scholarly journals Biosynthesis of chondroitin sulphate by a Golgi-apparatus-enriched preparation from cultures of mouse mastocytoma cells

1980 ◽  
Vol 190 (2) ◽  
pp. 307-313 ◽  
Author(s):  
J E Silbert ◽  
L S Freilich
Keyword(s):  
Golgi Apparatus ◽  
Suspension Culture ◽  
Glucuronic Acid ◽  
Chondroitin Sulphate ◽  
Fold Increase ◽  
Active Location ◽  
Polysaccharide Synthesis ◽  
Mastocytoma Cells

Mouse mastocytoma cells grown in suspension culture produce chondroitin 4-sulphate. A Golgi-apparatus-enriched fraction from these cells was prepared and examined for chondroitin-synthesizing activity. When Golgi-apparatus-enriched fractions were incubated with UDP-[14C]glucuronic acid and UDP-N-acetylgalactosamine, they demonstrated a greater than 13-fold increase in chondroitin-synthesizing activity over cell homogenates. Similar incubations with the addition of a pentasaccharide from chondroitin sulphate resulted in a greater than 40-fold increase in [14C]glucuronic acid-incorporating activity over cell homogenates. Other membrane fractions had much less activity, suggesting that the Golgi apparatus is the most active location for chondroitin biosynthesis. Products of the incubations indicated the formation of [14C]chondroitin glycosaminoglycan on endogenous primers and formation of [14C]-hexasaccharide and somewhat larger [14C]oligosaccharides on exogenous pentasaccharide acceptors. There was, however, a significant amount of large [14C]-chondroitin glycosaminoglycan formed on pentasaccharide, indicating that some pentasaccharide did serve as a true primer for polysaccharide synthesis.

scholarly journals Influence of Phytohormones on Monosaccharide Composition of Polysaccharides from Wheat Suspension Culture

2017 ◽  
Vol 19 (3) ◽  
pp. 231 ◽  
Author(s):  
N. Bishimbayeva ◽  
A. Murtazina ◽  
G. McDougall
Keyword(s):  
Chemical Composition ◽  
Suspension Culture ◽  
Culture Medium ◽  
Glucuronic Acid ◽  
Physiological Activity ◽  
Fold Increase ◽  
Monosaccharide Composition ◽  
Medium Composition ◽  
Plant Polysaccharides ◽  
Different Types

Plant polysaccharides with technical and physiologic traits attract researchers by their high physiological activity in regulation of the growth, development and protective reactions. Cell cultures allow to regulate chemical composition of synthesized substances by changing media composition and are widely used to enhance or change the biosynthesis of metabolites. The aim of this study was to investigate the influence of phytohormones 2,4-dichlorphenoxyacetic acid (2,4 –D) and abscisic acid (ABA) of culture medium on chemical composition of polysaccharides (PS), extracted from cells and extracellular liquid of wheat suspension culture. It was shown for the medium with ABA that monosaccharide composition of extracellular PS mainly represented by glucose (87%), whereas PS isolated from cells were rich for xylose and glucuronic acid. Monosaccharide composition of extracellular PS from media with 2,4-D showed 6-fold increase of arabinose, 8-fold ‒ of galactose, 5-fold ‒ of xylose and glucuronic acid, compared to extracellular PS from ABA medium. Composition of cellular PS from media with 2,4-D were mainly similar to ABA and differed by the increased amount of mannose (3-fold), and galacturonic acid (2,5-fold). Thus, regulative effect of the use of two different types of phytohormones was demonstrated on the biosynthesis of variously composed polysaccharides.

