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Properties of univalent fragments of rabbit antibody isolated by specific adsorption

1960; Elsevier BV; Volume: 88; Issue: 2 Linguagem: Inglês

10.1016/0003-9861(60)90229-0

1096-0384

Alfred Nisonoff, F.C. Wissler, D.L. Woernley,

Polymer Surface Interaction Studies

A method is described for the isolation of active nonprecipitating fragments of rabbit antihapten antibody by specific adsorption. The fragments, about 13 as large as the original molecule, were isolated from a papain hydrolyzate of specifically purified anti-p-azobenzoate antibody. The average equilibrium constant for their interaction with homologous hapten, and the index of heterogeneity were the same, within experimental error, as the values for untreated antibody. Thus the isolation procedure does not fractionate molecules according to their combining affinity. It is also evident that 23 of the molecule may be discarded without affecting the structural integrity of the combining site and that electrostatic effects on binding due to the residual bulk of the molecule are negligible. The average number of combining sites per molecule of the isolated fragments, determined by hapten-binding measurements, is 0.9–1.0, providing direct evidence that the active fragments produced by papain are univalent. The specific optical rotation of a dialyzed papain hydrolyzate is approximately the same as that of untreated rabbit γ-globulin, indicating that the fragments are compact molecular units; i.e., are not appreciably unfolded or denatured. This conclusion is supported by the reduction in specific viscosity and the unchanged heat stability which accompanies the hydrolysis.