Monoclonal antipeptide antibodies: affinity and kinetic rate constants measured for the peptide and the cognate protein using a biosensor technology.
The interaction of antipeptide antibodies with the corresponding peptide and the cognate protein has been compared using a novel biosensor technology (BIAcore, Pharmacia). The peptide corresponds to residues 110-135 of the coat protein of tobacco mosaic virus, known to encompass an alpha-helical region reactive with antiprotein antibodies. A panel of 33 monoclonal antibodies raised against the peptide was studied and the epitope recognized by these antibodies was determined by the pepscan method. Further discrimination between the antibodies was performed by measurements of association and dissociation kinetic constants. Several antibodies showed an heterogeneous binding profile when reacting with the 25 residue long peptide but not with a shorter 10 residue peptide suggesting that they recognized different conformational states in the longer peptide. Equilibrium affinity constants were calculated for five of the antibodies and were found to be 10-50 times higher for the peptide than for the protein, the difference being caused mainly by a lower association rate constant.
Zeder-Lutz
G
G
Laboratoire d'Immunochimie, Institut de Biologie Moléculaire et Cellulaire, Strasbourg, France.
Altschuh
D
D
Geysen
H M
HM
Trifilieff
E
E
Sommermeyer
G
G
Van Regenmortel
M H
MH
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D000915:Antibody Affinity; D000916:Antibody Diversity; D001666:Binding Sites, Antibody; D015374:Biosensing Techniques; D000939:Epitopes / Q000737:chemistry; D007700:Kinetics; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology; D008969:Molecular Sequence Data; D010455:Peptides / Q000276:immunology*; D011487:Protein Conformation; D014027:Tobacco Mosaic Virus / Q000276:immunology; D014764:Viral Proteins / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D010455:Peptides; D014764:Viral Proteins