Histochemical Differentiation of the Basement Membrane of the Mouse Seminiferous Tubule

1962 ◽  
Vol s3-103 (63) ◽  
pp. 385-391
Author(s):  
A. H. BAILLIE
Keyword(s):  
Basement Membrane ◽  
Protein Metabolism ◽  
Seminiferous Tubule ◽  
Glucuronic Acid ◽  
Chondroitin Sulphate ◽  
Hair Follicles ◽  
Ground Substance ◽  
Seminiferous Tubules ◽  
Active Protein ◽  
Pas Reaction

The ground substance of the testis of the albino mouse is PAS-positive but not metachromatic, and probably highly aggregated. The basement of the seminiferous tubules is intensely PAS-positive, metachromatic, and possibly not so highly aggregated. The reactivity of the ground substance to the PAS reaction and toluidine blue is tentatively ascribed to the presence of chondroitin sulphate C: this compound, previously known to contain N acetyl-galactosamine, glucuronic acid, tyrosine and tryptophane, is associated with arginine. The genesis of the basement membrane of the seminiferous tubule is shown to include the formation of a sheath of atypical elongated fibroblasts, the secretion of a PAS positive, metachromatic substance associated with arginine between this sheath and the seminiferous tubule, the appearance of mitochondria in the cells of the sheath, and lastly, the acquisition of alkaline phosphatase by these fibroblasts and its spread to the intervening ground substance. These changes are thought to be related to the structural and nutritional requirements of the seminiferous tubules. In its intense positive reaction to PAS and in its metachromasy, the basement membrane of the seminiferous tubule agrees with the ground substance adjacent to sites of active protein metabolism, such as growing tumours, embryonic organs, hair follicles, and skin.

scholarly journals Sequential degradation of a chondroitin sulphate trisaccharide by lysosomal enzymes from embryonic-chick epiphysial cartilage

1979 ◽  
Vol 179 (1) ◽  
pp. 7-13 ◽  
Author(s):  
B Ingmar ◽  
A Wasteson
Keyword(s):  
Lysosomal Enzymes ◽  
Glucuronic Acid ◽  
Enzyme Inhibitors ◽  
Functional Unit ◽  
Chondroitin Sulphate ◽  
Cartilage Degradation ◽  
Ester Group ◽  
Embryonic Chick ◽  
Reaction Products ◽  
Epiphysial Cartilage

The disulphated trisaccharide D-N-acetylgalactosamine sulphate-beta-D-glucuronic acid-beta-D-N-acetylgalactosamine sulphate prepared from 35S- or 14C-labelled chondroitin sulphate was incubated with a preparation of lysosomal enzymes from embryonic-chick epiphysial cartilage. Degradation was demonstrated by analysis of the reaction products. By use of the appropriate intermediate products as substrates, in conjunction with specific enzyme inhibitors, it was shown that the degradation proceeded sequentially from the non-reducing end. It was initiated by sulphatase (preferentially hydrolysing sulphate ester groups at the 6-position), followed by beta-N-acetylgalactosaminidase and beta-glucuronidase, converting the substrate into monosaccharides and inorganic sulphate. The latter enzyme preferentially attacked disaccharides carrying their sulphate ester group at C-4 of the hexosamine residue. Generation of chondroitin sulphate oligosaccharides may occur by the action of an endoglycosidase, previously demonstrated in embryonic-chick cartilage. Endo- and exo-enzymes may thus form a functional unit in lysosomal degradation of chondroitin sulphate.

scholarly journals Subcellular distribution of selenium in deficient mouse liver

1989 ◽  
Vol 258 (2) ◽  
pp. 541-545 ◽  
Author(s):  
R Reiter ◽  
R Otter ◽  
A Wendel
Keyword(s):  
Endoplasmic Reticulum ◽  
Plasma Membrane ◽  
Golgi Apparatus ◽  
Trace Element ◽  
Fold Increase ◽  
Subcellular Fractionation ◽  
Fast Response ◽  
Cell Compartment ◽  
Deficient Mice

