Alpha-Galactosyl trisaccharide epitope: Modification of the 6-primary positions and recognition by human anti-αGal antibody
- 82 Downloads
Galactose oxidase (EC 188.8.131.52, GAO) was used to convert the C-6′ OH of Galβ(1 → 4)Glcβ–OBn (5) to the corresponding hydrated aldehyde (7). Chemical modification, through dehydratative coupling and reductive amination, gave rise to a small library of Galβ(1 → 4)Glcβ–OBn analogues (9a–f, 10, 11). UDP-[6-3H]Gal studies indicated that α1,3-galactosyltransferase recognized the C-6′ modified Galβ(1 → 4)Glcβ–OBn analogues (9a–f, 10, 11). Preparative scale reactions ensued, utilizing a single enzyme UDP-Gal conversion as well as a dual enzymatic system (GalE and α1,3GalT), taking full advantage of the more economical UDP-Glc, giving rise to compounds 6, 15–22. Galα(1 → 3)Galβ(1 → 4)Glcβ–OBn trisaccharide (6) was produced on a large scale (2 g) and subjected to the same chemoenzymatic modification as stated above to produce C-6″ modified derivatives (23–30). An ELISA bioassay was performed utilizing human anti-αGal antibodies to study the binding affinity of the derivatized epitopes (6, 15–30). Modifications made at the C-6′ position did not alter the IgG antibody's ability to recognize the unnatural epitopes. Modifications made at the C-6″ position resulted in significant or complete abrogation of recognition. The results indicate that the C-6′ OH of the αGal trisaccharide epitope is not mandatory for antibody recognition. Published in 2004.
Unable to display preview. Download preview PDF.
- 8.Barresi F, Hindsgaul O, Chemically synthesized oligosaccharides, 1994.Asearchable table of glycosidic linkages, J CarbohydrChem 14, 1043-87 (1995).Google Scholar
- 13.McAuliffe JC, Hindsgaul O, Carbohydrate drugs-an ongoing challenge, Chem & Ind 5, 170-4 (1997).Google Scholar
- 19.Xu Y, Lorf T, Sablinski T, Gianello P, Bailin M, Monroy R, Kozlowski T, Awwad M, Cooper DKC, Sachs DH, Removal of anti-porcine natural antibodies from human and nonhuman primate plasma in vitro and in vivo by a Gal?(1?3)Gal?(1?4)Gle?-X immunoaffinity column, Transplantation 65, 172-9 (1998).PubMedCrossRefGoogle Scholar
- 23.Palcic MM, Hindsgaul O, Glycosyltransferases in the synthesis of oligosaccharide analogs, Trends Glycosci Glycotechnol 8, 37-49 (1996).Google Scholar
- 25.Ohrlein R, Glycosyltransferase-catalyzed synthesis of non-natural oligosaccharides, Top Curr Chem 200, 227-54 (1999).Google Scholar
- 26.Blanken WM, Van den E, Dirk H, Biosynthesis of terminal Gal? (1?3)Gal?(1?4)GlcN Ac-R oligosaccharide sequences on glycoconjugates. Purification and acceptor specificity of aUDPGal: N-acetyllactosaminide ?1,3-galactosyltransferase from calf thymus, J Biol Chem 260, 12927-34 (1985).PubMedGoogle Scholar
- 27.Mazur A, Galactose Oxidase. In: Enzymes in Carbohydrate Synthesis, edited by Bednarski MD (Oxford University Press, New York, 1991), pp. 99-110.Google Scholar