468 AMINO ACID RESIDUES 93–96 OF THE HUMAN GROWTH HORMONE MOLECULE ARE IMPORTANT FOR POLYCLONAL ANTISERUM RECOGNITION

Abstract

The precise regions of the human growth hormone (hGH) molecule that mediate its biologic effects and immunoreactivity are not known. To address this issue, we have introduced mutations into the coding region of a cloned hGH gene and expressed the resulting mutants in a eukaryotic expression vector. Two clones with alterations in the DNA sequences encoding amino acids 93-96 within the hGH molecule were created by the insertion of Bam HI DNA linkers into the Mst II site. The hGH derived from these mutants had characteristics indistinguishable from those of wild type hGH in the NB-2 cell lactogenic assay, the IM-9 cell radioreceptor assay, and in the binding of four anti-hGH monoclonal antibodies. However, significant differences were observed between the mutants and standard human pituitary GH or hGH produced by the normal gene in the binding of polyclonal hGH antisera. Both mutants bound to the polyclonal antibody poorly but did not displace labeled standard hormone in a proportionate manner.These results suggest that amino acids 93-96 of hGH regulate the expression of antigenic determinants that are detected by polyclonal antiserum, but these determinants are distinct from those recognized by the four monoclonal antibodies that were tested. Moreover, hGH molecules with abnormal immunologic reactivity may retain normal biologic activities in certain assays.