Selenium (Se)-deficient mice were labelled in vivo with single pulses of [75Se]selenite, and the intrahepatic distribution of the trace element was studied by subcellular fractionation. At 1 h after intraperitoneal injection of 3.3 or 10 micrograms of Se/kg body weight, 15% of the respective doses were found in the liver. Accumulation in the subcellular fractions followed the order: Golgi vesicular much greater than lysosomal greater than cytosolic = microsomal greater than mitochondrial, peroxisomal, nuclear and plasma-membrane fraction. At a dose of 3.3 micrograms/kg, more than 90% of the hepatic Se was protein-bound. When cross-contamination was accounted for, the following specific Se contents of the subcellular compartments were extrapolated: Golgi apparatus, 7.50 pmol/mg; cytosol, 0.90 pmol/mg; endoplasmic reticulum, 0.80 pmol/mg; mitochondria, 0.49 pmol/mg; nuclei, lysosomes, peroxisomes and plasma membrane, less than 0.4 pmol/mg. At 10 micrograms/kg, a roughly 2-3-fold increase in Se content of all fractions was found without major changes in the intrahepatic distribution pattern. An extraordinary rise in the cytosolic fraction was due to an apparently non-protein-bound Se pool. At 24 h after dosing, total hepatic Se had decreased to 6% of the initial dose and had become predominantly protein-bound. The 60% decrease in hepatic Se was reflected in a similar fall in the subcellular levels of the trace element. The Golgi apparatus still had the highest specific Se content, although accumulation was 5 times less than that after 1 h. The cytosolic pool accounted for 50% of the hepatic Se at both labelling times. After 1 h the Golgi apparatus was, with 19%, the second largest intrahepatic pool, followed by the endoplasmic reticulum with 16%. The high affinity and fast response of the Golgi apparatus to Se supplementation of deficient mice is interpreted in terms of a predominant function of this cell compartment in the processing and the export of Se-proteins from the liver.

scholarly journals The effect of steroids and nucleotides on solubilized bilirubin uridine diphosphate glucuronyltransferase

1970 ◽  
Vol 119 (3) ◽  
pp. 437-445 ◽  
Author(s):  
B. P. F. Adlard ◽  
G. H. Lathe
Keyword(s):  
Enzyme Activity ◽  
Glucuronic Acid ◽  
Liver Microsomes ◽  
Fold Increase ◽  
Rat Liver Microsomes ◽  
Uridine Diphosphate ◽  
Competitive Effects ◽  
Solubilized Enzyme ◽  
High Concentrations ◽  
Solubilized Form

1. It was confirmed that bilirubin glucuronyltransferase can be obtained in solubilized form from rat liver microsomes. 2. Michaelis–Menten kinetics were not followed by the enzyme with bilirubin as substrate when the bilirubin/albumin ratio was varied. High concentrations of bilirubin were inhibitory. 3. The Km for UDP-glucuronic acid at the optimum bilirubin concentration was 0.46mm. 4. Low concentrations of Ca2+ were inhibitory in the absence of Mg2+ but stimulatory in its presence; the converse applied for EDTA. 5. UDP-N-acetylglucosamine and UDP-glucose enhanced conjugation by untreated, but not by solubilized microsomes. 6. The apparent 9.5-fold increase in activity after solubilization was probably due to the absence of UDP-glucuronic acid pyrophosphatase activity in the solubilized preparation. 7. The activation of solubilized enzyme activity by ATP was considered to be a result of chelation of inhibitory metal ions. 8. The solubilized enzyme activity was inhibited by UMP and UDP. The effect of UMP was not competitive with respect to UDP-glucuronic acid. 9. A number of steroids inhibited the solubilized enzyme activity. The competitive effects of stilboestrol, oestrone sulphate and 3β-hydroxyandrost-5-en-17-one, with respect to UDP-glucuronic acid, may be explained on an allosteric basis.

scholarly journals Hydrazinolysis of heparin and other glycosaminoglycans

1984 ◽  
Vol 217 (1) ◽  
pp. 187-197 ◽  
Author(s):  
P N Shaklee ◽  
H E Conrad
Keyword(s):  
Chemical Modification ◽  
Carboxy Group ◽  
Uronic Acid ◽  
Glucuronic Acid ◽  
Chondroitin Sulphate ◽  
Acid Hydrazide ◽  
Iduronic Acid ◽  
Heparin Treatment ◽  
Hydrazine Sulphate ◽  
Low Rate

