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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)

A search was undertaken for bacteria which degrade chondroitin sulfate in nature and to find bacteria with a usefully high rate of chondroitinase (ChSase) productivity. First, 253 ChSase-producing bacteria were obtained from aquatic and land environments in Japan by aerobic and anaerobic screening methods. Identification according to Bergey's Manual of Determinative Bacteriology or Bain and Shewan (1968) permitted assignment of the majority of the isolates to seven genera, Aeromonas, Vibrio, Flavobacterium, Beneckea, Proteus, Micrococcus, and Arthrobacter. Next, ChSase productivities of all the isolates were compared with those of two established ChSase-producing stock strains, Proteus vulgaris NCTC 4636 and Flavobacterium heparinum ATCC 13125. As a result, special attention was given to production by a strain of Aeromonas sp. of large quantities of extracellular ChSase-AC. None of the isolates from the current study displayed significant ChSase-ABC productivity. Finally, ChSase-AC was prepared from the culture fluid of the Aeromonas strain by fractional precipitation with ammonium sulfate, chromatography on phospho-cellulose and diethylaminoethyl-cellulose, and gel filtration on Sephadex G-200. It was concluded that the Aeromonas strain may represent a profitable source of the enzyme ChSase-AC.
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PMID:Chondroitinase-producing bacteria in natural habitats. 80 22

Pentasaccharide 6-sulfate and hexasaccharide 6-sulfate were prepared from chondroitin 6-sulfate. Each oligosaccharide was incubated with a chick cartilage microsomal enzyme preparation and UDP [14C] glucuronic acid and/or UDP-N-[3H] acetylgalactosamine. As previously reported by other investigators, a single sugar was added from UDP-[14C] glucuronic acid to the nonreducing end of pentasaccharide 6-sulfate and from UDP-N-[3H] acetylgalactosamine to the nonreducing end of hexasaccharide 6-sulfate. The labeled oligosaccharides were characterized by gel chromatography and degradation by chondroitinase ABC followed by identification of products. The oligosaccharides in concentrations above their Km inhibited chondroitin synthesis on endogenous primers, reinforcing the assumption that the enzymes involved in the additions to exogenous oligosaccharides are the same as those involved in chondroitin polymerization. When either the pentasaccharide 6-sulfate or hexasaccharide 6-sulfate was incubated in reaction mixtures containing both of the sugar nucleotides there was generally growth of oligosaccharide by two or three sugars. With longer incubation under conditions of limiting oligosaccharide concentration, as many as 14 to 16 sugars could be added but no further chondroitin polymerization took place. Addition of each sugar was shown to depend upon the concentration of appropriate acceptor but was otherwise independent of the addition of the alternate sugar. No paired addition of sugars was noted. It was concluded that two specific enzymes are involved in alternate additions of sugars to the oligosaccharides and that the two enzymes have no apparent interaction with one another. It is suggested that the rapid polymerization to form large chondroitin chains which previously has been shown to take place on endogenous primers is facilitated by interaction of the two enzymes with a component of the endogenous primer. This component is not present in the exogenous oligosaccharides since they do not serve in the same fashion as primers for polymerization.
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PMID:Biosynthesis of chondroitin sulfate. Independent addition of glucuronic acid and N-acetylgalactosamine to oligosaccharides. 81 35

Nuclei isolated from rat liver were purified extensively and then subjected to extraction of glycosaminoglycans by the conventional method with a slight modification including the treatments with amylase, nucleases (DNAase and RNAase), and sialidase in addition to the pronase treatment. The nuclear glycosaminoglycan fraction thus prepared was subjected to chromatography on Dowes 1-X2 (Cl-) and electrophoresis before or after digestion with specific enzymes such as Streptomyces hyaluronidase, chondroitinase ABC and AC. These results together with the results of chemical analyses have revealed that the purified nuclei from rat liver contain glycosaminoglycans equivalent to 0.2-0.3 microgram hexuronic acid per mg DNA. A major component of the nuclear glycosaminoglycans has been identified as hyaluronic acid, while a minor component as chondroitin sulfate A (OR C). Preliminary investigations have shown that most of the nuclear glycosaminoglycans are associated with the chromatin fraction.
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PMID:Isolation and identification of glycosaminoglycans associated with purified nuclei from rat liver. 90 87

