Conformational changes induced by hapten in pig antibodies to the dinitrophenyl group. Analysis by temperature-perturbation and solvent-perturbation spectroscopy.
Pig antibodies to the dinitrophenyl group and fragments derived from them by limited proteolysis were studied by temperature-perturbation and solvent-perturbation spectroscopy with particular attention to differences between the number of perturbed chromophores in free antibodies and in antibody-hapten complexes. The position of the maxima in the difference spectra show that solvent-perturbed chromophores are exposed to water, but thermally perturbed chromophores are located in a microenvironment the polarity of which corresponds to 25-50% ethylene glycol. A significant fraction of tyrosine residues (65-90%) and tryptophan residues (20-45%) is perturbed by temperature. Much lower fractions, i.e. 35-45% of tyrosine residues and less than 15% of tryptophan residues, are perturbed by 20% glycerol. The numbers of perturbed chromophores in fragments constituting the molecule are lower than or equal to the numbers in the original molecule. The effect of hapten binding is significant only with one of the antibody types, the precipitating antibody. The number of thermally perturbed tyrosine residues is by about 17% lower in the liganded antibody. The absence of an analogous effect in the Fab fragment suggests that the fine conformational mechanism of signal transfer from the binding sites operates only in intact antibody molecules.
D000818:Animals; D000906:Antibodies / Q000276:immunology*; D004136:Dinitrobenzenes / Q000276:immunology*; D006241:Haptens / Q000276:immunology*; D009578:Nitrobenzenes / Q000276:immunology*; D011487:Protein Conformation / Q000187:drug effects*; D012997:Solvents; D013056:Spectrophotometry, Ultraviolet; D013552:Swine / Q000276:immunology*; D013696:Temperature; D014364:Tryptophan / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism
D000906:Antibodies; D004136:Dinitrobenzenes; D006241:Haptens; D009578:Nitrobenzenes; D012997:Solvents; D014443:Tyrosine; D014364:Tryptophan