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:3.2.1.36 (hyaluronidase)
4,606 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chondroitinase B and chondroitinase C were separated from an extract of Flavobacterium heparinum induced with chondroitin 6-sulfate by using column chromatography on hydroxylapatite. Chondroitinase C was eluted together with the activities of hyaluronidase, delta4,5glycosiduronase, and sulfatase. The latter two activities were eliminated exclusively by passing the crude chondroitinase C fraction through a phosphono-cellulose column pre-equilibrated with 0.07M sodium phosphate buffer (pH 6.8). Chondroitinase C was then purified by affinity chromatography using dermatan sulfate-bound AH-Sepharose 4B coated with the same glycosaminoglycan. Purification of the enzyme was achieved 18-fold and in 73% yield. On the other hand, the activities of delta4,5glycosiduronase and sulfatase were decreased to 50 and 60%, respectively, as compared with those in the crude chondroitinase B fraction, after passing the fraction through a column of phosphono-cellulose pre-equilibrated with 0.1M sodium phosphate buffer (pH 6.8). The remaining activities of these two enzymes were then eliminated from chondroitinase B by affinity chromatography with heparin-bound AH-Sepharose 4B coated with dermatan sulfate. In the affinity chromatography used in the present study, non-covalent coating of the glycosaminoglycan-bound (covalently) AH-Sepharose 4B with the same or another glycosaminoglycan was found to be important.
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PMID:Purification of chondroitinase B and chondroitinase C using glycosaminoglycan-bound AH-Sepharose 4B. 42 37

The specific binding and nature of the epitope recognized by monoclonal antibody (Mab) 1H10, which binds an antigen expressed on human cervical tumors, was characterized by enzyme digestion, lectin competition assay and immuno-electron microscopy. Membrane homogenates of CaSki cervical carcinoma cells were digested with various enzymes, then analysed by SDS-PAGE and immunoblotting. Cells grown on coverslips were treated with various enzymes and in situ binding of Mab 1H10 to cells was analysed by electron microscopy. The ability of lectin-conjugates to block Mab 1H10 binding to CaSki cells was also examined. Treatment of samples with sodium periodate abrogated antigen recognition by Mab 1H10. Neuraminidase and hyaluronidase digestion decreased but did not eliminate Mab 1H10 binding to cells in situ. Chondroitinase ABC digestion, in contrast, removed Mab 1H10 binding sites both in vitro and in situ. Trypsin and chymotrypsin digestion of cell membrane homogenates decreased the molecular weight of the Mab 1H10 antigen but did not decrease the binding intensity. Wheat germ agglutinin (WGA) strongly bound to CaSki cells and partially blocked Mab 1H10 binding, indicating that the antigen contains N-acetyl-galactosamine residues at or near the epitope recognized by Mab 1H10. Ricinus communis agglutinin (RCA) exhibited a similar binding pattern to WGA. However, concanavalin A bound only weakly to CaSki cells and was ineffective at blocking Mab 1H10 binding. The tumor-associated antigen recognized by Mab 1H10 is concluded to be a chondroitin sulphate glycoprotein or proteoglycan rather than a mucopolysaccharide or lipoprotein.
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PMID:Characterization of a human cervical carcinoma-associated antigen by lectin binding and immuno-electron microscopy. 142 5

Proteoglycans (PGs) from human burn hypertrophic scar of a patient with Ehlers-Danlos syndrome were extracted with 4M guanidinium chloride and purified by DEAE-cellulose chromatography. Differential ethanol precipitation of the PG fraction obtained after ion-exchange chromatography yielded two low mol.-wt. PGs, on rich in glucuronic acid (PGGLCA; Mr 66 kDa) and the other rich in iduronic acid (PGIDOA; Mr 48 kDa). In PGGLCA, 84% of the glycosaminoglycan chains are composed of GlcA----GalNAc(SO4) units, whereas in PGIDOA, the chains contain 95% IdoA----GalNAc(SO4) disaccharide units. Upon treatment with testicular hyaluronidase, the PGs gave different-sized oligosaccharides. Chondroitinase ABC digestion of PGGLCA or PGIDOA gave a single protein core (Mr approximately 20 kDa). The presence of glucosamine and sialic acid in PGGLCA and PGIDOA suggests that both contain N-linked oligosaccharides.
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PMID:Proteoglycans in human burn hypertrophic scar from a patient with Ehlers-Danlos syndrome. 159 19

