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)

We have examined the adhesion of primary Sertoli cells to a seminiferous tubule basement membrane (STBM) preparation in vitro. The STBM isolation procedure (Watanabe, T.K., L.J. Hansen, N.K. Reddy, Y.S. Kanwar, and J.K. Reddy, 1984, Cancer Res., 44:5361-5368) yields segments of STBM that retain their histotypic form in both three-dimensional tubular geometry and ultrastructural appearance. The STBM sleeves contain two laminae: a thick, inner basal lamina that was formed in vivo between Sertoli cells and peritubular myoid cells; and a thinner, outer basal lamina that was formed between myoid cells and sinusoidal endothelial cells. Characterization by immunofluorescence and SDS PAGE revealed that the isolated STBM retained fibronectin, laminin, and putative type IV collagen among its many components. When the STBM sleeves were gently shaken with an enriched fraction of primary Sertoli cells, the Sertoli cells bound preferentially to the lumenal basal lamina at the ends of the STBM sleeves. Few Sertoli cells bound to either the outer basal lamina of the STBM sleeves or to vascular extracellular matrix material which contaminated the STBM preparation. 3T3 cells, in contrast, bound to all surfaces of the STBM sleeves. Pretreatment of the STBM sleeves with proteases, 0.1 M Na metaperiodate, 4 M guanidine HCl, or heating to 80 degrees-90 degrees C inhibited lumenal Sertoli cell binding, but binding was not inhibited by chondroitinase ABC, heparinase, hyaluronidase, or 4 M NaCl. The lumenal Sertoli cell binding occurred in the presence or absence of added soluble laminin, but not fibronectin. The addition of soluble laminin, but not fibronectin, restored random binding of Sertoli cells to trypsinized STBM sleeves. Our in vitro model system indicates that Sertoli cells recognize differences in two basal laminae produced in vivo on either side of myoid cells.
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PMID:Sertoli cell binding to isolated testicular basement membrane. 352 69

Rat serosal mast cells, which synthesize only heparin proteoglycans as detected by intrinsic labeling with [35S]sulfate, were analyzed for the presence of intracellular chondroitin sulfate proteoglycans by chemical and immunochemical means. Rat serosal mast cells of greater than 99% purity were treated with Zwittergent 3-12 and 4 M guanidine HCl, and the extracted nonradiolabeled proteoglycans were purified by density gradient centrifugation. As assessed by quantification of the unsaturated disaccharides released from the proteoglycans by chondroitinase ABC treatment, 10(6) rat serosal mast cells contained 2.4-4.5 micrograms of chondroitin sulfate proteoglycans. Analysis of the chondroitinase ABC digests by high performance liquid chromatography revealed the unsaturated disaccharides delta Di-4S, delta Di-diSB, and delta Di-diSE which were derived from GlcA----GalNAc-4-SO4, iduronic acid-2-SO4----GalNAc-4-SO4, and GlcA----GalNAc-4,6-diSO4, respectively. The molar ratio of the monosulfated to disulfated disaccharides was approximately 2:1 with delta Di-diSE greater than delta Di-diSB. When analyzed with a mouse anti-chondroitin sulfate monoclonal antibody and fluorescein-labeled F(ab')2 goat anti-mouse IgG, approximately 91% of permeabilized and chondroitinase ABC-treated cells in the mast cell preparations exhibited intracellular fluorescence, and the pattern of staining indicated that the chondroitin sulfate molecules were located in the secretory granules. The specificity of the monoclonal antibody for the unsaturated double bond created by chondroitinase ABC treatment of the proteoglycan in situ was established by the absence of fluorescence when the chondroitinase ABC step was omitted or when heparinase digestion was substituted for chondroitinase ABC. Furthermore, the ability of the anti-chondroitin sulfate monoclonal antibody to mediate fluorescence in situ was markedly reduced by absorption with solid-phase chondroitin sulfate proteoglycan that had been chondroitinase ABC-treated, but not by absorption with undigested proteoglycan or with solid-phase heparin. The highly sulfated chondroitin sulfate proteoglycans of rat serosal mast cells are the same type synthesized by the rat mucosal mast cell subclass.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Secretory granules of heparin-containing rat serosal mast cells also possess highly sulfated chondroitin sulfate proteoglycans. 353 Dec 3

