar lining cells, staining for a minor sub-set of columnar lining cells, intensified staining within epithelial crypts, and staining of the entire tubular gland. Western blotting with the antibody Epi2 on eggshell matrix showed binding to molecules with the apparent molecular weight of eggshell matrix dermatan sulfate proteoglycan (eggshell DSPG). Immunoblots of cyanogen bromide-cleaved eggshell DSPG revealed broad band of reactivity that shifted to 25 kDa after chondroitinase digestion; indicating that the Epi2 binding site is located on a fragment which contains dermatan sulfate side chains. Immunogold labeling showed that Epi2 binds to secretory vesicles within the non-ciliated cells of the columnar epithelium, while the antibodies Tg1 and Tg2 bind to secretory vesicles of tubular gland cells. Immunogold labeling of demineralized shell matrix showed binding of Epi2, Tg1, and Tg2 to the matrix of the palisade layer, and showed little reactivity to other regions of the shell matrix. Quantification of the immunogold particles within the eggshell matrix revealed that antibodies Epi2 and Tg1 bind all calcified regions equally while antibody Tg2 has a greater affinity for the baseplate region of the calcium reserve assembly.
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PMID:Monoclonal antibodies to mineralized matrix molecules of the avian eggshell. 1110 57

The fluoroquinolone, ofloxacin, a widely used antimicrobial agent, has been shown to cause athropathogenic syndromes in juvenile animals. In the present study, the effect of ofloxacin on chondrogenesis in cartilage organoid cultures of limb-bud mesenchymal cells obtained from day-12 mouse embryos was investigated. Cultures treated with increasing concentrations of ofloxacin (10, 30 and 100 mug/ml medium) for 6 days showed no significant changes in overall protein content and dry weight. Collagen type II, as a specific marker for cartilage, and collagen type I, as a marker protein in the internodular loose connective tissue in this culture system, were estimated by an inhibition-ELISA after cyanogen-bromide digestion. The collagen type I content of treated cultures remained constant, but the collagen type II level decreased in a dose-dependent manner to about 40% of that of the controls. There was no detectable increase in the concentration of collagen type II in the medium suggesting that ofloxacin inhibits synthesis rather than stimulate degradation of collagen type II. Cultures treated with 100 mug ofloxacin/ml were further investigated by indirect immunofluorescence and electron microscopy. Anticollagen type II antibodies demonstrated irregularities, several defects in the cartilage nodules, and much weaker staining in the treated cultures compared with controls. Similar results were obtained with antibodies directed against the core protein of large chondroitin sulphate proteoglycan monomers. Using monoclonal antibodies specific for unsulphated, 4-sulphated and 6-sulphated disaccharide "stubs" that remain attached to the core protein after chondroitinase ABC digestion of proteoglycans, different changes in these glycosaminoglycans were observed. While unsulphated chondroitin seemed to disappear nearly completely from the cartilage matrix, the level of chondroitin 4-sulphate remained unchanged and in the case of chondroitin 6-sulphate a slight increase in staining intensity was observable in the ofloxacin-treated cultures. Ultrastructurally, there was a reduction in the number of collagenous fibrils in the cartilage matrix of treated cultures and necrotic chondroblasts could be demonstrated. The present results resemble, in some aspects, observations that have been made in vivo after ofloxacin treatment, indicating that this in vitro model may provide a suitable system for examining the mechanism of quinolone-induced athropathia.
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PMID:Effects of ofloxacin on chondrogenesis in murine cartilage organoid culture. 2073 46