NMR technique for assessing contributions of heavy and light chains to an antibody combining site.
Nuclear magnetic resonance (NMR) has been used extensively to study the structure of antibody combining sites. In recent studies we have observed the proton resonance spectra of the Fab fragment of a monoclonal anti-spin-label antibody derived from a hybridoma grown on various specifically deuterated amino acids. The broadening of the proton resonance signals by the paramagnetic hapten, together with selective deuteration, has allowed the identification of most of the amino acids in the combining-site region of this antibody and has also provided estimates of distances between amino-acid protons and the unpaired electron. Here we show how recombination of specifically deuterated heavy and light chains permits the assignment of single amino-acid proton resonance signals to either the heavy or light chain. In addition, the spectra of such recombinants demonstrate that their combining-site structures must be almost identical to the native structure.
D000818:Animals; D000911:Antibodies, Monoclonal; D001666:Binding Sites, Antibody; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D009682:Magnetic Resonance Spectroscopy; D051379:Mice; D013113:Spin Labels / Q000276:immunology; D013329:Structure-Activity Relationship; D014443:Tyrosine
D000911:Antibodies, Monoclonal; D007074:Immunoglobulin G; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D013113:Spin Labels; D014443:Tyrosine