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A fluorimetric study of the interactions of insolubilized human α-lactalbumin with galactosyl transferase (A-protein) and with anti-α-lactalbumin antibodies

par Prieels, Jean-Paul ;Barel, André
Référence BBA - Protein Structure, 393, 2, page (496-504)
Publication Publié, 1975-06

Article révisé par les pairs

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Résumé :

Intrinsic as well as extrinsic fluorescence of an immobilized protein was used for the study of the interactions between α-lactalbumin-Sepharose and protein ligands. The fluorescence peak of the human α-lactalbumin-agarose conjugate was shifted to the blue and quenched in the presence of the galactosyl transferase (A-protein), indicating the probable formation of a complex between both proteins. The natural fluorescence of human α-lactalbumin bound to Sepharose was specifically quenched in presence of antihuman α-lactalbumin antibodies. This change in fluorescence appears to be due to binding of the antibodies to the immobilized antigen. Furthermore, the extrinsic fluorescence of a bound dye such as 2-p-toluidinylnaphthalene-6-sulfonate was used to confirm the existence of binding between antibodies and α-lactalbumin-agarose, and to obtain values for the association constant. A value of 5.6·10+6 M-1 for the binding constant was reported, which compares favorably with other data obtained by equilibrium dialysis. © 1975.