Go Matsumura

Structures of carbohydrate units isolated from trout egg polysialoglycoproteins: short-cored units with oligosialosyl groups.

Abstract The eggs of rainbow trout contain a novel type of glycoproteins in which sialic acid (NeuGc) accounts for about 60% of the weight. Of more than 20 oligosaccharide fractions containing various numbers of NeuGc residues obtained by alkaline borohydride treatment of these polysialoglycoproteins followed by an anion-exchange chromatography, structures of a series of oligosaccharides, Galβ1–3[(NeuGcα2–8) n NeuGcα2–6]GalNAcol ( n = 0–3), have been determined. 1 H-NMR signals for H-3 protons of NeuGc residues in oligosialosyl groups were assigned using model NeuAc oligomers prepared by limited acid hydrolysis of colominic acid.

Endo-β-galactosidase of Eschericia freundii hydrolysis of pig colonic mucin and milk oligosaccharides by endoglycosidic action

Abstract The purified keratansulfate degrading enzyme from Eschericia freundii could hydrolyze desialyzed pig colonic mucin and milk oligosaccharides. Desialyzed pig colonic mucin was digested to produce GlcNAcβ(1→3)Gal, GlcNAc-6Sβ(1→3)Gal and resistant polymer. Lacto-N-tetraose and lacto-N-tetraitol were hydrolyzed endoglycosidically to release glucose and sorbitol, respectively. Therefore, this enzyme was found to be an endo-β-galactosidase of rather wide specificity.

Novel carbohydrate structures in trout egg glycoprotein: Occurrence of a neuraminidase-resistant N-glycolylneuraminosyl-(2→3)-N-acetylgalactosamine linkage

Summary Oligosaccharides with a new type of core structure, GalNAcβ1→4 Galβl→4[NGNA2→3]GalNAcβ1→3Ga1β1→3Ga1NAco1, have been isolated from trout egg glycoprotein. The occurrence of N-glycolylneuraminic acid (NGNA) linked 2→3 to the internal Ga1NAc residue is not known in any other glycoproteins and glycolipids. This NGNA residue was found to show some anomalous nature such as resistance to neuraminidases and unusual chemical shift values of H-3 protons in NMR spectra. A new feature is also found in the asialooligosaccharide structure, Ga1NAcβl→4Galβ1→4Ga1NAc, linked β1→3 to the linkage region disaccharide, Ga1β1→3Ga1NAc.

Stability of N‐acyl groups of neuraminic acid residues in 2→8 linked polymers toward methanolysis used in methylation analysis

Acid methanolysis has been found preferable to hydrolysis in methylation analysis of ohgosaccharide chains of hexosaminecontaining glycolipids [ 1], and of diand polysialosyl structures which have recently been found to occur in glycolipids [2] and glycoproteins f3-4 as well as in colominic acid [2,6]. Ahhough the stability of N-acyl groups of nonmethylated sialic acid toward rne~~olys~ has been described {7], no description on the corres~nd~g stability of methylated sialic acid occurs in the literature. While non-methylated N-acetylhexosamines undergo de-J%acetylation under the conditions of methanolysis used to cleave glycosidic linkages and re-N-acetylation is necessary before gas-liquid chromatography (GLC) IS], the applied methanolysis conditions for fully methylated carbohydrate did not accompany de&-acetylation of Nacetylglucosamine residues .f9]. In the course of me~ylation analysis of the structure of novel carbohydrate chains recently found in the glycoproteins from rainbow trout [49], the internal 8-Osubstituted N-glycolylneuraminic acid residues of methyhrted saccharides were found to undergo de-llracylation whereas the terminal fully methytated Nglycolylneuraminic acid residues do not under the conditions of methanolysis widely employed in similar studies, and suggested participa-

Stability of N-acyl groups in methylated α2 → 8-linked oligosialosyl chains toward methanolysis. Analysis by chemical ionization-mass spectrometry

Abstract The stability of N-acetyl group of methylated trisaccharide of N-acetylneuraminic acid toward methanolysis under conditions used in methylation analysis was investigated. The analysis of the products obtained after a reaction sequence, methylation-methanolysis-deuterioacetylation, by chemical ionization-mass spectrometry has led to unequivocal conclusion that N-acetyl group of internal 8-O-substituted residue of the methylated oligosialosyl compound is de-N-acetylated under conditions sufficient to cleave glycosidic linkages, whereas the fully methylated nonreducing terminal residue of neuraminic acid is completely resistant to de-N-acetylation. The reaction mechanism to explain these observations is presented.

