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

---

Query: EC:3.1.6.12 (chondroitinase)
2,183 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous studies on rabbit thrombomodulin (TM) revealed that certain anticoagulant activities expressed by TM depend on the presence of an acidic domain tentatively identified as a sulfated galactosaminoglycan (Bourin, M.-C., Ohlin, A.-K., Lane, D., Stenflo, J., and Lindahl, U. (1988) J. Biol. Chem. 263, 8044-8052). The glycan was released by alkaline beta-elimination, isolated by ion-exchange chromatography, and radiolabeled by partial N-deacetylation (hydrazinolysis) followed by re-N-[3H]acetylation. The labeled product behaved like standard chondroitin sulfate on ion-exchange chromatography, exhibited a Mr of 10-12 x 10(3) on gel chromatography, and was susceptible to degradation by chondroitinase and testicular hyaluronidase. The major labeled degradation products following digestion of the glycosaminoglycan with chondroitinase were identified, depending on the incubation conditions, either as 4/6-mono-O-sulfated, 4,5-unsaturated disaccharides (delta HexA-GalNAc(S] and N-acetylgalactosamine 4,6-di-O-sulfate (GalcNAc (diS], the latter component accounting for approximately 25% of the total label, or as a major fraction of labeled trisaccharide, with the predominant structure GalNAc(diS)-GlcA-GalNAc(diS). The terminal GalNAc(diS) unit (not substituted at C3) was shown to be more susceptible to N-deacetylation during hydrazinolysis than were the internal GalNAc units (substituted at C3), and thus was more extensively labeled, resulting in over-representation of this unit. It is concluded that rabbit TM is a chondroitin sulfate proteoglycan, which carries a single glycan side chain characterized by an unusual accumulation of sulfate groups at the nonreducing terminus. Metabolically 35S-labeled TM was isolated from cultured rabbit heart endothelial cells and characterized as a chondroitin sulfate proteoglycan which accounted for 1-2% of the total 35S-labeled cell-associated macromolecules. The isolated chondroitin sulfate showed weaker antithrombin-dependent anticoagulant activity, on a molar basis, than the intact TM proteoglycan. The anticoagulant action of TM thus depends on a unique form of functional collaboration between the different constituents of a glycoconjugate.
...
PMID:Isolation and characterization of the glycosaminoglycan component of rabbit thrombomodulin proteoglycan. 216 13

Thrombomodulin (TM), a major anticoagulant protein at the vessel wall, serves as a potent cofactor for the activation of Protein C by thrombin. Previous work has indicated that (rabbit) TM is a proteoglycan that contains a single polysaccharide chain, tentatively identified as a sulphated galactosaminoglycan, and furthermore suggested that this component may be functionally related to additional anticoagulant activities expressed by the TM molecule [Bourin, Ohlin, Lane, Stenflo & Lindahl (1988) J. Biol. Chem. 263, 8044-8052]. Results of the present study establish that (enzymic) removal of the polysaccharide chain abolishes the inhibitory effect of TM on thrombin-induced fibrinogen clotting as well as the promoting effect of TM on the inactivation of thrombin by antithrombin, but does not affect the ability of TM to serve as a cofactor in the activation of Protein C. Studies of yet another biological activity of rabbit TM, namely the ability to prevent the activation of Factor V by thrombin [Esmon, Esmon & Harris (1982) J. Biol. Chem. 257, 7944-7947], confirmed that TM markedly delays the conversion of the native 330 kDa Factor V precursor into polypeptide intermediates, and further into the 96 kDa heavy chain and 71-74 kDa light-chain components of activated Factor Va. In contrast, the activation kinetics of a similar sample of Factor V incubated with thrombin in the presence of chondroitinase ABC-digested TM did not differ from that observed in the absence of TM. It is concluded that the inhibitory effect of TM on Factor V activation also depends on the presence of the polysaccharide component on the TM molecule.
...
PMID:Functional role of the polysaccharide component of rabbit thrombomodulin proteoglycan. Effects on inactivation of thrombin by antithrombin, cleavage of fibrinogen by thrombin and thrombin-catalysed activation of factor V. 216 42

