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)

Monoclonal antibodies have been developed that recognize epitopes in native chondroitin sulfate chains. One of these antibodies, CS-56, reportedly recognizes chondroitin 4- and 6-sulfates. However, this antibody, and four other anti-chondroitin sulfate antibodies, 4C3, 4D3, 6C3 and 7D4, do not recognize epitopes in chondroitin sulfate chains from Swarm rat chondrosarcoma proteoglycan, an indication that native chondroitin sulfate epitopes are more structurally complex than the standard 0-, 4-, and 6-sulfated disaccharide repeats that constitute the backbone of chondroitin sulfate chains. A series of limited chondroitinase digestions was performed on the large aggregating proteoglycan monomer extracted from embryonic chick chondrocyte cultures to identify the digestion parameters required to release the different native chondroitin sulfate epitopes. Some epitopes were more accessible to enzymatic digestion than other epitopes. The approximate location of epitopes was determined by measuring the size of undigested oligosaccharides retained on the core protein following a limited digestion, and correlating this with the level of immunoreactivity for the different antibodies. These analyses identified the locations of three different antigenic domains. Domain 1 resides at the linkage region and contains epitopes for two of the five antibodies, and a portion of the epitopes for a third antibody. Domain 2 lies in the interior of the chain and contains epitopes for three of the five antibodies. Domain 3 resides at the non-reducing terminus and does not contain epitopes for any of the anti-chondroitin sulfate antibodies used in this study. These results indicate that specific native chondroitin sulfate epitopes are non-randomly distributed within the linear framework of chondroitin sulfate chains.
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PMID:Structural domains in chondroitin sulfate identified by anti-chondroitin sulfate monoclonal antibodies. Immunosequencing of chondroitin sulfates. 750 64

We report herein identification of a novel ligand for CD44, a cell surface glycoprotein implicated in tumor metastasis, lymphocyte differentiation and homing. A mouse T cell line CTLL-2 transfected with cDNA encoding a hemopoietic form of mouse CD44 exhibited a new self-adhesive phenotype, forming large aggregates. The aggregation was blocked by anti-CD44 mAb but little affected by hyaluronidase, indicating the involvement of CD44 and its non-hyaluronate ligand in the cell aggregation. The ability to induce CD44-dependent aggregation was observed in culture supernatants of CTLL-2 and its CD44 transfectants. Immunoprecipitation analysis using a CD44-Ig chimeric molecule indicated that CTLL-2 and its transfectants synthesized a macromolecule (gp600) which bound specifically to CD44. gp600 was readily labeled with radioactive sulfate and treatment of gp600 with chondroitinase ABC or AC II generated a lower molecular weight species (18-22 kDa), suggesting that gp600 consists of a small core protein heavily modified with chondroitin sulfate glycosaminoglycan side chains. However, when binding of CD44 was tested in vitro to chondroitinase-sensitive purified glycosaminoglycans, such as chondroitin-4-sulfate, chondroitin-6-sulfate and dermatan sulfate, no binding was demonstrable, suggesting either that a novel type of chondroitinase-sensitive glycosaminoglycan is recognized by CD44 or that association of the glycosaminoglycan with a core protein is required for recognition by CD44.
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PMID:A novel ligand for CD44 is sulfated proteoglycan. 751 79

We have identified a novel ligand for CD44, a cell surface glycoprotein implicated in tumor metastasis and lymphocyte homing. When the mouse T cell line CTLL-2 was transfected with cDNA encoding a hemopoietic form of mouse CD44, CTLL-2 cells exhibited a new self-adhesive phenotype, forming large aggregates. The aggregation was blocked by neutralizing anti CD44 monoclonal antibody but unaffected by hyaluronidase, indicating the involvement of CD44 and its non-hyaluronate ligand in the cell aggregation. The ability to induce CD44-dependent aggregation was found in culture supernatants of CTLL-2 and its CD44 transfectants. The use of CD44-immunoglobulin chimeric protein revealed that CTLL-2 and its transfectants synthesized a large-molecular weight protein (gp600) which bound specifically to CD44. The gp600 was readily labeled with radioactive sulfate, and treatment of gp600 with chondroitinase ABC or ACII generated a lower molecular weight species (18-22 kDa), suggesting that gp600 consists of a small core protein with chondroitin sulfate glycosaminoglycan side chains. However, binding of CD44 to glycosaminoglycans such as chondroitin 4-sulfate, chondroitin 6-sulfate, and dermatan sulfate was undetectable, suggesting either that a novel chondroitin-type glycosaminoglycan is recognized by CD44 or that a particular configuration of the glycosaminoglycan is required for recognition by CD44.
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PMID:A sulfated proteoglycan as a novel ligand for CD44. 2292 40

