Fractionation of a hyaluronic acid preparation in a density gradient. The isolation and identification of a chondroitin sulphate

Silpananta, Panee; Dunstone, J. R.; Ogston, A. G.

doi:10.1042/bj1040404

Cited by 40 publications

(25 citation statements)

References 22 publications

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“…Franek and Dunstone (1966), Silpananta et al (1967) and Creeth and Denborough (1970) consider that protein and polysaccharide sedimenting together in caesium chloride gradients are covalently linked glycoproteins, and they have applied this method to separate protein and glycoprotein quantitatively.…”

Section: Discussionmentioning

confidence: 99%

A Water-soluble Arabinogalactan-Peptide From Wheat Endosperm

Fincher¹,

Stone²

1974

Aust. Jnl. Of Bio. Sci.

149

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The water-soluble polymeric components of wheat endosperm have been extracted by two different procedures and their chemical composition studied in detail. Water extracts of wheat flour that had first been treated with hot 80% ethanol contained only 2% protein, but if the ethanol treatment was omitted up to 20% of the extracted polymeric fraction was low-molecular-weight non-dialysable protein material. Density-gradient ultracentrifugation in caesium chloride solutions indicated that most of this protein was free, whereas the 2% protein in the water extract of ethanol-treated flottr was firmly bound to a polysaccharide. This bound protein (a peptide) was characterized by high levels of hydroxyproline (16-20% of the amino acids present).Fractional precipitation of the water-soluble components with ammonium sulphate enabled the separation of a high-molecular-weight arabinoxylan (xylose : arabinose = 1·6 -1 . 9) and a lowmolecular-weight arabinogalactan (galactose :. arabinose =approx. 1· 5), the latter. being associated with the hydroxyproline-rich peptide.

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Section: Discussionmentioning

confidence: 99%

A Water-soluble Arabinogalactan-Peptide From Wheat Endosperm

Fincher¹,

Stone²

1974

Aust. Jnl. Of Bio. Sci.

149

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“…Earlier biochemical work revealed the presence of chondroitin sulfate in synovial fluid (19)(20)(21) and showed that there were increases in the CS content relative to that of hyaluronic acid (2Q,21). Others have described cartilage proteoglycan-like fragments in normal ox synovial fluid of molecular weight 250,000 (22). Recently, using a polyclonal antibody to cartilage proteoglycan, increased amounts of proteoglycan were detected by enzyme-linked immunosorbent assay in the synovial fluids of patients with reactive arthritis (23).…”

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confidence: 99%

The immunologic detection and characterization of cartilage proteoglycan degradation products in synovial fluids of patients with arthritis

Witter

Webber

et al. 1987

Arthritis & Rheumatism

102

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Antibodies were used in radioimmunoassays with gel chromatography to detect the hyaluronic acidbinding region, core protein, and keratan sulfate of human cartilage protoeglycan in the synovial fluids of patients with rheumatoid arthritis, juvenile rheumatoid arthritis, and osteoarthritis. All fluids contained proteoglycan that was mainly included on Sepharose CL4B; this result indicates cleavage of proteoglycan (which is normally excluded). The hyaluronic acidbinding region was the smallest and most commonly detected fragment. It was relatively free of keratan sulfate and core protein, and it could sometimes bind to hyaluronic acid. Other larger fragments containing core protein and/or keratan sulfate were detected in every fluid.

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“…AnaIytical density gradient sedimentation was performed in a Beckman model E analytical ultracentrifuge [5,6] Serological activity was determined by the capacity of the substances to inhibit in specific haemagglutination tests [17]. Fig.…”

Section: Sedimentationmentioning

confidence: 99%

Fractionation in a Density Gradient of some Glycoproteins from Human Ovarian Cyst Fluids

Dunstone

1969

European Journal of Biochemistry

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Equilibrium density gradient sedimentation in caesium chloride has been used as an eiTective method of separating the components of sulphite-reduced sparingly soluble glycoproteins from two human ovarian cyst fluids. An almost quantitative recovery of the carbohydrate and peptide constituents has been obtained. Some fractions had overall chemical compositions similar to those isolated previously by Dunstone and Morgan in 1965, while others contained much greater proportions of amino acids, especially of aspartic acid, glutamic acid and cystine.The results suggest that the sparingly soluble glycoproteins are large macromolecules containing serologically active carbohydrate chains, bound to specific peptide fragments which form an integral part of larger macromolecules, the insolubility of which is due to the presence of -S-S-bonds.The experiments emphasize the value of equilibrium density gradient sedimentation in the study of complex proteinpolysaccharide systems.Human blood group-specific glycoproteins are obtained from pseudomucinous ovarian cyst fluids by extraction of the freeze-dried fluids with 90--95O/, w/w phenol [l-31. I n most instances, water-soluble glycoproteins that have been well-characterized, and sparingly soluble glycoproteins about which little is known, are obtained. As a result of preliminary studies of the sparingly soluble gel-like substances Dunstone and Morgan [4], have suggested that these are large macromolecules composed of a t least two types of amino acid-containing structure associated through inter-molecular -S-S-bonds. One of these structures was identified, after reduction of the gel-like material, as a typical water-soluble blood group-specific glycoprotein. Although the other type of structure was not isolated as a discrete molecular species, the experiments suggested that it should contain a relatively high proportion of amino acids, especially of aspartic acid, glutamic acid and cystine. These conclusions were considered tentative, because the recoveries were not quantitative and because only a limited number of preparations were examined.Equilibrium density gradient sedimentation in caesium chloride is an effective method for the quantitative separation of proteinpolysaccharides with minimal degradation [5-81. The present paper describes the fractionation, by this method, of sulphitereduced, sparingly soluble glycoproteins from two human ovarian cyst fluids. Excellent recoveries of the carbohydrate and amino acid constituents were obtained. Some of the fractions had overall chemical compositions similar to those isolated previously The experiments emphasize the value of equilibrium density gradient sedimentation in the study of complex proteinpolysaccharide systems. MATERIAL AND METHODSOvarian Cyst Glycoproteins These were prepared as described by Dunstone and Morgan [a]. The sparingly soluble glycoproteins obtained from cyst fluids no.485 (groupA) and no. 531 (group B)' were reduced with sulphite at room temperature as follows [9] : 100 mg of the glycoprotein were suspende...

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Fractionation of a hyaluronic acid preparation in a density gradient. The isolation and identification of a chondroitin sulphate

Cited by 40 publications

References 22 publications

A Water-soluble Arabinogalactan-Peptide From Wheat Endosperm

A Water-soluble Arabinogalactan-Peptide From Wheat Endosperm

The immunologic detection and characterization of cartilage proteoglycan degradation products in synovial fluids of patients with arthritis

Fractionation in a Density Gradient of some Glycoproteins from Human Ovarian Cyst Fluids

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