Thrombomodulin acts as a cofactor for protein C activation by thrombin (PC activation cofactor activity) and inhibits thrombin-induced fibrinogen clotting (direct anticoagulant activity). In addition, rabbit thrombomodulin has been shown to promote thrombin inactivation by antithrombin (AT-dependent anticoagulant activity). However, a non-acidic form (i.e. non-retarded on ion-exchange chromatography) of thrombomodulin generated by limited proteolysis retained only the PC activation cofactor activity. The acidic form (retarded on ion-exchange chromatography) of thrombomodulin is now shown to prevent the rapid inactivation of thrombin by antithrombin in the presence of heparin, presumably by preventing the formation of the ternary thrombin-AT-heparin complex. This effect was not observed with non-acidic thrombomodulin. When submitted to chondroitinase digestion, thrombomodulin was converted into an essentially non-acidic form that lacked both the AT-dependent and the direct anticoagulant activities but showed a PC activation cofactor function indistinguishable from that of native thrombomodulin. This chondroitinase-digested form did not prevent the catalytic effect of heparin on the inhibition of thrombin by AT. It is concluded that the acidic domain of rabbit thrombomodulin, a chondroitin (dermatan) sulfate glycosaminoglycan, interacts with a site of the thrombin molecule that is not involved in the protein C activation cofactor function, but is essential to the cleavage of fibrinogen or binding of heparin.
...
PMID:Effect of rabbit thrombomodulin on thrombin inhibition by antithrombin in the presence of heparin. 254 98

35S-labelled heparins were recovered from adipose tissue, hearts, lungs, peritoneal cavities and skins of rats given H2(35)SO4. Their purification involved incubation with Pronase, precipitation with cetylpyridinium chloride in 1.0 M-NaCl, gradient elution from DEAE-Sephacel and incubation with chondroitinase ABC. Each product was divided into proteoglycan and "depolymerization products' fractions by gel filtration on Bio-Gel A-15m. Heparin chains were released from a portion of each proteoglycan fraction by beta-elimination with NaOH. Proteoglycans, chains and depolymerization products were separated by gradient elution from a column of antithrombin-agarose into fractions with no affinity, low affinity and high affinity for antithrombin. The relative sizes of the products were determined by gel filtration on columns of Bio-Gel A-50m, A-15m, A-1.5m and A-0.5m. Skin was the major source of heparin and contained the largest proteoglycans and the lowest proportion of depolymerization products. Lungs contained the smallest proteoglycans, the smallest depolymerization products and the highest proportion of depolymerization products. The highest proportions of proteoglycans, chains and depolymerization products with high affinity for antithrombin were found in adipose tissue. The lowest proportions of each of these fractions were found in the peritoneal cavity. The data suggest that there was relatively little biosynthesis of sites with high affinity for antithrombin in peritoneal-cavity mast cells and that heparin catabolism was most active in lungs. Each source of heparin was unique with respect to both biosynthesis and subsequent breakdown of its proteoglycans.
...
PMID:Rat heparins. A study of the relative sizes and antithrombin-binding characteristics of heparin proteoglycans, chains and depolymerization products from rat adipose tissue, heart, lungs, peritoneal cavity and skin. 382 37

