Reduced immunogenicity of beta-lactoglobulin by conjugation with carboxymethyl dextran differing in molecular weight.
To reduce the immunogenicity of beta-lactoglobulin (beta-LG), two beta-LG-carboxymethyl dextran (CMD) conjugates (Conj. 40 and Conj. 162) were prepared by using water-soluble carbodiimide (EDC). The molar ratios of beta-LG to CMD in Conj. 40 and Conj. 162 were 8:1 and 7:1, respectively. Each conjugate maintained approximately 50% of the retinol binding activity of beta-LG. Structural analyses by intrinsic fluorescence, CD spectra, and ELISA with monoclonal antibodies indicated that the surface of beta-LG in each conjugate was covered by CMD without great disruption of native conformation. By conjugation with CMD, the antibody response to beta-LG was reduced in BALB/c, C3H/He, and C57BL/6 mice, which was eminent in Conj. 162. The results of B cell epitope scanning using overlapping synthesized peptides showed that the linear epitope profiles of the conjugates were similar to those of beta-LG, whereas the antibody response to each epitope was reduced, which was eminent in Conj. 162. It was concluded that conjugation with CMD of higher molecular weight is effective in reducing the immunogenicity of beta-LG and that masking of epitopes by CMD is responsible for the reduced immunogenicity.
Department of Applied Biological Science, Faculty of Agriculture, Tokyo University of Agriculture and Technology, Tokyo 183-8509, Japan.
K
K
Kobayashi
A
A
Hirano
A
A
Ohta
T
T
Yoshida
K
K
Takahashi
M
M
Hattori
D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis*; D000917:Antibody Formation; D001402:B-Lymphocytes / Q000276:immunology*; D003911:Dextrans / Q000737:chemistry / Q000276:immunology*; D000939:Epitopes / Q000737:chemistry / Q000276:immunology; D005260:Female; D007782:Lactoglobulins / Q000737:chemistry* / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013045:Species Specificity
D000911:Antibodies, Monoclonal; D003911:Dextrans; D000939:Epitopes; D007782:Lactoglobulins; C014392:carboxymethyl dextran