Hapten-induced changes in pig anti-dansyl antibodies revealed by EPR spectra of spin-labelled antibodies.
Pig anti-Dns antibodies were labelled by 2, 2, 6, 6- tetramethyl-N1-oxylpiperidine-4-amino (N-dichlorotriazine) (label SL I) or by 4-amino-2,2,6,6-tetramethyl-piperidine-N1-oxyl (label SL II). SL I presumably reacted with Fab parts, whereas SL II was attached to the carbohydrate moiety of the Fc part of the antibody molecule. The new method of intramolecular mobility estimation based on viscosity-dependent correlation times of the N-O group of spin-labelled proteins was used, making it possible to separate the correlation times of the spin-label relative to the protein carrier (tauR) and of the labelled protein itself (tauM). The altered shape of the EPR spectrum observed upon binding of epsilon-Dns-lysine indicates that the relative fluctuations of the liganded antibody domains within the Fab part are reduced. The character of temperature dependence of the correlation times of the labelled antibody changed upon hapten binding as well, suggesting an altered interaction between antibody subunits. Evidence of hapten-induced changes of both tauR and tauM was obtained at 1 degrees C and 5 degree C using the label SL II bound to the Fc part. The character of the changes can be interpreted in terms of mobilization of the domains and of altered interaction between labelled carbohydrate and the protein moiety. At 5 degree C the addition of 3% of D2O to the antibody labelled by SL II brought about a substantial effect qualitatively similar to that induced by the hapten.
D000818:Animals; D000906:Antibodies; D002417:Cattle; D003619:Dansyl Compounds / Q000276:immunology*; D004578:Electron Spin Resonance Spectroscopy; D006241:Haptens; D011487:Protein Conformation; D013113:Spin Labels; D013552:Swine
D000906:Antibodies; D003619:Dansyl Compounds; D006241:Haptens; D013113:Spin Labels