Antibodies specific for the antigenic domain 1 of glycoprotein B (gpUL55) of human cytomegalovirus bind to different substructures.
Glycoprotein B (gB, gpUL55) is a major antigen for the induction of neutralizing antibodies against human cytomegalovirus, making it an attractive antigen for active and passive immunoprophylaxis. The immunodominant region on gB is the antigenic domain 1 (AD-1), a complex structure which requires a minimal linear amino acid sequence of more than 75 amino acids (aa 552-635) for antibody binding. We have analyzed the fine specificity of neutralizing and nonneutralizing AD-1-binding monoclonal antibodies. Point mutations were introduced into AD-1 and mutants were expressed as bacterial fusion proteins. The antigens were analyzed in immunoblots using a panel of 13 human and murine monoclonal antibodies. Complete loss of binding of all antibodies was observed with mutations at cysteine residues 573 and 610 as well as with a combinatorial exchange of prolines at position 577 and 613. The remaining mutations had different effects on antibody binding. Six individual recognition patterns were observed, indicating various antigenic substructures on AD-1. Changing the Fc portions of 3 murine monoclonal antibodies to human IgG1 showed that neutralization of AD-1-binding immunoglobulins is exerted by different mechanisms. Dependent on the recognized substructure within AD-1, avidity-dependent as well as Fc portion-mediated effects were observed.
Schoppel
K
K
Institut für Klinische und Molekulare Virologie, Universität Erlangen-Nürnberg, Germany.
Hassfurther
E
E
Britt
W
W
Ohlin
M
M
Borrebaeck
C A
CA
Mach
M
M
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000914:Antibodies, Viral / Q000276:immunology*; D000918:Antibody Specificity; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology*; D001483:Base Sequence; D002478:Cells, Cultured; D003587:Cytomegalovirus / Q000276:immunology*; D017931:DNA Primers; D018985:Epitopes, B-Lymphocyte / Q000276:immunology; D006801:Humans; D016056:Immunodominant Epitopes / Q000276:immunology; D007141:Immunoglobulin Fc Fragments / Q000276:immunology; D007128:Immunoglobulin Fragments / Q000276:immunology; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009500:Neutralization Tests; D011993:Recombinant Fusion Proteins / Q000276:immunology; D014407:Tumor Cells, Cultured; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D017931:DNA Primers; D018985:Epitopes, B-Lymphocyte; D016056:Immunodominant Epitopes; D007141:Immunoglobulin Fc Fragments; D007128:Immunoglobulin Fragments; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; C040495:glycoprotein B, Simplexvirus; C055966:immunoglobulin Fv