Dermatan sulfate-chondroitin sulfate copolymers have been isolated from human umbilical cord as a major galactosaminoglycan component of this tissue. The galactosaminoglycan fraction was obtained from this tissue by papain [EC 3.4.22.2] digestion followed by precipitation with cetylpyridinium chloride in a yield of 700 mg per 100 g of dry tissue. Ethanol fractionation resolved 4-5 subfractions differing in relative content of L-iduronic acid and D-glucuronic acid. No galactosaminoglycan containing either solely L-iduronic acid or D-glucuronic acid was obtained. The copolymeric structure of the material in each subfraction was demonstrated by analysis of oligosaccharide fragments obtained by chondroitinase-AC [EC 4.2.2.5] digestion. All the polymers contained repeating disaccharide units, D-glucuronosyl-N-acetylgalactosamine, D-glucuronosyl-N-acetylgalactosamine 4-sulfate, D-glucuronosyl-N-acetyl-galactosamine 6-sulfate, and L-iduronosyl-N-acetylgalactosamine 4-sulfate, of which D-glucuronosyl-N-acetylgalactosamine 6-sulfate and L-iduronosyl-N-acetylgalactosamine 4-sulfate were predominant. Both iduronic acid- and glucuronic acid-containing units were arranged in clusters. The presence of a considerable amount of nonsulfated disaccharide units was noted. The copolymers show extensive polydispersity in electrophoresis on cellulose acetate and gel chromatography on Sephadex G-200.
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PMID:Dermatan sulfate-chondroitin sulfate copolymers from ambilical cord. Isolation and characterization. 97 51

Further investigation was carried out on the action patterns of two chondroitinase-AC [EC 4.2.2.5.] preparations obtained from Arthrobacter aurescens and Flavobacterium heparinum. To infer the action patterns of the chondroitinases, we proposed a new method for the calculation of the degree of multiple attack, based on the concept established by Robyt and French ((1967) Arch. Biochem. Biophys. 122, 8-16). It was shown that the degree of multiple attack (DM) is represented by the ratio of the initial velocity of number-average degree of scission to that of viscosity-average degree of scission. By this method, DM for A-Chase was estimated to be 3.03 and for F-chase, 1.31.
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PMID:Action of chondroitinases. II. Numerical calculation of the degree of multiple attack. 101 15

Radioactivity was significantly incorporated from ascorbate 2-[35S]sulfate into chondroitin sulfate by embryonic chick cartilage epiphyses. The extent of incorporation was comparable with that from inorganic [35S]sulfate. The radioactive chondroitin sulfate formed from ascorbate 2-[35S]sulfate gave two radioactive disaccharides on chondroitinase-ABC [EC 4.2.2.4] digestion. The incorporation was markedly decreased by inorganic sulfate. The time course of incorporation from ascorbate 2-[35S]sulfate and inorganic [35S]sulfate into chondroitin sulfate and the constituent disaccharides suggest that the incorporation rates from the two radioactive substances are different.
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PMID:Sulfate incorporation from ascorbate 2-sulfate into chondroitin sulfate by embryonic chick cartilage epiphyses. 103 15

The bone inducing factor derived from BF osteosarcoma was purified in the following manner. Step 1. The sarcoma, grown in CBA mice, was excised and lyophilized. Step 2. The powder was washed with chilled acetone. Step 3. The acetone-treated powder was then homogenized with chilled distilled water. Step 4. Washing with 0.15M KCl. Step 5. The precipitate was incubated in in 0.2 N NH2OH, pH7.0, for 48 H at 25 degrees. After Step 5, the bone-forming activity showed a slight increase; however, the factor remained insoluble. The properties of the factor were as follows. The factor is relatively relatively heat stable; the osteogenic activity survived the treatment at 75 degrees for 15 min or at 55 degrees for 19 h. The activity was easily lost by mechanical shaking. Incubation with DNase, RNase, neuraminidase, chondroitinase ABC and beta-galactosidase left the osteogenic activity intact, but treatment with either pronase or collagnease destroyed this activity. The results suggest that the factor may be a protein. The activity was seen with the lyophilized BF osteosarcoma cells (without matrix), and it is probable that the factor was exclusively synthesized in the cells. The bone formation, observed across a millipore filter when living BF osteosarcoma enclosed in a millipore chamber was implanted in mice, suggests the synthesis and secretion of the factor from the cells.
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PMID:Studies on a factor responsible for new bone formation from osteosarcoma in mice. 105 58

