Chemoenzymatic synthesis of spacer-linked oligosaccharides for the preparation of neoglycoproteins.
In the present work, the combination of chemical and enzymatic methods to obtain neoglycoproteins is described. Three bovine serum albumin (BSA)-conjugates, BSA-[GalNAc alpha-], BSA-[Gal(beta 1-3)GalNAc(alpha-], and BSA-[Neu5Ac(alpha 2-3)Gal(beta 1-3)GalNAc(alpha-], were prepared. alpha GalNAc derivatives were galactosylated employing crude beta-galactosidase from bovine testes. The use of oversaturated donor solutions (pNP beta Gal) enhanced the yields up to 60%. This method was verified using divalent structures as acceptors, that rendered di- and tri-galactosylated products. Further treatment of the disaccharides with CMP-Neu5Ac and alpha 2-3 sialyltransferase from pork liver led to formation of trisaccharides. Finally, mono-, di-, and trisaccharides were coupled to BSA employing a thiolic group introduced into the protein for Michael addition to a maleinimide group in the spacer-arm of the saccharide components. The results were monitored by HPLC and MALDI-TOF.
Gonzalez Lio
R
R
Institut für Organische Chemie, Universität Hamburg, Germany.
Thiem
J
J
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002417:Cattle; D004187:Disaccharides / Q000138:chemical synthesis* / Q000737:chemistry; D005690:Galactose; D006023:Glycoproteins / Q000138:chemical synthesis* / Q000737:chemistry; D007202:Indicators and Reagents; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis* / Q000737:chemistry; D009904:Optical Rotation; D012710:Serum Albumin, Bovine; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014312:Trisaccharides / Q000138:chemical synthesis* / Q000737:chemistry; D001616:beta-Galactosidase
D004187:Disaccharides; D006023:Glycoproteins; D007202:Indicators and Reagents; D009844:Oligosaccharides; D014312:Trisaccharides; D012710:Serum Albumin, Bovine; D001616:beta-Galactosidase; D005690:Galactose