Heparin, carboxy-group-reduced heparin, several sulphated monosaccharides and disaccharides formed from heparin, and a tetrasaccharide prepared from chondroitin sulphate were treated at 100 degrees C with hydrazine containing 1% hydrazine sulphate for periods sufficient to cause complete N-deacetylation of the N-acetylhexosamine residues. Under these hydrazinolysis conditions both the N-sulphate and the O-sulphate substituents on these compounds were completely stable. However, the uronic acid residues were converted into their hydrazide derivatives at rates that depended on the uronic acid structures. Unsubstituted L-iduronic acid residues reacted much more slowly than did unsubstituted D-glucuronic acid or 2-O-sulphated L-iduronic acid residues. The chemical modification of the carboxy groups resulted in a low rate of C-5 epimerization of the uronic acid residues. The hydrazinolysis reaction also caused a partial depolymerization of heparin but not of carboxy-group-reduced heparin. Treatment of the hydrazinolysis products with HNO2 at either pH 4 or pH 1.5 or with HIO3 converted the uronic acid hydrazides back into uronic acid residues. The use of the hydrazinolysis reaction in studies of the structures of uronic acid-containing polymers and the implications of the uronic acid hydrazide formation are discussed.

scholarly journals Biosynthesis of glycosaminoglycans in bovine cornea. The effect of uridine diphosphate xylose

1970 ◽  
Vol 120 (4) ◽  
pp. 719-723 ◽  
Author(s):  
C. Balduini ◽  
A. Brovelli ◽  
A. A. Castellani
Keyword(s):  
Glucuronic Acid ◽  
Chondroitin Sulphate ◽  
Glucose Dehydrogenase ◽  
Uridine Diphosphate ◽  
Regulatory Effect ◽  
Keratan Sulphate ◽  
Polysaccharide Chain

1. The role of UDP-xylose in the regulation of corneal glycosaminoglycan biosynthesis was investigated. Bovine corneas were incubated with [U-14C]-glucose in the presence and in the absence of the nucleotide, and the radioactivity of chondroitin, chondroitin sulphate and keratan sulphate, as well as of their monosaccharide constituents, was determined. 2. A decrease in the rate of biosynthesis of chondroitin and chondroitin sulphate and an increase in that of keratan sulphate were observed in the samples incubated with UDP-xylose. 3. The UDP-glucuronic acid isolated after the incubation in the presence of UDP-xylose showed a noticeable decrease in the amount of radioactivity incorporated; this result suggests that UDP-xylose inhibits the UDP-glucose dehydrogenase, causing an accumulation of UDP-glucose and consequently an increase in the formation of UDP-galactose and keratan sulphate. 4. Galactose and galactosamine isolated from the polysaccharides showed variations in the amount of radioactivity incorporated in accordance with those observed for the macromolecules; this fact confirms that in the system we used in vitro a real biosynthesis of the polysaccharide chain took place and that the regulatory effect of UDP-xylose was active at the monosaccharide level.

scholarly journals Sulphated glycosaminoglycans in regenerating rat liver

1980 ◽  
Vol 188 (3) ◽  
pp. 769-773 ◽  
Author(s):  
M Edward ◽  
W F Long ◽  
H H Watson ◽  
F B Williamson
Keyword(s):  
Molecular Mechanisms ◽  
Temporal Trend ◽  
Chondroitin Sulphate ◽  
Heparan Sulphate ◽  
Fold Increase ◽  
Sham Operation ◽  
Dermatan Sulphate ◽  
Weight Percentage ◽  
Biphasic Pattern ◽  
Sulphated Glycosaminoglycans

The total weight percentage glycosaminoglycan content of rat liber was found to increase by 50% in the first 30 h after partial hepatectomy. The content returned to near normal by the third day, but then increased again to a second maximum at 5-6 days, only to gradually decline to normal by the ninth day, when regeneration was nearly complete. This biphasic pattern was most marked in the chondroitin sulphate A/C component, with a 6-fold increase by the sixth day. Dermatan sulphate showed the same temporal trend, whereas heparan sulphate remained relatively unaltered. No such changes were detected in the livers of rats subjected to sham operation. The possible molecular mechanisms underlying the apparent link between cellular glycosaminoglycan content and proliferative tendency are discussed.

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