Proteoglycans metabolically labelled with [35S]sulphate and [3H]glucosamine or [3H]leucine were isolated from the incubation medium and cell layer of human adult mesangial cells and glomerular visceral epithelial cells using sequential DEAE chromatography purification steps followed by gel-filtration chromatography. The proteoglycan composition of each peak was analysed by treatment with HNO2, chondroitinase ABC or chondroitinase AC followed by chromatography on Sephadex G-50 columns. Heparan sulphate proteoglycan (HSPG) and dermatan sulphate proteoglycan were detected in both the culture medium and cell layer of mesangial cells. Culture medium of glomerular visceral epithelial cells contained HSPG and a second proteoglycan with the properties of a hybrid molecule containing HS and chondroitin sulphate (CS). The cell layer contained HSPG and CSPG. Detailed analysis of the hybrid molecule revealed that it had an apparent molecular mass of 400 kDa. SDS/PAGE of hybrid molecules, after treatment with heparitinase and chondroitinase ABC, revealed a core protein of 80 kDa. Using 1.8% polyacrylamide/0.6% agarose-gel electrophoresis, we deduced that the HS and CS were independently attached to one core protein. Because glomerular-basement-membrane HSPG is thought to be derived from mesangial cells and glomerular visceral epithelial cells and this molecule is involved in several kidney diseases, we investigated its synthesis in more detail. Anti-(rat glomerular-basement-membrane HSPG) monoclonal antibodies (JM403) and anti-(human glomerular-basement-membrane HSPG) polyclonal antibodies (both antibodies known to react with the large basement-membrane HSPG, perlecan) reacted strongly with HSPG obtained from both mesangial cells and glomerular visceral epithelial cells. However, the hybrid molecule did not react with these antibodies, suggesting that the HS side chain and the core protein were different from glomerular-basement-membrane HSPG. To quantify HS we performed an inhibition ELISA using mouse antibodies specific for glomerular-basement-membrane HS glycosaminoglycan side chains. Glomerular visceral epithelial cells produced significantly higher levels of HS (between 197.56 and 269.40 micrograms/72 h per 10(6) cells) than mesangial cells (between 29.8 and 45.5 micrograms/72 h per 10(6) cells) (three different cell lines; n = 3; P < 0.001). HS production by these cells was inhibited by cycloheximide, revealing that it was synthesized de novo. Expression of perlecan mRNA, demonstrated using reverse transcriptase PCR, was different in the two cell types. We conclude that glomerular visceral epithelial cells and mesangial cells have characteristic patterns of proteoglycan production. Glomerular visceral epithelial cells produced a hybrid proteoglycan containing CS and HS independently attached to its core protein.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Proteoglycan production by human glomerular visceral epithelial cells and mesangial cells in vitro. 753 59

After transection of adult mouse sciatic nerve, the expression of a chondroitin sulphate epitope recognized by the monoclonal antibody 473-HD (mAb 473-HD) was found to be up-regulated. The epitope was localized immunocytochemically mainly in Schwann cell basal laminae and, more weakly, also in the endoneurium. In cultures of mouse dorsal root ganglion cells, Schwann cells expressed high levels but fibroblasts only low levels of the epitope. To identify the molecule(s) carrying this chondroitin sulphate epitope, human sciatic nerves were extracted with phosphate-buffered saline and shown to contain two chondroitin sulphate proteoglycans of apparent molecular weights of 130 and 900 kDa. The 900 kDa and, more weakly, the 130 kDa proteoglycan were reactive with mAb 473-HD, which was found to recognize chondroitin-6-sulphate as epitope. Following chondroitinase ABC treatment of the 130 kDa proteoglycan, a core protein of approximately 45 kDa was seen and shown to react with polyclonal antibodies against the chondroitin-dermatan sulphate proteoglycan decorin from human fibroblasts. Chondroitinase ABC treatment of the 900 kDa proteoglycan yielded a core protein with a molecular weight of approximately 400 kDa that was recognized by polyclonal antibodies against recombinantly expressed fusion proteins from human versican. After transection of adult mouse sciatic nerves, the distal nerve stumps showed up-regulation of the chondroitin-6-sulphate epitope of the 900 kDa proteoglycan, whereas the core protein of this proteoglycan did not show any detectable change in the level of expression. In contrast, the core protein of the 130 kDa proteoglycan was up-regulated in expression. These observations suggest that versican- and decorin-like molecules may contribute to successful regeneration in the peripheral nervous system of mammals.
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PMID:Up-regulation of a chondroitin sulphate epitope during regeneration of mouse sciatic nerve: evidence that the immunoreactive molecules are related to the chondroitin sulphate proteoglycans decorin and versican. 754 29

