The effect of dodecyl sulfate on immunoglobulin hapten binding.
The instantaneous effect of dodecyl sulfate (DDS), in the mM concn range, on the binding of monovalent hapten by immunoglobulin was examined. Fluorescence measurements were utilized to study the effect of the detergent on sheep antiserum generated against thyroxin (T4) and against methamphetamine. Haptens were conjugated with the thiocyanate derivative of fluorescein in order to determine hapten binding on the basis of increased fluorescence polarization for the fluorescein-thiocarbamyl-hapten adducts (FT4 or FA) bound to immunoglobulin. Incubation of anti-T4-serum with DDS for 1 hr before the addition of FT4 resulted in diminished binding. The effect occurred at DDS concns greater than 0.1 mM and was essentially complete at a DDS conc of 1 mM. A kinetic study demonstrated a two stage process. An initial, rapid stage, with a half time less than 30 sec accounted for a reduction of immunoglobulin binding by 75%. The remaining 25% binding capacity was lost during a second, much slower phase with a half-time of about 11/2 hr. Prior hapten binding inhibited the effect of DDS. The degree of protection from combining site denaturation afforded by prior hapten binding was limited by the dissociation rate of bound hapten. The major, rapid phase was completely and immediately reversible by dilution. Prolonged incubation in DDS resulted in irreversible denaturation. The overall rate of DDS denaturation of the entire immunoglobulin molecule, as revealed by changes in the circular dichroism spectrum of a sheep gamma globulin fraction, was considerably slower than the denaturation rate of the combining site.
D000818:Animals; D004306:Dose-Response Relationship, Immunologic; D019793:Fluorescein; D005452:Fluoresceins; D006241:Haptens / Q000276:immunology*; D007106:Immune Sera / Q000276:immunology; D007136:Immunoglobulins / Q000276:immunology*; D008694:Methamphetamine / Q000276:immunology; D012756:Sheep; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology*; D013974:Thyroxine / Q000276:immunology; D013997:Time Factors
D005452:Fluoresceins; D006241:Haptens; D007106:Immune Sera; D007136:Immunoglobulins; D012967:Sodium Dodecyl Sulfate; D008694:Methamphetamine; C028913:dodecyl sulfate; D013974:Thyroxine; D019793:Fluorescein