CROSS-REACTIVITY OF ANTIBODIES TO RETRO-INVERSO PEPTIDOMIMETICS WITH THE PARENT PROTEIN HISTONE H3 AND CHROMATIN CORE PARTICLE - SPECIFICITY AND KINETIC RATE-CONSTANT MEASUREMENTS

A series of monoclonal antibodies has been generated against an hexape ptide of sequence IRGERA corresponding to the C-terminal residues 130- 135 of histone H3 and three analogues of this model peptide. The analo gues correspond to the D-enantiomer, containing only D-residues, and t wo retro-peptides containing NH-CO bonds instead of natural amide pept ide bonds. The chirality of each residue was maintained in the retrope ptide and inverted in the retro-inverso-peptide. Monoclonal antibodies were generated from mice immunized with the analogues coupled to neut ral small unilamellar liposomes containing monophosphoryl lipid A as a djuvant. The reactivity of antibodies with the four analogues and with the parent protein H3 was studied in enzyme-linked immunosorbent assa y and in a biosensor system. The equilibrium affinity constant (K(a)lp ha) toward the retro-inverso-peptide of two out of three antibodies of IgG1 isotype induced against the L-hexapeptide was 7-75-fold higher t han toward the homologous L-peptide. The range of K-alpha, values of f our antibodies of IgG1 and IgG2a isotypes generated against the retro inverso-peptide was 0.6-1.9 x 10(9) M(-1) for both the retro-inverso- and L-peptides. Furthermore, antibodies to the L- and retroinverso-pep tides cross reacted strongly (in some cases better than with the homol ogous peptide) with the parent histone H3 and with chromatin subunits containing H3. The results are thus promising in respect to the potent ial use of retro inverso-analogues, which are particularly stable, in the design of much more potent synthetic vaccines or to generate antib ody probes.