Altering the antigenicity of proteins.
To better understand the binding interaction between antigen and antibody we need to distinguish protein residues critical to the binding energy and mechanism from residues merely localized in the interface. By analyzing the binding of monoclonal antibodies to recombinant wild-type and mutant myohemerythrin (MHr) proteins, we were able to test the role of individual critical residues at the highly antigenic site MHr-(79-84), within the context of the folded protein. The results directly show the existence of antigenically critical residues, whose mutations significantly reduce antibody binding to the folded protein, thus verifying peptide-based assignments of these critical residues and demonstrating the ability of buried side chains to influence antigenicity. Taken together, these results (i) distinguish the antigenic surface from the solvent-exposed protein surface before binding, (ii) support a two-stage interaction mechanism allowing inducible changes in protein antigens by antibody binding, and (iii) show that protein antigenicity can be significantly reduced by alteration of single critical residues without destroying biological activity.
Alexander
H
H
Division of Biological Sciences, University of Missouri, Columbia 65211.
Alexander
S
S
Getzoff
E D
ED
Tainer
J A
JA
Geysen
H M
HM
Lerner
R A
RA
D000595:Amino Acid Sequence; D000936:Antigen-Antibody Complex; D000941:Antigens / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D001666:Binding Sites, Antibody; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000737:chemistry* / Q000235:genetics / Q000276:immunology; D005813:Genes, Synthetic; D006422:Hemerythrin / Q000031:analogs & derivatives* / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016297:Mutagenesis, Site-Directed; D010449:Peptide Mapping; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000276:immunology; D015183:Restriction Mapping
D000936:Antigen-Antibody Complex; D000941:Antigens; D000939:Epitopes; D006422:Hemerythrin; D011994:Recombinant Proteins; C008571:myohemerythrin