Mutations in the third alpha-helix of bovine growth hormone dramatically affect its intracellular distribution in vitro and growth enhancement in transgenic mice.
To investigate the relationship between the secondary structure of the third alpha-helix (amino acids 109-126) of bovine growth hormone (bGH) and the biological activity of the molecule, proline or glycine residues have been used as substitutes for native amino acids at positions 114, 118, 121, and 126, respectively. Mutations at the positions 114, 118, and 121 resulted in a dramatic decrease in bGH secretion by transiently transfected mouse L cells whereas the substitution of glycine for glutamate at position 126 (bGH-E126G) did not affect secretion. Immunofluorescence staining revealed that those nonsecretory bGH mutations possessed a different intracellular location as compared with wild-type bGH or the mutated secretory forms of bGH. Similar results were seen in the distribution of these mutated bGH molecules in transfected rat GH-3 cells. Transgenic mice that express wild-type bGH or bGH-E126G grew to approximately 1.6 times the mass of nontransgenic littermates. Transgenic mice that express two nonsecretory forms of mutated bGHs were found to lack the enhanced mouse growth phenotype in spite of elevated levels of serum bGH. These results suggest that the secondary structure in the third alpha-helix of bGH may be important for efficient intracellular targeting in vitro and in growth promotion in transgenic mice.
Chen
W Y
WY
Department of Zoology, Ohio University, Athens 45701.
Wight
D C
DC
Chen
N Y
NY
Coleman
T A
TA
Wagner
T E
TE
Kopchick
J J
JJ
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D005260:Female; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation; D013006:Growth Hormone / Q000235:genetics* / Q000378:metabolism; D007739:L Cells; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010641:Phenotype; D011487:Protein Conformation; D051381:Rats; D014162:Transfection
D013006:Growth Hormone