MOLECULAR REQUIREMENTS FOR POLYAMINES BINDING TO THE ANTISPERMINE MONOCLONAL-ANTIBODY SPM8-2

The monoclonal antispermine antibody Spm8-2 was obtained by immunizing mice with a thyroglobulin-spermine conjugate. The molecular requireme nts for polyamines binding to this antibody were investigated by ELISA binding and inhibition tests, using a variety of natural polyamines a nd synthetic polyamine analogs, Four major structural determinants are important for the binding of polyamines by the antibody: (1) terminal amino groups: N-alkylation of both terminal amino groups of the polya mines leads to an important drop in the affinity for the antibody; (2) number of methylene groups spacing the amino groups: the four carbon chains appear to present the optimum length since the antibody binds p olyamines with repeats of the aminobutyl moiety more actively than the ir homologues with shorter or longer carbon chains; (3) number of amin o groups: the affinity of Spm8-2 for free homologous polyamines varied in the following order: pentamines > tetramines > triamines > diamine s, showing the importance of the number of positive charges of the pol yamines in the antibody-antigen reaction; the importance of charges is further emphasized by the dependence of antibody binding on the ionic strength of the medium; (4) N-acylation of one terminal amino group: the antibody binds more actively N-1-acetylspermidine than spermidine or spermine, The binding properties of Spm8-2 suggest the presence of two recognition sequences, one selective for N-acylaminopropyl moietie s, the second for the aminobutyl moiety.