The proteoglycans (PG) of bovine fetal tendon (4-8 months in utero) were extracted with 4 M guanidine HCl and partially purified by ion exchange chromatography. Proteoglycans from fetal tendon were virtually entirely small molecules (Kav approximately equal to 0.55 by Sepharose CL-4B chromatography). These small proteoglycans had dermatan sulfate glycosaminoglycan chains and a core protein (after chondroitinase ABC digestion) with Mr approximately equal to 45,000 on sodium dodecyl sulfate-polyacrylamide gels. By electrophoretic mobility, immunocross-reactivity, and V8 protease sensitivity, these proteoglycans were determined to be of both PG I and PG II types. In contrast, adult tendon contains only the PG II type of small proteoglycan. Proteoglycans synthesized by fetal tendon explant cultures were, by both chromatographic and electrophoretic mobilities, somewhat larger than those extracted from the same tissue. There was no difference in the spectrum of proteoglycans observed between those regions of fetal tendon destined to receive only tensional forces (proximal) and those regions that will be subjected as well to compressive forces (distal) in the adult. These observations indicate that the proteoglycan content and synthetic capability of all regions of fetal tendon are constant and significantly different from those of both the tensional and fibrocartilaginous regions of adult tendon.
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PMID:Proteoglycans of fetal bovine tendon. 365 32

Proteoglycans were extracted in good yield from the proximal, fibrous portion of adult bovine tendon with 4 m guanidine HCl. They comprise less than 1% of the dry weight of the tissue. Using CsCl density gradient centrifugation, gel chromatography, and ion exchange chromatography, two populations of proteoglycans were separated and purified from other tissue proteins. One was a large, chondroitin sulfate proteoglycan with high buoyant density in CsCl. This component appeared to be composed of two or three subpopulations as detected by agarose/polyacrylamide electrophoresis, although they could not be effectively separated from one another for individual characterization. As a group, the large proteoglycans eluted from Sepharose CL-2B with Kav from 0.1-0.5 and their core protein had Mr greater than 200,000 with high contents of glutamic acid, serine, and glycine. The glycosaminoglycan chains had a weight average Mr of 17,000 and more than 98% of the uronic acid was glucuronic acid. This group comprised only 12% of the total proteoglycan of the tissue. The other 88% of the proteoglycans appeared to represent one group of small molecules that eluted from Sepharose CL-2B at Kav = 0.70. They demonstrated buoyant densities in a CsCl gradient ranging from greater than or equal to 1.51 to 1.30 g/ml. Their core protein had an apparent Mr = 48,000 following removal of the glycosaminoglycan chains by digestion with chondroitinase ABC. This core protein had a particularly high content of aspartic acid/asparagine and leucine. The glycosaminoglycan chains had a weight average Mr of 37,000 and were dermatan sulfate containing 73% iduronic acid. Those molecules found at highest buoyant density appeared to have additional glycosaminoglycan chains that were shorter. Proteoglycans were also extracted from the pressure-bearing distal region of this tendon, where contents of proteoglycan per wet weight of tissue were 3-fold higher and as much as 50% of this was as large as the large proteoglycans from the proximal tissue. Preparations of large proteoglycans from both tendon regions contained molecules capable of interacting with hyaluronic acid.
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PMID:Characterization of proteoglycans from adult bovine tendon. 401 75

An explant culture of 15 cynomolgus monkey corneas was incubated with [35S]sulfate and [2-3H]mannose as labeling precursors. A 4 M guanidine HCl extract of the corneal stromas was prepared and combined with a 4 M guanidine HCl extract of stromas from 300 unlabeled corneas. The keratan sulfate proteoglycans in the combined extracts were purified by a combination of DEAE-cellulose chromatography, chondroitinase ABC digestion to remove chondroitin-dermatan sulfate proteoglycans, and elution from immobilized concanavalin A. The purified keratan sulfate proteoglycan was digested with papain and the digest was eluted on DEAE-Sephacel. The unbound fraction contained 59% of the 3H activity and consisted of intact oligosaccharide-peptides. The bound fraction, consisting of keratan sulfate chains linked to peptides, eluted during a linear 0-0.75 M NaCl gradient as a peak centered at approximately 0.6 M NaCl and contained 41% of the 3H and all of the 35S activity in the original proteoglycan. The chains were digested with endo-beta-galactosidase, and the digest was eluted on DEAE-Sephacel with a linear 0-0.75 M NaCl gradient. Most of the sulfated digestion fragments from the chains eluted as several distinct peaks during the gradient. All the 3H activity eluted in the unbound volume along with a small proportion of 35S activity. This unbound fraction was eluted on Bio-Gel P-10 to give a 3H peak (Kav = 0.46) well resolved from the remaining 35S activity which eluted near the total volume. This 3H peak contained the oligosaccharide-peptides derived from the linkage region between the keratan sulfate chains and the core protein. Structural analyses of the linkage region oligosaccharides and the intact oligosaccharides (Nilsson, B., Nakazawa, K., Hassell, J.R., Newsome, D.A., and Hascall, V.C. (1983) J. Biol. Chem. 258, 6056-6063) in combination with the 3H-labeling data suggest that the intact keratan sulfate proteoglycans contain an average of about one intact oligosaccharide per keratan sulfate linkage site.
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PMID:Purification of keratan sulfate proteoglycan from monkey cornea. 622 39