Km and Vmax. were determined for the degradation by chondroitinase of chondroitin 4-sulphate, 4-sulphate-proteoglycna, chondroitin 6-sulphate, dermatan sulphate and hyaluronic acid. Degradation of chondroitin 4-sulphate was inhibited by hyaluronic acid but not by keratan sulphate. The results are discussed with regard to the use to the use of chondroitinase as a sleective reagent for the degradation of tissue glycosaminoglycans.
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PMID:The kinetic of degradation of chondroitin of sulphates and hyaluronic acid by chondroitinase form Proteus vulgaris. 109 55

Culture of chondrocytes in the presence of 4-methylumbelliferyl beta-D-xyloside resulted in a synthesis of protein-free, fluorogenic chondroitin sulfate which was heterogeneous on DEAE-cellulose chromatography. Degradation of the major chromatographic fraction with chondroitinase-ABC yielded, in addition to a large quantity of delta4-glucuronic acid-containing disaccharides, two fluorogenic oligosaccharides of different size. Quantitative analysis showed that delta4-glucuronic acid, galactose, xylose, and 4-methylumbelliferone were present in the small oligosaccharide fragment in a molar ratio of 1:2:1:1. Since these analytical data are analogous to those reported for glycopeptides derivedfrom proteochondroitin sulfates, it may be suggested that 4-methyl-umbelliferyl beta-D-xyloside replaces the need for xylosyl protein core in the normal synthesis of proteochondroitin sulfate with a resultant production of the unusual polysaccharide bearing the added xyloside at the reducing end.
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PMID:Synthesis of a fluorogenic mucopolysaccharide by chondrocytes in cell culture with 4-methylumbelliferyl beta-D-xyloside. 111 97

L-[14C]Iduronic acid-containing sulfated galactosaminoglycans were formed by incubation of a fibroblast particulate fraction with UDP-D[14C]glucuronic acid, UDP-N-acetylgalactosamine, and sulfate donor (3'-phosphoadenylylsulfate). The formation of L-iduronic acid was strongly promoted by concomitant sulfation of the polymer. In the absence of sulfate donor 5 to 10% of the [14C]uronic acid residues were L-iduronic acid. However, when 3'-phosphoadenylylsulfate was included in the incubation mixture the amount of L-iduronic acid in the product increased 3 to 5-fold. Furthermore, approximately the same quantity of L-[14C]iduronic acid was recovered from the product formed in a pulse-chase experiment where incorporation of 14C-isotope preceded sulfation. It was therefore concluded that C-5 inversion of D-glucuronic acid to L-iduronic acid occurred on the polymer level as shown previously for the biosynthesis of heparin (Hook, M., Lindahl, U., Backstrom, G., Malmstrom, A., AND Fransson, L-A., J. Biol. Chem. (1974) 249, 3908). This conclusion was supported by the finding that no L[14C]iduronic acid could be detected in the UDP-hexuronic acid pool during this experiment. Nonsulfated and sulfated [14C]galactosaminoglycan products were degraded separately with chondroitinase-AC. The non-sulfated products afforded primarily disaccharide and a small amount of tetrasaccharide, while the sulfated products yielded, in addition, a considerable amount of larger oligosaccharides. Tetrasaccharides from nonsulfated products contained L-iduronic acid indicating that C-5 inversion at solitary sites can occur in the absence of sulfation of adjacent hexosamine moieties. The larger oligosaccharides obtained after chondroitinase-AC digestion of sulfated products yielded L-iduronic acid upon acid hydrolysis and were susceptible to chondroitinase-ABC digestion. The split products were almost exclusively 4-sulfated disaccharides. These results demonstrate that formation of blocks of L-iduronic acid-containing repeat periods is associated with 4-sulfation of adjacent hexosamine moieties.
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PMID:Biosynthesis of dermatan sulfate. I. Formation of L-iduronic acid residues. 112 48


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