Biological characterization of purified native 20-kDa human growth hormone

Abstract

Because of the propensity of the 20-kDa variant of human growth hormone (GH) to aggregate with itself and with 22-kDa human GH, it has been difficult to prepare monomeric 20-kDa GH in highly purified form. This has been a major complicating factor in determining whether 20-kDa GH has a biological activity profile distinct from that of 22-kDa GH. In the present study, native 20-kDa GH was isolated from a human GH dimer concentrate and purified by a procedure that included column electrophoresis in agarose suspension as a final separation step. This procedure yielded highly purified monomeric 20-kDa GH, which was contaminated to an extent of less than 1% with 22-kDa GH, and which exhibited only a small degree of dimerization upon storage. The native 20-kDa GH was quite active in stimulating growth in hypophysectomized rats, when growth was assessed by body weight gain, longitudinal bone growth, the stimulation of sulfation of cartilage, and the elevation of serum IGF-1 level. However, in all of these growth assays, the 20-kDa GH was somewhat less active than the native 22-kDa GH to which it was compared; e.g., in the body weight gain and longitudinal bone growth assays, it had an estimated potency of 0.6 relative to the 22-kDa GH. The 20-kDa GH exhibited substantial diabetogenic activity when tested for the ability to raise fasting blood glucose concentration and to impair glucose tolerance in ob/ob mice. Also, the native 20-kDa GH had significant in vitro insulin-like activity, although its potency was approximately 20% that of the native 22-kDa GH to which it was compared. Thus, the biological activity profile of native 20-kDa GH differs from that of 22-kDa GH primarily in that insulin-like activity is markedly attenuated.

Optimizing conditions for production of high levels of soluble recombinant human growth hormone using Taguchi method

“…It was used as a therapeutic agent against growth-retarded GH deficiency (GHD), Turner syndrome and End-stage renal disease in children and also used against SHOX (short stature homeobox gene) deficiency and AIDS-associated wasting in adults [4,11,13]. Using of pituitary-derived hGH has been prohibited when its association with Creutzfeldt-Jakob disease was proved [2].…”

22-kD Growth hormone exclusion assay: a new approach to measurement of non-22-kD growth hormone isoforms in human blood

“…In serum collected during spontaneous GH secretion, approximately 12% of total GH was non-22 GH isoforms, but the percentage ranged from 7.2% to19.4% in individual samples. Investigations using immunoaffinity extraction combined with polyacrylamide gel electrophoresis (PAGE) and RIA or immunoprecipitation followed by PAGE with immunoblotting have shown that among the monomeric variants of GH, 22 GH corresponds to 76-85% of plasma GH, 20 GH to 7-16% and acidic isoforms to 5-10%…”

Inhibition of growth hormone bioactivity by recombinant human growth hormone-binding protein in the eluted stain assay system

“…This preparation exhibits a variable degree of contamination with 22 kDa hGH, which can complicate the interpretation of GHBP studies (Baumann & Shaw 1990a). In rats, 20 kDa hGH has been reported to have full growth-promoting activity and somatomedin-generating potency in vivo when com¬ pared with 22 kDa hGH (Spencer et al 1981, Mosier & Lewis 1982, Closset et al 1983) but reduced diabetogenic activity (Kostyo et al 1986). This demonstrated that the variants may differ in their biological activity profiles.…”