LOCATION OF AN EPITOPE DEFINED BY AN ENHANCING MONOCLONAL-ANTIBODY TO GROWTH-HORMONE - SOME STRUCTURAL DETAILS AND BIOLOGICAL IMPLICATIONS

We have previously shown that a murine monoclonal antibody (MAb OA15) prepared against ovine GH (oGH) can enhance the somatogenic activity of bovine GH (bGH), as determined by increased incorporation of (SO42-)-S-35 into costal cartilage of hypopituitary Snell dwarf mice in vivo We have now established that MAb OA15 can also enhance the biological activity of oGH and porcine GH (pGH) in vivo. Using multiple pin peptide synthesis techniques a set of overlapping immobilized octamers representing the entire bGH sequence were synthesized and tested for their ability to bind MAb OA15 using an enzyme-linked immunosorbent assay, the pattern of binding showed that OA15 defined a functionally continuous epitope comprising residues 91-102. This region includes the C-terminal end of helix 2 plus a portion of the adjacent loop linking helices 2 and 3. Polyclonal antisera to a synthetic peptide representing this epitope mimicked the ability of MAb OA15 to enhance oGH, bGH, and pGH. Window size analysis showed that the heptapeptide 94-100 (SRVFTNS) represents the minimum unit to retain full recognition of MAb OA15. The fact that pGH, bGH, and oGH have identical sequences in this region also explains the ability of OA15 to both crossreact with and enhance the biological activity of each of these GHs. Replacement net analysis (where each residue in the heptamer is substituted with each of the 19 naturally occurring L-amino acids) demonstrated that residues R95 end T98 are critical for antibody binding and also indicated that the substitution of V96 with I, as found in rat GH, would permit the observed binding of OA15 to this hormone. Analogs of the epitope peptide with enhanced binding to OA15 were also revealed. Our data help to explain the observed pattern of cross-reactivity of OA15 with heterologous antigen and may be of assistance in the development of peptide vaccines for the endocrine manipulation of animal performance.