Abstract
The specificity of goat antibodies to rabbit muscle AMP-deaminase was tested by immunoinhibition of enzymatic activity. Over most of the immunotitration curve, inhibition of the rabbit muscle enzyme was greater than that of the enzyme from chicken breast muscle; however, at high ratios of antiserum to enzyme, inhibition of the avian enzyme was virtually complete, while inhibition of the rabbit muscle deaminase was partially reversed. These observations were interpreted in terms of differential binding of activating and inhibiting antibodies. The role of lattice formation in inhibition of enzymatic activity by rabbit antibody was investigated by comparing the effect of intact antibody and univalent fragments, isolated from papain digests. Preparations of fragments Fab and Fab', as well as untreated antibody, completely inhibited the avian enzyme; however, the molar ratios of fragments Fab and Fab' to enzyme required to attain 50% inhibition were 2–8-fold greater than those required for untreated antibodies. In each case, fragment Fab was a more effective inhibitor of the avian deaminase than fragment Fab', thus, lattice formation contributes significantly to, but is not required for, immunoinhibition. Parallel studies of the effect of these untreated rabbit antibodies and soluble fragments on the enzymatic activity of rabbit muscle deaminase showed that enzyme inhibition by divalent antibody as always less than 100%, and in contrast with inhibition of the bird enzyme, fragment Fab' was consistently a more potent inhibitor of rabbit muscle AMP-deaminase than fragment Fab. AMP-deaminases from both rabbit skeletal muscle and chicken breast muscle were quantitatively precipitated by antibodies isolated from rabbits which were immunized with avian muscle deaminase. The degree of cross reaction of rabbit AMP-deaminase with these antibodies, relative to the amount of antibody precipitated at equivalence by the homologous avian enzyme (100%), ranged from 19–49%, depending on the particular antibody preparation. Direct binding studies, using Sepharose-conjugated rabbit antibody to chicken muscle enzyme, showed that the affinity of the rabbit enzyme for antibody was approximately 7-fold less than that of the avian muscle AMP-deaminase.
Published Version
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