The biologic properties of two major proteoglycans of bovine aorta, heparan sulfate proteoglycan and chondroitin sulfate-dermatan sulfate proteoglycan were compared. The heparan sulfate proteoglycan was isolated either by elastase digestion or by 4.0 M guanidine hydrochloride extraction, of aorta tissue, fractionated by CsCl isopycnic centrifugation and purified by chondroitinase ABC treatment. The first method resulted in considerably greater yield (about 70% of the total heparan sulfate proteoglycan of the tissue) than the second procedure (12% of total). The chondroitin sulfate-dermatan sulfate proteoglycan was obtained by 4.0 M guanidine-HCl extraction of aorta tissue followed by CsCl isopycnic centrifugation. The chemical composition of both heparan sulfate proteoglycan preparations was similar. Unlike the chondroitin sulfate-dermatan sulfate proteoglycan, which eluted in the void volume of Sepharose CL-6B column, the heparan sulfate proteoglycan preparations were each resolved into a high molecular weight fraction (kav = 0.18 and 0.13) and a low molecular weight fraction (kav = 0.47 and 0.36). The heparan sulfate proteoglycan preparations exhibited significantly more potent anticoagulant and platelet aggregation inhibitory activities than the chondroitin sulfate-dermatan sulfate proteoglycan. The protein core of the proteoglycan molecules did not seem to be essential for their hemostatic properties. The complex forming ability of the heparan sulfate proteoglycan with serum low density lipoproteins (LDL) was much less than that of chondroitin sulfate-dermatan sulfate proteoglycan in the presence and absence of Ca2+. Interaction between heparan sulfate proteoglycan and LDL was also much more sensitive to changes in the ionic strength of the medium than that of chondroitin sulfate-dermatan sulfate proteoglycan and the lipoprotein. Since the total sulfate content of both proteoglycans is almost similar, the smaller molecular size and hence the lower overall charge density of the heparan sulfate proteoglycan appears to be partly responsible for its low affinity for LDL. The differences in biologic properties of the two proteoglycans might have implications in the pathophysiology of cardiovascular diseases.
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PMID:Hemostatic properties and serum lipoprotein binding of a heparan sulfate proteoglycan from bovine aorta. 622 69

The structural and immunological properties of the glycosaminoglycans and the core proteins of bovine nasal cartilage proteoglycan and the proteoglycans produced by rabbit articular chondrocytes in spinner and monolayer culture were compared. Culture medium with 35SO4- or 3H-serine-labeled proteoglycan was mixed with bovine nasal cartilage 4M guanidine-HCl extract and digested with trypsin. The proteoglycan fragments were then isolated by DEAE-cellulose chromatography and fractionated by dissociative CsCl density gradient centrifugation. Approximately 90% of the 35SO4 incorporated into proteoglycan by the cultured chondrocytes was in chondroitin sulfates and about 5% in keratan sulfate. Although there was considerable overlap in the Sepharose 4B elution of the tryptic proteoglycan fragments of highest buoyant density, some monolayer-produced proteoglycan fragments eluted earlier and some spinner-produced proteoglycan fragments eluted later than the proteoglycan fragments from bovine nasal cartilage. These differences in apparent fragment size could relate to differences in glycosaminoglycan chain length, since the glycosaminoglycans released by treatment with alkali from monolayer-produced proteoglycan in part eluted from Sepharose 4B earlier and those from spinner-produced proteoglycan in part eluted later than the chondroitin sulfate chains released from bovine cartilage proteoglycan. After digestion with chondroitinase ABC, 3H-serine-labeled high density tryptic proteoglycan fragments from monolayer and spinner culture yielded Sepharose 6B elution profiles which were similar to each other but did not coincide with the peaks of carbazole reactivity found with similarly treated fragments of bovine nasal cartilage proteoglycan. Cross-reactivity was demonstrated by radioimmunoautography between bovine cartilage and rabbit chondrocyte proteoglycan fragments restricted to gradient fractions of low buoyant density, but immunological cross-reactivity was not found for the antigens associated with the keratan sulfate-rich and chondroitin sulfate-bearing tryptic fragments of bovine nasal cartilage proteoglycan. These studies indicate that the proteoglycan core proteins produced by rabbit articular chondrocytes in monolayer and spinner culture are, in part, different from the core protein of bovine nasal cartilage proteoglycan and that the three proteoglycans differ in the length of some of their chondroitin sulfate chains.
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PMID:A comparison of the proteoglycans produced by rabbit articular chondrocytes in monolayer and spinner culture and those of bovine nasal cartilage. 622 50