Monoclonal antibodies were raised against human glial hyaluronate-binding protein (GHAP), a major CNS-specific glycoprotein known to bind hyaluronate in vitro. Frozen sections of dog and human spinal cord were digested with Streptomyces hyaluronidase in order to ascertain whether GHAP is bound to hyaluronate in vivo. Digestion with hyaluronidase, prior to staining of the sections by conventional indirect immunofluorescence, led to a drastic reduction in the intensity of the staining reaction. Chondroitinase ABC (protease-free) was also effective in bringing about the release of GHAP from tissue sections. This enzyme also degrades hyaluronate. The effects of the chondroitinase were completely reversed by the addition of 1 mM Zn2+, a known inhibitor of this enzyme. The intact protein was released into the soluble fraction of human brain homogenates by testicular hyaluronidase. An immunoreactive species of 70 kD was released into the soluble fraction of dog spinal cord homogenates by Streptomyces hyaluronidase. Dog GHAP was isolated from spinal cord by means of ion exchange and affinity chromatography. This protein bound efficiently to hyaluronate in vitro. Dog and human GHAP had identical isoelectric points and similar peptide maps but different molecular weights. Dog GHAP (70 kD) was larger than its human counterpart (60 kD). These findings imply that GHAP exists in association with hyaluronate in CNS white matter. Immunoelectron microscopy revealed that GHAP fills the space between myelin sheaths in dog spinal cord white matter. One is led to conclude therefore that an hyaluronate based extracellular matrix exists in CNS white matter.
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PMID:Extracellular matrix of central nervous system white matter: demonstration of an hyaluronate-protein complex. 171 74

In order to visualize by light microscopy the glycosaminoglycans (GAGs) in the rat tongue mucosa, the tissue was fixed with cuprolinic-blue (CB)-aldehyde and the staining enhanced by autometallographic (AM) procedure. As other polyanions were also detected, enzymatic digestions with hyaluronidase, chondroitinase ABC and pronase were performed on these tissues in order to test the specificity of the staining. Chondroitinase ABC caused a dramatic decrease of silver grains in the lamina propria whereas hyaluronidase and pronase induced only discrete or no modification. This supported the concept that the GAGs visualized by CB and autometallography in this area as dermatan sulphate. The other polyanions (mostly DNA and RNA) seen in the epithelial layers were unaffected by these enzyme treatments.
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PMID:Autometallographic visualization of glycosaminoglycans in the tongue mucosa of rats using cuprolinic blue and enzymatic digestions. 186 53

The distribution of sulfated proteoglycans in Bruch's membrane of the human eye was evaluated histochemically using Cupromeronic Blue in combination with specific enzyme digestions and nitrous acid treatment. Five distinct categories of filament-shaped profiles were present following staining with this dye. Type 1 (90 +/- 13 nm long and 7 +/- 1 nm in diameter) (mean +/- S.D.) and type 2 (43 +/- 7 nm long and 5 +/- 1 nm in diameter) filaments were associated with collagen fibrils in the inner and outer collagenous zones. Type 3 profiles (70 +/- 18 nm long and 8 +/- 1 nm in diameter) were present in two locations--along the cortical border of the central elastic zone and within the basal infoldings of the pigment epithelium. Type 4 (60 +/- 11 nm long and 6 +/- 1 nm in diameter) and type 5 (200 +/- 100 nm long and 100 +/- 50 nm in diameter) filaments were associated with the basal laminae of the retinal pigment epithelium and choriocapillaris. Chondroitinase AC treatment eliminated the staining of type 1 filaments. Chondroitinase ABC treatment eliminated the staining of both type 1 and type 2 filaments. Nitrous acid eliminated the staining of type 4 and type 5 filaments. Incubations with keratanase or hyaluronidase did not alter the staining of any filament type. Type 3 filaments were resistant to all enzyme digestions and nitrous acid treatment. These results are consistent with an interpretation that Bruch's membrane contains chondroitin sulfate, dermatan sulfate and heparan sulfate-type proteoglycans. Proteoglycans containing chondroitin sulfate (type 1) and dermatan sulfate (type 2) are associated uniquely with collagen fibrils. Heparan sulfate type proteoglycans (types 4 and 5) are associated with the basal lamina of the pigment epithelium and choriocapillaris. The identity of type 3 profiles, which were resistant to all enzyme and nitrous acid digestions employed, could not be established at this time.
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PMID:Sulfated proteoglycans in Bruch's membrane of the human eye: localization and characterization using cupromeronic blue. 212 81

Enzymes, which degrade elements of the extracellular environment, were studied for their actions upon stereocilia and their cross-linkages by scanning electron microscopy. Chondroitinase, hyaluronidase and keratanase, which attack carbohydrate moieties of the extracellular matrix, had little effect upon hair bundles. Collagenase and plasmin (fibrinolysin) also had only marginal effects. Elastase produced dramatic effects upon hair bundles. Both lateral and tip links were degraded resulting in separation and splaying of stereocilia. Many stereocilia showed no marked loss of rigidity, although some were bent or kinked. In general, inner hair cells were the most susceptible to elastase followed by row 3, row 2, row 1 of the outer hair cells. The proteolytic enzyme trypsin did not noticeably disrupt the hair bundles. Protease caused loss of rigidity and fracture of stereocilia resulting in considerable collapse of hair bundles. Crosslinkages between stereocilia were less noticeably degraded. These results indicate that both lateral and tip links of stereocilia comprise a proteinaceous moiety which could be elastin or some chemically related structure.
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PMID:Action of elastase, collagenase and other enzymes upon linkages between stereocilia in the guinea-pig cochlea. 216 89

