scholarly journals Very-high-field n.m.r. studies of bovine lung heparan sulphate oligosaccharides produced by nitrous acid deaminative cleavage. 13C-n.m.r. study of methylene resonances: degree and positions of C-6 sulphation

1983 ◽  
Vol 211 (3) ◽  
pp. 677-682 ◽  
Author(s):  
P N Sanderson ◽  
I A Nieduszynski ◽  
T N Huckerby
Keyword(s):  
Nitrous Acid ◽  
High Field ◽  
Uronic Acid ◽  
Glucuronic Acid ◽  
General Structure ◽  
Heparan Sulphate ◽  
Glcnac Residue ◽  
Very High

Oligosaccharides with the general structure UA-(GlcNAc-GlcUA-)m-aManOH (m = 1-5) (where UA represents uronic acid, GlcNAc N-acetylglucosamine, GlcUA glucuronic acid and aManOH anhydromannitol) were prepared from low-sulphated heparan sulphates of bovine lung origin by nitrous acid deaminative cleavage followed by reduction. Analysis of the methylene signals in the 100 MHz 13C-n.m.r. spectrum of the tetrasaccharide (m = 1) shows that, whereas the extent of C-6 O-sulphation in the GlcNAc is approx. 65%, in the aManOH [formerly a GlcNSO3 (N-sulphoglucosamine) residue in the parent heparan sulphate] it is only approx. 10%. In the higher oligosaccharides (m = 2-5) the gross extent of C-6 O-sulphation of GlcNAc residues falls systematically with increasing oligosaccharide size, whereas that in the aManOH residues remains below 10%. There is also evidence that the C-6 O-sulphation of the GlcNAc residues is confined to the GlcNAc residue adjacent to the non-reducing terminal uronic acid residue. It is therefore tentatively proposed that the GlcNAc in the sequence -GlcNSO3-UA-GlcNAc- might be a favoured substrate for the 6-O-sulphotransferase. It is concluded that in the low-sulphated heparan sulphates GlcNSO3 residues that do not occur in (GlcNSO3-UA-)n blocks tend to have a significantly smaller extent of C-6 O-sulphation than do GlcNAc residues that occur in -GlcNSO3-UA-GlcNAc-GlcUA-GlcNSO3-sequences.

scholarly journals Very-high-field n.m.r. studies of bovine lung heparan sulphate tetrasaccharides produced by nitrous acid deaminative cleavage. Determination of saccharide sequence, uronate composition and degrees of sulphation

1984 ◽  
Vol 223 (2) ◽  
pp. 495-505 ◽  
Author(s):  
P N Sanderson ◽  
T N Huckerby ◽  
I A Nieduszynski
Keyword(s):  
Nitrous Acid ◽  
High Field ◽  
General Structure ◽  
Heparan Sulphate ◽  
Gel Filtration ◽  
Exchange Chromatography ◽  
Very High ◽  
Specific Sequences

Tetrasaccharides with the general structure UA-GlcNAc-GlcUA-aManOH (where UA represents uronate, GlcNAc N-acetylglucosamine, GlcUA glucuronate and aManOH anhydromannitol) were prepared from low-sulphated heparan sulphates of bovine lung origin by complete nitrous acid deaminative cleavage followed by reduction and fractionated by gel filtration. Ion-exchange chromatography of the tetrasaccharides yielded three major fractions in approximate yields of 37%, 45% and 14%. These were shown to be non-, mono- and di-sulphated respectively. Complete structural characterization of the tetrasaccharide fractions by quantitative high-field n.m.r. spectroscopy showed that each fraction contained only two discrete species and led to the following observations. (1) All of the uronate residues in the tetrasaccharides (and in larger oligosaccharides) are unsulphated, and hence sulphated iduronate [IdUA(2SO3)] must occur exclusively within -GlcNSO3-IdUA(2SO3)-GlcNSO3- sequences (where GlcNSO3 represents N-sulpho-glucosamine) in the parent polymers. (2) The GlcNAc residues in the tetrasaccharides are more highly C-6-O-sulphated than are the aManOH residues, and furthermore sulphation on the aManOH appears to occur only where the GlcNAc is also sulphated. (3) Where the GlcNAc is C-6-O-sulphated, iduronate is the major non-reducing terminal residue, whereas glucuronate predominates in this position if the GlcNAc is unsulphated. The quantitative data obtained are used to determine the degree of C-6-O-sulphation of glucosamine residues in specific sequences within the parent heparan sulphates.

