Determination of the specificity of monoclonal antibodies against Schistosoma mansoni CAA glycoprotein antigen using neoglycoconjugate variants.
The immunogenic O-glycan of circulating anodic antigen (CAA) is a high-molecular-mass polysaccharide with the unique -->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GalpNAc-(1--> repeating unit. To obtain information at the molecular level about the specificity of monoclonal antibodies against CAA, the immunoreactivity of two series of bovine serum albumin-coupled synthetic oligosaccharides related to the CAA O-glycan was monitored using ELISA and surface plasmon resonance spectroscopy. The importance of the axial hydroxyl group of beta-D-GalpNAc for antibody binding was investigated using the following series of analogues: beta-D-GlcpA-(1-->3)-beta-D-GlcpNAc-(1-->O); beta-D-GlcpNAc-(1-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GlcpNAc-(1-->O); and beta-D-GlcpA-(1-->3)-beta-D-GlcpNAc-(1-->6)-[beta-D-GlcpA-(1-->3)]-beta-D-GlcpNAc-(1-->O). In the second series of analogues, beta-D-Glcp6S-(1-->3)-beta-D-GalpNAc-(1-->O), beta-D-GalpNAc-(1-->6)-[beta-D-Glcp6S-(1-->3)]-beta-D-GalpNAc-(1-->O), and beta-D-Glcp6S-(1-->3)-beta-D-Gal-pNAc-(1-->6)-[beta-D-Glcp6S-(1-->3)]-beta-D-GalpNAc-(1-->O), the native beta-D-GlcpA moiety was replaced by beta-D-Glcp6S to evaluate the influence of the nature of the charge on antibody recognition. Comparison of the immunoreactivity of these series with that measured for conjugates containing corresponding synthetic CAA fragments showed that the antibody binding levels can be correlated to the antibody specificity to CAA fragments. For the most reactive antibodies, the structural changes chosen (beta-D-GalpNAc replaced by beta-D-GlcpNAc, and beta-D-GlcpA replaced by beta-D-Glcp6S) completely eradicated the binding.
Carvalho de Souza
Adriana
A
Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, Padualaan 8, NL-3584 CH Utrecht, The Netherlands.
van Remoortere
Alexandra
A
Hokke
Cornelis H
CH
Deelder
André M
AM
Vliegenthart
Johannes F G
JF
Kamerling
Johannis P
JP
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology*; D000918:Antibody Specificity; D000947:Antigens, Helminth / Q000737:chemistry / Q000276:immunology*; D002240:Carbohydrate Sequence; D002417:Cattle; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000737:chemistry / Q000276:immunology*; D015801:Helminth Proteins / Q000737:chemistry / Q000276:immunology*; D006801:Humans; D018796:Immunoconjugates / Q000737:chemistry / Q000276:immunology*; D007700:Kinetics; D051379:Mice; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000138:chemical synthesis / Q000737:chemistry / Q000276:immunology*; D012550:Schistosoma mansoni / Q000276:immunology*; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000276:immunology; D020349:Surface Plasmon Resonance
D000911:Antibodies, Monoclonal; D000947:Antigens, Helminth; D006023:Glycoproteins; D015801:Helminth Proteins; D018796:Immunoconjugates; D009844:Oligosaccharides; C090907:circulating anodic antigen, Schistosoma mansoni; D012710:Serum Albumin, Bovine