Immunological and biochemical characteristics of a 100,000 MW biglycan-like haemopoietic factor, purified from thymic myoid cells 871207B, were studied to distinguish them from macrophage colony-stimulating factor (M-CSF), which they resemble in activity and biochemical properties. Rabbit antibody raised against a synthetic peptide fragment (J-1) designed from amino acid sequences specific to the 100,000 MW factor responded to 871207B cells, the conditioned medium of 871207B, and capillary-like structures in the thymus, but not to M-CSF producer L-929 cells or the conditioned medium of L-929 cells. In contrast, M-CSF epitope was detected in L-929 cells and the conditioned medium cells but not in 871207B cells or the conditioned medium, even after enzymatic digestion of glycosaminoglycan chains. Treatment of the 100,000 MW factor with chondroitinase ABC and AC produced a 50,000 MW component. Digestion of this product with N-glycanase resulted in a 40,000 MW protein component. These results suggest that the 100,000 MW factor is a proteoglycan consisting of a core protein with an apparent molecular mass of 40,000 MW, a 50,000 MW chondroitin sulphate chain and 10,000 MW N-linked oligosaccharide chains. A small amount of a 40,000 MW monocytic cell growth activity was also found in the 871207B cell-conditioned medium. An enzymatically obtained 40,000 MW factor, the conditioned medium 40,000 MW factor, and the 100,000 MW factor were specifically eluated from an anti-J-1 IgG-immobilized affinity column with monocytic cell growth activity, suggesting that the biological activity resides in the 40,000 MW core protein. The 100,000 MW factor induced the proliferation and differentiation of monocytic lineage cells from a variety of sources, such as bone marrow cells, peritoneal exudated cells and brain microglia cells.
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PMID:Immunological and biochemical characterization of biglycan-like haemopoietic factor. 754 45

Following injury to the adult CNS, the expression of a number of extracellular matrix molecules increases in regions of reactive gliosis. This glial matrix includes certain chondroitin sulfate proteoglycans (CS-PGs) which have been correlated with an inhibition of axon outgrowth. In order to test the influence of glial associated CS-PGs on neurite elongation directly, we sought to determine whether enzymatic modification of injury-induced CS-PGs could enhance neurite outgrowth across the surface of intact glial scars formed in vivo after implanting nitrocellulose filters into the cortex of adult rats. This gliotic tissue was subsequently explanted in vitro and used as a substrate for growing embryonic retinal neurons. Treatment of adult explants with chondroitinase ABC led to a significant increase in mean neurite length over the scar surface. Heparitinase treatment caused a much smaller, although significant, increase in neurite outgrowth. This suggested that more than one type of PG was present or that a single PG with both CS and HS side chains was upregulated. Western analysis revealed that a PG(s) with a core protein between 180 and 400 kDa was found to be relatively more abundant in areas of reactive gliosis induced to form in adult rather than neonatal animals. Simultaneous treatment of adult glial scars with chondroitinase and antibodies to the beta 1, beta 2 chain of laminin partially reversed the growth-enhancing effect of enzymatic digestion alone. These data demonstrate that the increase in neurite outgrowth along the surface of reactive astrocytes following enzymatic modification of injury-induced PGs was due, in part, to the presence of laminin. Thus, in this model of gliosis, particular PGs may act as inhibitors of neurite outgrowth by attenuating the potential for axon elongation that could occur due to the concomitant expression of growth-promoting molecules in regions of reactive gliosis.
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PMID:Injury-induced proteoglycans inhibit the potential for laminin-mediated axon growth on astrocytic scars. 758 32