scholarly journals Structural studies on heparan sulphate from human lung fibroblasts. Characterization of oligosaccharides obtained by selective periodate oxidation of d-glucuronic acid residues followed by scission in alkali

1980 ◽  
Vol 191 (1) ◽  
pp. 103-110 ◽  
Author(s):  
Ingrid Sjöberg ◽  
Lars-Ȧke Fransson
Keyword(s):  
Uronic Acid ◽  
Glucuronic Acid ◽  
Human Lung ◽  
General Structure ◽  
Heparan Sulphate ◽  
Periodate Oxidation ◽  
Exchange Chromatography ◽  
Lung Fibroblasts ◽  
Human Lung Fibroblasts ◽  
Iduronic Acid

1. 3H- and 35S-labelled heparan sulphate was isolated from monolayers of human lung fibroblasts and subjected to degradations by (a) deaminative cleavage and (b) periodate oxidation/alkaline elimination. Fragments were resolved by gel- and ion-exchange-chromatography. 2. Deaminative cleavage of the radioactive glycan afforded mainly disaccharides with a low content of ester-sulphate and free sulphate, indicating that a large part (approx. 80%) of the repeating units consisted of uronosyl-glucosamine-N-sulphate. Blocks of non-sulphated [glucuronosyl-N-acetyl glucosamine] repeats (3–4 consecutive units) accounted for the remainder of the chains. 3. By selective oxidation of glucuronic acid residues associated with N-acetylglucosamine, followed by scission in alkali, the radioactive glycan was degraded into a series of fragments. The glucuronosyl-N-acetylglucosamine-containing block regions yielded a compound N-acetylglucosamine–R, where R is the remnant of an oxidized and degraded glucuronic acid. Periodate-insensitive uronic acid residues were recovered in saccharides of the general structure glucosamine–(uronic acid–glucosamine)n–R. 4. Further degradations of these saccharides via deaminative cleavage and re-oxidations with periodate revealed that iduronic acid may be located in sequences such as glucosamine-N-sulphate→iduronic acid→N-acetylglucosamine. Occasionally the iduronic acid was sulphated. Blocks of iduronic acid-containing repeats may contain up to five consecutive units. Alternating arrangements of iduronic acid- and glucuronic acid-containing repeats were also observed. 5. 3H- and 35S-labelled heparan sulphates from sequential extracts of fibroblasts (medium, EDTA, trypsin digest, dithiothreitol extract, cell-soluble and cell-insoluble material) afforded similar profiles after both periodate oxidation/alkaline elimination and deaminative cleavage.

scholarly journals Structure of heparan sulphate oligosaccharides and their degradation by exo-enzymes

1979 ◽  
Vol 183 (3) ◽  
pp. 711-720 ◽  
Author(s):  
A Linker
Keyword(s):  
Structural Analysis ◽  
Nitrous Acid ◽  
Uronic Acid ◽  
Glucuronic Acid ◽  
Heparan Sulphate ◽  
Major Fraction ◽  
Sulphate Content ◽  
Acid Degradation ◽  
Iduronic Acid ◽  
Heparan Sulphate Chain

Oligosaccharides obtained from heparan sulphate by nitrous acid degradation were shown to be degraded sequentially by beta-D-glucuronidase or alpha-L-iduronidase followed by alpha D-N-acetylglucosaminidase. Structural analysis of the tetrasaccharide fraction showed the following. (1) N-Acetylglucosamine is preceded by a non-sulphated uronic acid residue that can be either D-glucuronic of L-iduronic acid, but followed by a glucuronic acid residue. (2) The N-acetylglucosamine in the major fraction is sulphated. (3) Very few if any of the uronic acid residues are sulphated (4). The results indicate that the area of the heparan sulphate chain where disaccharides containing N-acetylglucosamine and N-sulphated glucosamine residues alternate is higher in sulphate content than expected and that the sulphate groups are mainly located on the hexosamine units.

scholarly journals Molecular distinctions between heparan sulphate and heparin. Analysis of sulphation patterns indicates that heparan sulphate and heparin are separate families of N-sulphated polysaccharides

1985 ◽  
Vol 230 (3) ◽  
pp. 665-674 ◽  
Author(s):  
J T Gallagher ◽  
A Walker
Keyword(s):  
Uronic Acid ◽  
Heparan Sulphate ◽  
Analytical Data ◽  
Gel Filtration ◽  
Published Data ◽  
Sulphated Polysaccharides ◽  
Close Proximity ◽  
Sulphated Glycosaminoglycans ◽  
Very High ◽  
High Voltage Electrophoresis

