Abstract
The carbohydrate-binding specificity of Aleuria aurantia lectin was investigated by analyzing the behavior of a variety of fucose-containing oligosaccharides on an A. aurantia lectin-Sepharose column. Studies with complex-type oligosaccharides obtained from various glycoproteins by hydrazinolysis and their partial degradation fragments indicated that the presence of the alpha-fucosyl residue linked at the C-6 position of the proximal N-acetylglucosamine moiety is indispensable for binding to the lectin column. Binding was not affected by the structures of the outer chain moieties nor by the presence of the bisecting N-acetylglucosamine residue. These results indicated that A. aurantia lectin-Sepharose is useful for the group separation of mixtures of complex-type asparagine-linked sugar chains. Studies of glycosylated Bence Jones proteins indicated that this procedure is also applicable to intact glycoproteins. The behavior of oligosaccharides isolated from human milk and the urine of patients with fucosidosis indicated that the oligosaccharides with Fuc alpha 1----2Gal beta 1----4GlcNAc and Gal beta 1----4(Fuc alpha 1----3)GlcNAc groups interact with the lectin, but less strongly than complex-type sugar chains with a fucosylated core. Lacto-N-fucopentaitol II, which has a Gal beta 1----3(Fuc alpha 1----4)GlcNAc group, interacts less strongly than the above two groups with the matrix. Oligosaccharides with Fuc alpha 1----2Gal beta 1----3GlcNAc and Gal beta 1----4GlcNAc beta 1----3Gal beta 1----4(Fuc alpha 1----3)GlcNAc groups showed almost no interaction with the matrix.
Highlights
The detailed investigation of the sugar binding specificity of aurantia lectin (AAL) described inthispaper, revealed thatthe AALSepharose column can be used as an effective tool for the fractionation of glycoproteins as well as oligosaccharides according to their contenot f a-fucosyl linkages
Behavior of Oligosaccharides Related to Complex-type Asparagine-linked Sugar Chains-When each of the 9 tritiumlabeled oligosaccharides (I-IX) liberated from the glycoproteins byhydrazinolysiswasapplied to an AAL-Sepharose column, allof those with a n a-fucosyl residue on their reducing terminal N-acetylglucosamine residue were bound to the column andelutedwith buffer containing 0.5 mM fucose
The actual elution profiles of the three biantennary oligosaccharides, I, 11, and 111 are shown as solid lines inFig. 1,A, B, and C, respectively. These results indicated that the binding of oligosaccharides is not altered by the number of outer chain moieties nor by the presence of the bisecting N-acetylglucosamine residue
Summary
Materials-AAL was purified from the fruiting bodies of A. aurantia according to the method reported previously [13]. Purified AAL was coupled to Sepharose 4B by the procedure of March et al [14]. Oligosaccharides XII, XIV, XV, XVIII-XXI, XXV, and XXVI were isolated from human milk as described previously [17,18] and labeled by reduction with NaB[3H]4(31). Oligosaccharide XXII was isolated from the N-3 fraction [17] of milk obtained from an Le(a-b-), nonsecretor individual.’. The abbreviations used are: AAL, Aleuria aurantia lectin; subscript OT is used to indicate NaB[3H]r-reducedoligosaccharides (all sugars mentioned in this paper wereof D-configuration except for fucose whichhas an L-configuration). Structures of typical oligosaccharides widely distributed in various glycoproteins, and their behavioro n a n AAL-Sepharose column
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