Glycosaminoglycan (GAG) was extracted from the porcine thyroid gland with a buffer containing 5.3 M guanidine-HCl and proteolytic enzyme inhibitors and was fractionated by subsequent isodensity CsCl centrifugation. 60% of uronic acid positive materials was accumulated in the bottom one-fourth fraction with high buoyant density. More than 90% of this uronic acid positive material in the thyroid tissue was heparin or heparan sulfate (sensitive to nitrous acid treatment) and the rest was chondroitin sulfate or dermatan sulfate (sensitive to chondroitinase ABC treatment). When the accumulated high buoyant density GAG was analyzed on a Sepharose CL-6-B column, approximately 14% of the heparin sulfate were in the macromolecular portion as a form of proteoglycan because it was destroyed by the papain digestion or alkaline borohydride treatment which extensively digests protein or releases GAG from protein by the elimination reaction, respectively. This study demonstrates the existence of heparin sulfate proteoglycan in thyroid tissue for the first time.
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PMID:Presence of heparan sulfate proteoglycan in thyroid tissue. 623 Nov 80

Retinas were labeled in culture with [3H]glucosamine or [3H]leucine and [35S]sulfate and extracted sequentially with physiologically balanced saline and 4 M guanidine HCl. They were dialyzed into associative conditions (0.5 M NaCl) and chromatographed on agarose columns. Under these conditions, some of the proteoglycans were associated in massive complexes that showed low buoyant densities when centrifuged in CsCl density gradients under dissociative conditions (4 M guanidine HCl). Much of the label in these complexes was in molecules other than proteoglycans. Most of the proteoglycans, however, were included on the agarose columns, where they appeared to be constitutionally of low buoyant density. They resisted attempts to separate potential low buoyant density contaminants from the major proteoglycans by direct CsCl density gradient centrifugation or by the fractionation of saline or 8 M urea extracts on diethylaminoethyl-Sephacel. The diethylaminoethyl-Sephacel fractions were either subjected to CsCl density gradient centrifugation or were chromatographed on Sephacryl S-300, in both cases before and after alkaline cleavage, to confirm the presence of typical O-linked glycosaminoglycans. The medium and balanced salt extracts were enriched in chondroitin sulfate and other sulfated macromolecules, possibly highly sulfated oligosaccharides, that resisted digestion by chondroitinase ABC but were electrophoretically less mobile than heparan sulfate. Guanidine HCl or urea extracts of the residues were mixtures of high and low density proteoglycans that were enriched in heparan sulfate.
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PMID:Low buoyant density proteoglycans from saline and dissociative extracts of embryonic chicken retinas. 636 28

Heparan sulfate fractions were isolated from three normal human livers and three cancerous human liver tissues, and their polyanionic properties were examined using electrophoresis, sequential partition fractionation, and chemical analyses. More than 60% of total glycosaminoglycans from normal human liver and about 30% from cancerous liver tissue were found to be heparan sulfate from their resistance to exhaustive digestion with chondroitinase ABC and their susceptibility to nitrous acid treatment. The heparan sulfate isolated from cancerous liver tissue afforded a lower sulfate/uronic acid molar ratio (0.58 to 0.65) than did normal human liver heparan sulfate (0.76 to 0.80). Also, the former showed lower electrophoretic mobility in 0.1 M HCl and a different partition fractionation profile in comparison with the latter. These differences in charge density of the macromolecule were not detected on the chondroitin sulfate and/or dermatan sulfate fractions isolated from normal human liver and cancerous liver tissue.
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PMID:Changes in charge density of heparan sulfate isolated from cancerous human liver tissue. 644 94


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