Polysaccharide was isolated from human spleen mastocytoma by proteolytic digestion, precipitation with cetylpyridinium chloride, digestion with chondroitinase ABC, and ion-exchange chromatography on DEAE-cellulose. The final product (0.7 mg per g of starting material, MW 8000) behaved like standard heparin on ion-exchange chromatography and on electrophoresis, and contained D-glucuronic acid, L-iduronic acid, D-glucosamine and sulfate in the proportions expected for heparin. Affinity chromatography on antithrombin-Sepharose separated a distinct high-affinity fraction (4-5% of the total material). Structural analysis of this fraction showed that about 10% of the D-glucosamine residues were N-acetylated, the remainder N-sulfated. The anticoagulant activity of the isolated heparin was 71 B.P. units per mg (whole-blood system), or 30 units per mg (anti-thrombin and chromogenic substrate). 205 and 10-15 units per mg (chromogenic assay) were found for high and low affinity fractions, respectively. These results demonstrate conclusively the occurrence of heparin in a human tissue.
...
PMID:Isolation and characterization of heparin from human mastocytoma tissue. 678 Oct 95

It has been found that dermatan polysulfates (DPS) I, II and III isolated from hagfish notochord, hagfish skin and shark skin, respectively, and chemically sulfated dermatan sulfate exhibit considerable anticoagulant activity in the "activated partial thromboplastin time (APTT)" system. On comparing the activities with the various compositions, including disaccharide units produced by the digestion with chondroitinase-ABC, it was shown that the activity of these dermatan polysulfates depends not only on the total sulfate content but also on the content of sulfated L-iduronic acid residues. The activity seemed to decrease for molecular weight of below 10,000. The effect of these dermatan polysulfates on th inactivation of the clotting enzymes, factor Xa and thrombin, by antithrombin II (AT-III) was also studied using chromogenic substrates for the assay of the enzyme activities. The dermatan polysulfates showed an inhibitory effect on thrombin-AT-III, as estimated by the APTT assay, in contrast with the effect on factor Xa-AT-III which was found to be very small.
...
PMID:Anticoagulant activity of dermatan polysulfates. 680

Heparin, NAcHep, DS, and CS were labeled with deuterium by N-reacetylating, with the deuterated acetic anhydride (CD3CO)2O, GAGs previously N-deacetylated (by hydrazinolysis) to the desired extent. Degrees of deuteration of the present preparations, as determined by 2H- and 1H-NMR were 15%, 51%, 49%, and 79% for heparin, NAcHep, DS, and CS, respectively. The NMR analysis (including the 13C spectra) of the labeled products indicated that deuterium labeling did not involve any substantial modification of the GAG structures. Also NMR signals associated with specific sequences of heparin for antithrombin and of DS for heparin cofactor II were essentially the same in the unlabeled and in the deuterated GAGs. The substantial retention of the original structure was confirmed by data on the degree of sulfation (by conductimetry) and on the electrophoretic mobility in acid buffer. On the other hand, HPLC/SEC data indicated some depolymerization of heparin and DS in the N-deacetylation step of the labeling reactions. HPLC/MS spectrometry permitted a clear identification of disaccharide and tetrasaccharide fragments obtained from deuterated GAGs by enzymic (heparinase, chondroitinase ABC) or chemical depolymerization (deaminative cleavage, Smith degradation), opening new prospects for studies of human pharmacokinetics, with differentiation of exogenous from endogenous GAGs.
...
PMID:Preparation and characterization of deuterium-labeled glycosaminoglycans. 799 88

While checking anticoagulant activities in crude fractions from Wakan-Yakus (traditional herbal drugs), we detected antithrombin activity in the polysaccharide fraction of the leaves of Artemisia princeps Pamp. A sulfated polysaccharide purified from the crude fractions by ion-exchange chromatography on DEAE-cellulose and gel filtration on Sepharose 6B potentiated the heparin cofactor II (HC II)-dependent antithrombin activity but not the antithrombin activity of antithrombin III (AT III). The polysaccharide enhanced the HC II-thrombin reaction more than 6000-fold. The apparent second-order rate constant of thrombin inhibition by HC II increased from 3.8 x 10(4) (in the absence of the polysaccharide) to 2.5 x 10(8) M-1 min-1 in the presence of 25-125 micrograms/ml of the polysaccharide. In human plasma, the polysaccharide accelerated the formation of thrombin-HC II complex. The stimulating effect on HC II-dependent antithrombin activity was almost totally abolished by treatment with chondroitinase AC I, heparinase or heparitinase, while chondroitinase ABC or chondroitinase AC II had little or no effect. These results suggest that the polysaccharide is a glycosaminoglycan-like material with properties that are quite distinct from heparin or dermatan sulfate.
...
PMID:Selective activation of heparin cofactor II by a sulfated polysaccharide isolated from the leaves of Artemisia princeps. 856 35

