Purification, Composition and Immunochemical Properties of Arabinogalactan-Protein H Active Glycopeptides fromEuonymus Sieboldiana Seeds

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Abstract
Seven blood group H active glycopeptides were isolated from the defatted seeds of Euonymus Sieboldiana by a procedure that included extraction with cold phosphate-buffered saline, fractionation with ammonium sulfate, chromatography with CM- and DEAE Sepharose CL 6B®, and gel filtration on Sepharose 4B® and Sephacryl S-200®. Each glycopeptide, except three macromolecular ones, eluted as a single symmetrical peak from the Sephacryl S-200® chromatography. On the basis of their elution from a column that had been calibrated with FITC-dextrans as molecular weight markers, these glycopeptides had molecular weights of approximately 110,000, 15,000 and 8,000. All isolated H active glycopeptides inhibited eel anti-H reagent at concentrations of 1 to 63 μg/ml and exhibited a single precipitation band against the eel reagent on immunodiffusion. Furthermore, all of them contained galactose and arabinose as major sugars, with fucose, rhamnose, mannose, glucose and glucosamine as minor sugars. Amino acid analysis of the four H active glycopeptides retarded by the Sepharose 4B® column revealed the presence of hydroxyproline together with large amounts of threonine and serine. However, the three excluded macromolecular H active glycopeptides contained both threonine and serine, but no hydroxyproline.