ECE2008 Poster Presentations Growth and development (14 abstracts)
There is a high heterogeneity of human growth hormone (hGH), variation rising from the different genes in pituitary and placenta, alternative splicing, post-translational modifications, oligomerization and binding to the growth hormone binding protein (GHBP). Distinguishing between different variants remains problematic and there is still quite little information about the proportion of these variants in circulation of both healthy individuals and of patients with growth hormone disorders. Studying the abundance of dimer and oligomers in different physiological and pathological conditions is highly interesting since these forms of hGH will have longer half lives in circulation. In this experiment, 22 kDa hGH was expressed by HEK 293 cells and the hGH secreted in supernatant was studied. The western blot has shown, that in addition to monomer, hGH was also produced as dimer and oligomer. The dimer and oligomers are partially reduced by the reducing agent breaking down the disulfide bonds. Nevertheless, the dimer and oligomers can be detected even in reducing conditions, implicating that part of the dimer and oligomers are formed covalently otherwise than by disulfide bonding. A non-denaturing method based on fractionation with FPLC followed by high-sensitivity time-resolved immunofluorometric assays has turned out promising for the study of dimer and oligomer. The peaks measured by the immunoassays for monomeric versus dimeric and oligomeric hGH offered valuable information on the proportion of these forms. The function and structure of hGH dimer and oligomers as well as their relationship to hGH monomer need to be distinguished by further studies.