Allosteric effects of monoclonal antibodies on human growth hormone

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Abstract
We have previously shown that a monoclonal antibody (MAb) recognizing the human growth hormone (hGH) antigenic domain left exposed after binding to lactogenic receptors enhanced hGH binding probably through allosteric effects on the hormone binding site. Since receptors displaying different specificities would not recognize exactly the same hGH region, we explored whether some of our MAb could affect hGH binding to somatogenic receptors from rabbit liver and to human liver hGH-specific receptors. The effect of MAbAE5, AC8 and F11 on hGH binding was measured by determining the formation of125I-MAb:hGH:receptor complexes using two different experimental approaches. Results from procedure A, which involved the previous binding of the hormone to microsomes before adding125I-MAb, indicated that the hGH domain defined by epitopes AE5, AC8 and F11 is uncovered in the various hormone:receptor complexes. Procedure B was devised to reveal any alteration in the hGH molecule induced by the MAb. In this case preformed125I-MAb:hGH complexes were added to microsomes. Data showed that125I-MAb AE5:hGH complexes bound better to the various receptors than125I-MAb AE5 to hGH:receptor complexes. On the contrary, hGH previously bound to125I-MAb AC8 or125I-MAb F11 was less recognized by the receptors than the free hormone. Furthermore, binding of MAb AE5 or MAb F11 to hGH 20 K (a natural hGH variant lacking residues 32–46) also enhanced its affinity to the various receptors whereas MAb AC8 did not inhibit hGH 20 K binding. Results indicated that MAb recognizing the hGH antigenic area that remains unmasked after binding to different membrane-bound receptors are able to affect hormone binding site. MAb would induce either positive or negative allosteric changes in the hormone region involved in its binding to lactogenic, somatogenic and hGH-specific receptors.