An improved micro method for measuring sulfated glycosaminoglycans (S-GAG) in chondrocyte cultures using 1,9-Dimethylmethylene Blue (DMB) has been developed. By increasing the protein concentration in the DMB assay a soluble GAG-DMB complex is prolonged. Without bovine serum albumin (BSA) in the phosphate-buffered saline (PBS) medium, the half time for loss of absorbance was 18 min; with 1% BSA-PBS there was no loss of absorbance over this time period. The limit of detection in a 96 well microtiter plate assay was 2 micrograms/ml; for a cuvette assay it was 1 microgram/ml. Collagen, DNA and RNA did not interfere with this assay. Hyaluronate caused an increase in absorbance at 530 nm that was lost by preincubating with Streptomyces hyaluronidase. The increase in absorbance was due to a turbidity change because there was no color shift from 600 to 530 nm but rather a uniform increase in absorbance between 400 to 700 nm. To validate the assay, the S-GAG was measured in conditioned medium from primary bovine articular chondrocyte monolayer cultures. A protein synthesis inhibitor, cycloheximide, blocked proteoglycan synthesis by greater than 90%. A cytokine, Interleukin-1 alpha, caused a dose-dependent decrease in proteoglycan accumulation. Chondroitinase ABC digestion of the chondrocyte conditioned medium completely prevented reactivity with the DMB. By preincubating samples with specific enzymes, different types of S-GAG can be measured with this assay. This assay can be used to measure changes in proteoglycans synthesized by chondrocytes.
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PMID:An improved method for determining proteoglycans synthesized by chondrocytes in culture. 237 28

Sulfated glycoconjugates were ultrastructurally localized within embryonic chick marrow by using the high iron diamine-silver proteinate stain. Stain was concentrated in the extravascular, granulopoietic compartment, indicating that granulopoiesis, but not erythropoiesis, proceeded in a highly sulfated environment. It was likely that most of the stainable material represented sulfated proteoglycans since staining was abrogated by predigesting tissue with enzymes and other treatments known to degrade specific glycosaminoglycan chains. Chondroitinase/hyaluronidase digestion resulted in the removal of most of the stainable material associated with the extracellular matrix and a portion of the stainable material associated with fibroblastic cell surfaces. Unaffected material lay in close proximity to fibroblastic cell membranes. Heparitinase/heparinase digestion had essentially the opposite effect. Sulfated material associated with matrix components was largely unaffected, but the fibroblastic plasmalemmal material was now absent. These results suggest that there are at least two categories of sulfated proteoglycans in the granulopoietic compartment, each differentially distributed. The plasmalemmal material likely represented heparan sulfate which in this tissue appeared to be associated in a uniform layer with fibroblastic stromal cell membranes and not with blood or endothelial cell membranes. Material identified as chondroitin sulfates was found within patches of amorphous matrix that was located on fibroblastic stromal cell surfaces and that was interspersed with fibrils in the extracellular matrix. Chondroitin sulfates were sparsely distributed on granulocytic cell surfaces.
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PMID:Ultrastructural localization of heparan sulfate and chondroitin sulfates associated with granulopoiesis in embryonic chick bone marrow. 244 89

There is little information available concerning the effects of orthodontic forces on glycosaminoglycans (GAG) of alveolar bone. The present study identifies changes in Alcian blue staining intensity in rat alveolar bone undergoing resorption resulting from a heavy (25g) tipping force applied to the adjacent teeth by a separating spring. One day after force application, bone from treated animals (internal control and experimental sides) demonstrated more intense staining with Alcian blue, pH 1.0 (p less than 0.005) and pH 2.5 (p less than 0.05) than external controls (untreated animals). By day 3, the intensity of Alcian blue staining of treated alveolar bone was similar to untreated. Chondroitinase AC, ABC and testicular hyaluronidase predigestion did not completely block the staining reaction, suggesting that both GAG and noncollagenous proteins were demonstrated. Mean cross-sectional areas of the interdental septum of the experimental side were nearly 44% less than that of the internal control side after 3 days and nearly 62% less after 5 days. The study suggested that alterations in bone GAG levels occurred prior to tooth movement as histochemical changes occurred after force application but before initiation of significant septal resorption. A precise appraisal of the types of macromolecules effected awaits future biochemical analysis. The results of the present work strongly suggest the use of an external control group for future studies, as Alcian blue staining reactions of the internal control side of treated animals were not similar to those of external controls.
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PMID:Effects of orthodontic tooth movement on the Alcian blue staining patterns of rat alveolar bone: an histochemical study. 298 Feb 6


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