Interaction of monoclonal anti-peptide antibodies with lysozyme.
The interaction of monoclonal anti-peptide antibodies with the free peptide and its protein counterpart has been evaluated for hen egg white lysozyme and the peptide constituting residues 38 to 45. Fluorescence methodology has been developed for the measurement of association constants based on resonance energy transfer between the excited tryptophan of antibody and bound peptide ligand conjugated to a fluorescent probe. Five antibodies, four IgM and one IgG, have been assayed by ELISA, and have demonstrated binding to the adsorbed peptide alone, to the adsorbed lysozyme alone, or to both. Multivalent interaction with the adsorbed ligand is a key factor in the efficacy of binding. Measurement of binding constants in homogeneous solution, by equilibrium dialysis and energy transfer, demonstrated that lysozyme was bound to an IgG antipeptide antibody with an association constant (4 X 10(2) M-1) 200-fold less than that for the free peptide (8 X 10(4) M-1). It was also inferred for IgM that an association constant of the order of 10(2) M-1 was sufficient to effect selective interaction in a system providing multivalent interaction. The shared conformations between protein and peptide, implied by the specific reactivity of the anti-peptide antibody with the protein, points to structural fluctuations of the surface regions and residues of globular proteins.
Hirayama
A
A
Takagaki
Y
Y
Karush
F
F
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000937:Antigen-Antibody Reactions; D001666:Binding Sites, Antibody; D001667:Binding, Competitive; D002645:Chickens / Q000276:immunology; D002999:Clone Cells / Q000276:immunology / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000276:immunology; D005455:Fluorescent Antibody Technique; D007075:Immunoglobulin M / Q000502:physiology; D051379:Mice; D008807:Mice, Inbred BALB C; D009113:Muramidase / Q000276:immunology*; D010047:Ovalbumin / Q000276:immunology*; D010455:Peptides / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D007075:Immunoglobulin M; D010455:Peptides; D010047:Ovalbumin; D009113:Muramidase