The binding of the amyloid protein precursor (APP) to heparan sulfate proteoglycans has been shown to stimulate the neurite-promoting activity of APP. In this study, proteoglycans that bind with high affinity to APP were characterized. Conditioned medium from cultures of postnatal day 3 mouse brain cells was applied to an affinity column containing a peptide homologous to a heparin-binding domain of APP. A fraction 17-fold enriched in proteoglycans was recovered by elution with a salt gradient. APP bound saturably and with high affinity to the affinity-purified proteoglycan fraction. Scatchard analysis of the binding showed that APP bound to high- and low-affinity sites with equilibrium dissociation constants of 1.4 x 10(-11) and 6.5 x 10(-10) M, respectively. APP, in conjunction with the affinity-purified proteoglycan fraction, promoted neurite outgrowth. The affinity-purified proteoglycan fraction contained a heparan sulfate proteoglycan and a chondroitin sulfate proteoglycan. Digestion of the affinity-purified fraction with heparitinase I revealed a core protein of 63-69-kDa molecular mass, whereas digestion with chondroitinase ABC revealed a core protein of 100-110 kDa. The results suggest that expression of specific APP-binding proteoglycans may be an important step in the regulation of the neurite outgrowth-promoting activity of APP.
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PMID:Affinity purification of proteoglycans that bind to the amyloid protein precursor of Alzheimer's disease. 759 8

The Alzheimer amyloid precursor (APP) protein is a member of a family of glycoproteins that includes the amyloid precursor-like proteins (APLPs). Previously, we showed that in C6 glioma cell cultures, secreted APP nexin II occurs as the core protein of a chondroitin sulfate proteoglycan (CSPG). Here, we report that among seven untransfected cell lines, expression of secreted APP CSPG was restricted to two cell lines of neural origin, namely, C6 glioma and Neuro-2a neuroblastoma (N2a) cells. Addition of dibutyryl cyclic AMP in N2a cultures, a treatment that induces the neuronal phenotype in these cells, resulted in a significant reduction in the amount of the secreted APP CSPG, although secretion of APP was only marginally affected. Growth in the presence of serum increased the size of the secreted APP CSPG, suggesting that the number and/or length of the chondroitin sulfate (CS) chains attached to the core APP varies with growth conditions. Extensive mapping with epitope-specific antibodies suggested that a CS chain is attached within or proximal to the A beta sequence of APP. In contrast to the restricted expression of the APP CSPG, expression of secreted APLP2 CSPGs was observed in all cell lines examined. After chondroitinase treatment, two core proteins of approximately 100 and 110 kDa were obtained that reacted with an APLP2-specific antiserum, suggesting that non-transfected cell lines contain at least two endogenous APLP2 CSPGs, probably derived by alternative splicing of the APLP2 KPI domain. The fraction of the APLP2 proteins in the CSPG form was dependent on the particular cell line examined.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Expression of the chondroitin sulfate proteoglycans of amyloid precursor (appican) and amyloid precursor-like protein 2. 761 33

A chondroitin sulfate-dermatan sulfate proteoglycan was isolated from bovine aorta intima by extraction of the tissue with 4 M guanidine hydrochloride. The proteoglycan was purified by CsCl isopycnic centrifugation followed by gel filtration and ion exchange chromatography. A monoclonal antibody C8F4 was developed to this core protein. The characteristics and specificity of the antibody were studied by an enzyme-linked immunosorbent assay (ELISA) using an alkaline phosphatase conjugated antibody (goat anti-mouse IgG). The antibody binding to the core protein was found specific and optimal at pH 7.0. The antibody recognizes either intact chondroitin sulfate-dermatan sulfate proteoglycan monomer, chondroitinase ABC digested monomer or chemically deglycosylated proteoglycan. Free chondroitin sulfates, keratan sulfate and hyaluronic acid did not compete for the antigenic sites in ELISA. Limited hydrolysis of the core protein by trypsin resulted in three peptides and only the peptide with a molecular weight M(r) = 40,000 was found capable of binding to hyaluronic acid. The antibody C8F4 recognized this hyaluronic acid binding peptide but did not recognize the other two peptides suggesting that the epitope(s) for this antibody is in the hyaluronic acid-binding region of the core protein. The antibody recognized the core proteins from bovine nasal cartilage proteoglycan and human aorta proteoglycan but did not recognize bovine aorta link protein, bovine serum albumin, human serum albumin, human transferrin, collagen Type I and fibronectin. The antibody was found useful to localize proteoglycans in atherosclerotic lesions in human aorta by immunohistochemical techniques.
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PMID:A monoclonal antibody that recognizes hyaluronic acid binding region of aorta proteoglycans. 768 Dec 90


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