The structure of a neural specific carbohydrate epitope of horseradish peroxidase recognized by anti-horseradish peroxidase antiserum.
Antiserum raised against horseradish peroxidase (HRP) recognizes a neural specific carbohydrate antigen in Drosophila and other insects. The epitopic activity of the carbohydrate moiety of HRP recognized by anti-HRP antiserum was measured by a newly developed enzyme-linked immunosorbent assay, in which HRP glycopeptides conjugated with bovine serum albumin were coated onto the wells and then reacted with goat anti-HRP antiserum. HRP sugar moieties released by almond glycopeptidase A digestion of HRP pepsin digests were subjected to pyridylamination. Pyridylamino oligosaccharides were separated into seven fractions by reverse-phase high performance liquid chromatography. The major fraction, which comprised about 80% of the total sugars, reacted strongly with anti-HRP antiserum. The carbohydrate structure of this fraction was determined by sugar composition analysis and 600-MHz 1H NMR spectroscopy as follows: Man alpha 1----6(Man alpha 1----3)(Xyl beta 1----2)Man beta 1----4GlcNAc beta 1----4(Fuc alpha 1----3)GlcNAc. Analyses of reactivity with anti-HRP antiserum of various oligosaccharide derivatives obtained from the major fraction by exoglycosidase digestion and partial acid hydrolysis indicated that alpha 1----6-linked mannose and alpha 1----3-linked fucose are predominantly involved in the epitopic structure.
Kurosaka
A
A
Department of Biotechnology, Faculty of Engineering, Sangyo University, Japan.
Yano
A
A
Itoh
N
N
Kuroda
Y
Y
Nakagawa
T
T
Kawasaki
T
T
D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes; D006001:Glycoconjugates / Q000276:immunology*; D006735:Horseradish Peroxidase / Q000737:chemistry / Q000276:immunology*; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009474:Neurons / Q000276:immunology*; D009844:Oligosaccharides / Q000276:immunology
D000939:Epitopes; D006001:Glycoconjugates; D009844:Oligosaccharides; D006735:Horseradish Peroxidase