We examined the ability of unfractionated heparin to modulate the procoagulant activities of stimulated endothelial cells (EC). Confluent human venous umbilical EC were incubated with heparin before or after stimulation, then rinsed extensively to eliminate any heparin in the solution. EC, stimulated for 4 h with endotoxin and interleukin 1 beta, expressed tissue factor and prothrombinase activities. When EC were treated with heparin (6 and 60 micrograms/ml) during the last 10 min of the stimulation period, EC-related procoagulant activities were inhibited in a dose-dependent manner (80-90% inhibition at 60 micrograms/ml). The inhibition was antithrombin-dependent and it disappeared after heparin removal in less than 15 min at 37 degrees C but persisted at 4 degrees C. When EC were treated with heparin (60 micrograms/ml) for 24 h then extensively washed before stimulation, the anticoagulant effect was more modest (50% inhibition). The effect was antithrombin-dependent. Inhibition was maximum after 18-24 h of pretreatment of EC with heparin and was stable for at least 7 h. The cell surface displayed a "heparin-like" activity: treatment by heparin doubled the rate of thrombin-antithrombin complex formation and this effect was heparinase sensitive and chondroitinase ABC insensitive. Thus, heparin modulates the procoagulant properties of stimulated EC according to two distinct mechanisms. The first one is rapid and transient, probably related to the presence of heparin molecules bound at the membrane surface. The second is delayed and persistent, and our results suggest that it is mediated by an increase in the membrane heparan sulfate molecules.
...
PMID:Heparin reverses the procoagulant properties of stimulated endothelial cells. 871

Calcium spirulan (Ca-SP), a novel sulfated polysaccharide isolated from the blue-green alga Spirulina platensis, enhanced the antithrombin activity of heparin cofactor II (HC II) more than 10000-fold. The apparent second-order rate constant of thrombin inhibition by HC II was calculated to be 4.2 x 10(4) M-1 min-1 in the absence of Ca-SP, and it increased in the presence of 50 micrograms/ml Ca-SP to 4.5 x 10(8) M-1 min-1. Ca-SP effectively induced the formation of a thrombin-HC II complex in plasma. In the presence of Ca-SP, both the recombinant HC II variants Lys173-->Leu and Arg 189-->His, which are defective in interactions with heparin and dermatan sulfate, respectively, inhibited thrombin in a manner similar to native rHC II. This result indicates that the binding site of HC II for Ca-SP is different from the heparin- or dermatan sulfate-binding site. When we removed the calcium from the Ca-SP, the compound did not exert any antithrombin activity. Furthermore, Na-SP, which was prepared by replacement of the calcium in Ca-SP with sodium, accelerated the antithrombin activity of HC II as Ca-SP did. We therefore suggest that the molecular conformation maintained by Ca or Na is indispensable to the antithrombin activity of Ca-SP. The HC II-dependent antithrombin activity of Ca-SP was almost totally abolished by treatment with chondroitinase AC I, heparinase or heparitinase, but not by treatment with chondroitinase ABC and chondroitinase AC II, suggesting that a heparin- or dermatan sulfate-like structure is not responsible for the activation of HC II by Ca-SP. Ca-SP is therefore thought to be a unique sulfated polysaccharide which shows a strong antithrombin effect in an exclusively HC II-dependent manner.
...
PMID:Heparin cofactor II-dependent antithrombin activity of calcium spirulan. 887 66


1 2 Next >>

---