Comparison of relationships between the chemical structures and mobilities of hyaluronate oligosaccharides in thin-layer and high-performance liquid chromatography

Abstract The R M values of odd- and even-numbered hyaluronate oligosaccharides, comprised of N-acetylglucosamine and glucuronic acid residues, were determined by thin-layer chromatography (TLC). Previous retention time data of the acidic oligosaccharides obtained by high performance liquid chromatography (HPLC) were converted into R M values. By dividing the oligosaccharide structures into several fragments, the contributions of these fragments to chromatographic mobility (group constants) were estimated essentially from the difference between the R M values of two oligomers having appropriate structures. The group constants of the bridging oxygen atoms at the β1,4- and -1,3-glycosidic linkages of these oligomers were identical in HPLC but not in TLC. In the two types of chromatography, the mobility of a given hyaluronate oligosaccharide could be explained by a linear combination of group constants and the R M value of N-acetylglucosamine or glucuronic acid, with the exception that the R M value of the uronic acid in TLC was anomalous.

Determination of Stokes radii and molecular masses of sodium hyaluronates by Sephacryl gel chromatography

Abstract On the basis of available volumes (Kav) of a chondroitin sulphate fraction and some proteins with known Stokes radii (R) in Sepharose (6B, 4B and 2B) gel chromatography, it was found that Ogstons equation correlating Kav with R was also applicable to Sepharose gel in addition to the Sephadex gel previously examined. Therefore, R values for sodium hyaluronate samples were determined in a similar manner to that already described for R values for hyaluronate oligosaccharides by Sephadex gel chromatography, using bovine serum albumin as a standard of known size and the radius (2.5 nm) of agarose chain forming the gel networks. The molecular masses (M) of these samples, estimated from their intrinsic viscosities according to Mark-Houwink equations, were in the range 3.9 · 104−3.0 · 105. Their Stokes radii determined by gel chromatography were in the range 5.9–14.4 nm. A simple relationship of R = 0.030 M 1 2 was maintained for the hyaluronates with M below 1.3 · 105 (R = 11 nm). The plots of R against M 1 2 gave R = 4.82 + 0.0175 M 1 2 for M above 1.3 · 105. By use of these relationships, the molecular mass hyaluronate can be determined from its Stokes radius under the experimental conditions.

The effect of link protein on the rheological properties of solutions of proteoglycan-hyaluronic acid aggregates

Abstract Proteoglycan monomers, prepared from bovine nasal cartilage, were reassembled with a commercial hyaluronic acid, with or without link protein. The effects of link protein on the rheological properties of proteoglycan aggregates were studied over the concentration range 0.1–2.0 g/100 ml. Aggregate formation of proteoglycan monomer with hyaluronic acid increased the dependence on shear rate and shifted the Newtonian region to lower shear rates, compared to that observed with a proteoglycan monomer solution at the same concentration. The dynamic viscoelasticity also increased. In the presence of the link protein there is no Newtonian flow region over the shear rate ranges at a concentration of 1 g/100 ml or above and the value of the loss tangent remained low over the angular frequencies examined. Link protein strengthens the intermolecular interactions increasing the relaxation time of the aggregate solution. It increases the co-operativity of the binding of the proteoglycan monomers to the hyaluronic acid chain allowing a dynamic network structure to be developed in solution. The rheology is strongly dependent on the weight ratio of the proteoglycan to hyaluronic acid.

Determination of stokes radii of hyaluronate oligosaccharides by Sephadex gel chromatography

Abstract Even-numbered hyaluronate oligosaccharides ranging from di- to hexadecasaccharides were chromatographed on Sephadex G-25, -50 and 075, and their available volumes (Kav) were examined. Log Kav ratios between any two samples of these oligomers were constant for the three Sephadex types used. The relationships and the use of the log Kav of a substance of known size simplified the processes for calculating Stokes radii of hyaluronate oligosaccharides.