Heparan sulphate and heparin are chemically related alpha β-linked glycosaminoglycans composed of alternating sequences of glucosamine and uronic acid. The amino sugars may be N-acetylated or N-sulphated, and the latter substituent is unique to these two polysaccharides. Although there is general agreement that heparan sulphate is usually less sulphated than heparin, reproducible differences in their molecular structure have been difficult to identify. We suggest that this is because most of the analytical data have been obtained with degraded materials that are not necessarily representative of complete polysaccharide chains. In the present study intact heparan sulphates, labelled biosynthetically with [3H]glucosamine and Na2(35)SO4, were isolated from the surface membranes of several types of cells in culture. The polysaccharide structure was analysed by complete HNO2 hydrolysis followed by fractionation of the products by gel filtration and high-voltage electrophoresis. Results showed that in all heparan sulphates there were approximately equal numbers of N-sulpho and N-acetyl substituents, arranged in a similar, predominantly segregated, manner along the polysaccharide chain. O-Sulphate groups were in close proximity to the N-sulphate groups but, unlike the latter, the number of O-sulphate groups could vary considerably in heparan sulphates of different cellular origins ranging from 20 to 75 O-sulphate groups per 100 disaccharide units. Inspection of the published data on heparin showed that the N-sulphate frequency was very high (greater than 80% of the glucosamine residues are N-sulphated) and the concentration of O-sulphate groups exceeded that of the N-sulphate groups. We conclude from these and other observations that heparan sulphate and heparin are separate families of N-sulphated glycosaminoglycans.

scholarly journals Distribution of sulphate and iduronic acid residues in heparin and heparan sulphate

1974 ◽  
Vol 137 (1) ◽  
pp. 33-43 ◽  
Author(s):  
Magnus HÖÖk ◽  
Ulf Lindahl ◽  
Per-Henrik Iverius
Keyword(s):  
Uronic Acid ◽  
Glucuronic Acid ◽  
Heparan Sulphate ◽  
Molar Ratio ◽  
Human Aorta ◽  
Hydroxyl Groups ◽  
Free System ◽  
Molar Ratios ◽  
Iduronic Acid ◽  
A Cell

1. A method was developed for determination of the uronic acid composition of heparin-like glycosaminoglycans. Polymers or oligosaccharides are degraded to monosaccharides by a combination of acid hydrolysis and deamination with HNO2. The resulting uronic acid monosaccharides (accounting for about 70% of the uronic acid contents of the starting materials) are isolated and converted into the corresponding aldono-1,4-lactones, which are separated by g.l.c. The calculated ratios of glucuronic acid/iduronic acid are reproducible within 5%. 2. Samples of heparin from pig intestinal mucosa (molar ratio of sulphate/disaccharide unit, 2.40) and heparan sulphate from human aorta (sulphate/disaccharide ratio, 0.46) were subjected to uronic acid analysis. l-Iduronic acid constituted 77% and 19% respectively of the total uronic acid contents. 3. The correlation between the contents of sulphate and iduronic acid indicated by this finding also applied to the fractionated deamination products of the two polymers. The sulphated fragments varied in size from disaccharide to octasaccharide (or larger) and showed sulphate/disaccharide molar ratios in the range of 0.05–2.0. The proportion of iduronic acid increased with increasing ester sulphate contents of the oligosaccharides. 4. Previous studies on the biosynthesis of heparin in a cell-free system have shown that l-iduronic acid residues are formed by C-5 epimerization of d-glucuronic acid units at the polymer level; the process requires concomitant sulphation of the polymer. The results obtained in the present structural study conform to these findings, and suggest further that similar mechanisms may operate in the biosynthesis of heparan sulphate. The epimerization reaction appears to be linked to the sulphation of hydroxyl groups but does not seem to require sulphation of the target uronic acid residues. The significance of sulphamino groups in relation to the formation of iduronic acid is unknown.

