Evidence that the N-terminus of human growth hormone is involved in expression of its growth promoting, diabetogenic, and insulin-like activities.
Recent work with various point and deletion mutants of human GH (hGH) has suggested that the proximal N-terminal end of the hormone molecule is important for its growth promoting action. This study was conducted to examine the growth promoting, diabetogenic, and insulin-like activities of two N-terminal mutants of hGH, the deletion mutant Des-7 hGH (met8, ala11), and a chimeric mutant of bovine GH (bGH) and hGH containing the N-terminal 13 amino acids of bGH (met, ala 1-13/14-191, asp11). The CD spectra of these mutants are similar to that of wild-type hGH and they retain lactogenic activity on Nb2 lymphoma cells, whereas their ability to bind to somatogenic receptors on IM-9 lymphocytes and bovine liver membranes is markedly reduced. In this study, growth promoting activity of the mutants was assessed using the 9-day weight gain test in hypophysectomized rats. Des-7 hGH had a potency of 0.03 IU/mg protein in this assay, whereas the potency of the bGH/hGH chimera was 0.71 IU/mg. Diabetogenic activity was tested in the ob/ob mouse, using the elevation of fasting blood glucose and the worsening of glucose tolerance after a 3-day course of treatment as end-points. Both Des-7 hGH and the bGH/hGH chimera had reduced diabetogenic activity compared to that of biosynthetic wild-type hGH, consistent with their reduced growth activity. Insulin-like activity was assessed by testing the in vitro ability of the mutants to stimulate [14C] glucose oxidation by epididymal adipose tissue of hypophysectomized rats. Des-7 hGH had about 1% the activity of wild-type hGH, whereas the chimera was about 20% as active. When Des-7 hGH was added to the incubation medium along with wild-type hGH in ratios of 5, 12.5, or 25:1 (Des-7 hGH:hGH), the insulin-like action of hGH was significantly inhibited, indicating that the mutant is a modest antagonist of the insulin-like action of hGH. When the ability of Des-7 hGH to compete with [125I] hGH for binding to isolated rat adipocytes was tested, the mutant was about 10% as effective as wild-type hGH. Thus, Des-7 hGH appears to be more effective in binding to adipocyte GH receptors than in triggering an insulin-like response, perhaps accounting for its modest antagonistic activity. The results of this study suggest that the proximal N-terminal end of the hGH molecule is involved in the expression of the growth promoting, diabetogenic and insulin-like activities of GH.
Towns
R
R
Department of Physiology, University of Michigan Medical School, Ann Arbor 48109.
Kostyo
J L
JL
Vogel
T
T
Sakal
E
E
Tchelet
A
A
Maher
R
R
Gertler
A
A
D000273:Adipose Tissue / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D000818:Animals; D001786:Blood Glucose / Q000378:metabolism*; D005260:Female; D013006:Growth Hormone / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D007328:Insulin / Q000502:physiology*; D008297:Male; D051379:Mice; D008817:Mice, Mutant Strains; D008820:Mice, Obese; D009154:Mutation / Q000235:genetics; D011487:Protein Conformation; D051381:Rats; D011986:Receptors, Somatotropin / Q000187:drug effects / Q000502:physiology; D015430:Weight Gain / Q000187:drug effects*
D001786:Blood Glucose; D007328:Insulin; D011986:Receptors, Somatotropin; D013006:Growth Hormone