Binding of lead and copper(II) cations to hemicelluloses of rye bran

1986 ◽  
Vol 51 (10) ◽  
pp. 2250-2258 ◽  
Author(s):  
Rudolf Kohn ◽  
Zdena Hromádková ◽  
Anna Ebringerová
Keyword(s):  
Uronic Acid ◽  
Equilibrium Solution ◽  
Glucuronic Acid ◽  
The Other ◽  
Carboxyl Groups ◽  
Stoichiometric Ratio ◽  
Molar Ratios ◽  
Free Ions ◽  
Uronic Acid Content ◽  
The Relationship

Several fractions of acid hemicelluloses isolated from rye bran were characterized by molar ratios of saccharides (D-Xyl, L-Ara, D-Glc, D-Gal) and 4-O-methyl-D-glucuronic acid and protein content. Binding of Pb2+ and Cu2+ ions to these acid polysaccharides was considered according to function (M)b = f([M2+]f), expressing the relationship between the amount of metal (M)b bound to 1 g of the substance and the concentration of free ions [M2+]f in the equilibrium solution and according to the association degree β of these cations with carboxyl groups of uronic acid at a stoichiometric ratio of both components in the system under investigation. Acid hemicelluloses contained only a very small portion of uronic acid ((COOH) 0.05-0.18 mmol g-1); the model polysaccharide, 4-O-methyl-D-glucurono-D-xylan of beech, was substantially richer in uronic acid content ((COOH) 0.73 mmol g-1). Consequently, the amount of lead and copper bound to acid hemicelluloses is very small ((M)b 0.017-0.025 mmol g-1) at [M2+]f = 0.10 mmol l-1. On the other hand, much greater amount of cations ((M)f 0.09-0.10 mmol g-1) was bound to the glucuronoxylan. The association degree β was like with the majority of samples (β = 0.31-0.38). The amount of lead and copper(II) bound to acid hemicelluloses from rye bran is several times lower than that bound to dietary fiber isolated from vegetables (cabbage, carrot), rich in pectic substances.

Very high field EPR study of a molecular nanomagnet

1998 ◽  
Vol 177-181 ◽  
pp. 709-710 ◽  
Author(s):  
A.L. Barra ◽  
A. Caneschi ◽  
D. Gatteschi ◽  
R. Sessoli
Keyword(s):  
High Field ◽  
Very High

SECMA 1®, a mitogenic hexapeptide from Ulva algeae modulates the production of proteoglycans and glycosaminoglycans in human foreskin fibroblast

1998 ◽  
Vol 17 (1) ◽  
pp. 18-22 ◽  
Author(s):  
R Ennamany ◽  
D Saboureau ◽  
N Mekideche ◽  
E E Creppy
Keyword(s):  
Glucuronic Acid ◽  
De Novo ◽  
Heparan Sulphate ◽  
Guanidine Hydrochloride ◽  
Sodium Deoxycholate ◽  
Effective Concentration ◽  
Skin Fibroblasts ◽  
Human Skin Fibroblasts ◽  
Dermatan Sulphate ◽  
Salt Extract

SECMA 1® is a polypeptide purified from a green algeae of the Ulva species by several gel chromatographies, showing the following sequence (Glu-Asp-Arg-Leu-Lys-Pro). In order to determine the effect of SECMA 1® on human skin fibroblasts extracellular matrix, proteoglycans (PGs) and glycosaminoglycans (GAGs) were assayed after 24 h incubation of 20 day-old foreskin fibroblasts at the 2nd passage. The results revealed that most of [35S]sulphate was associated with fibroblast membranes, which contained (67%) of the total de novo synthesized sulphated PGs, in two distinct forms: one hydrophilic (39%), and one hydrophobic (28%). The remaining `matrix' retained 5% of proteoglycans. The remaining 35S-label may represent the free label in the cytosol. After 24 h incubation of skin fibroblasts with different concentrations of SECMA 1® (2, 4 and 10 μg/ml), the [35S] sulphate incorporation into PGs of Salt-extract, sodium deoxycholate (DOC) extract and Guanidine hydrochloride (GuA-HCl)-extract was increased significantly ( P<0.005) with 4 μg/ml, as compared to untreated control. The most effective concentration (4 μg/ml) increased the different [35S]sulphate PGs extracts (NaCl, DOC and GuA-HCl) by respectively (66; 17 and 75%). The relative contents of iduronic and glucuronic acid in the GAG produced by skin fibroblasts were estimated. No effect of SECMA 1® on the incorporation of [35S]sulphate into Heparan sulphate was found. The incorporation of [35S]sulphate into (chondroïtine sulphate + heparan sulphate) and (chondroïtine sulphate + dermatan sulphate) was increased by respectively 37% and 11% by SECMA 1® (4 μg/ml).

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