Amino acid residues in the third alpha-helix of growth hormone involved in growth promoting activity.
The third amphiphilic alpha-helix of GH has been found to be an important motif in the biological activities of the molecule. To further characterize this growth-promoting domain, three bovine (b) GH analogs were designed: one contained a scrambled third amphiphilic alpha-helix (SAH); a second contained a scrambled hydrophilic region of the helix (SAP); and a third contained a scrambled hydrophobic region of the helix (SAB). Transgenic mice that expressed these mutated bGH genes were produced. SAH transgenic mice displayed a phenotype identical to nontransgenic littermates. SAB transgenic mice grew slightly larger than nontransgenic littermates but remained smaller than bGH transgenic mice. On the other hand, SAP transgenic mice exhibited a dwarf phenotype. We subsequently generated individual amino acid substitutions in the hydrophilic region of the helix. The results from the growth rates of corresponding transgenic mice demonstrated that most bGH analogs with individual amino acid substitution within the third alpha-helix retained wild type-like growth-promoting activity except those with alterations at positions 115, 119, 122, and 123. Together these residues are predicted to form a cleft in the helix. To further substantiate the importance of the cleft, we deleted Gly 119 (delta 119). This resulting bGH analog was inactive in vivo as well as in in vitro assays. These results indicated that the primary structure of the third alpha-helix is critical for GH's growth-promoting activity and Gly 119 is a crucial amino acid in this region. Three adjacent amino acids, Asp 115, Ala 122, and Leu 123, also contribute to the growth-enhancing ability of the molecule.
Chen
W Y
WY
Edison Biotechnology Institute, Department of Biological Sciences, Ohio University, Athens 45701, USA.
Chen
N Y
NY
Yun
J
J
Wight
D C
DC
Wang
X Z
XZ
Wagner
T E
TE
Kopchick
J J
JJ
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D013006:Growth Hormone / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000378:metabolism; D007739:L Cells; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D016296:Mutagenesis; D019000:Phosphotyrosine; D017433:Protein Structure, Secondary; D011869:Radioligand Assay; D011994:Recombinant Proteins / Q000737:chemistry / Q000494:pharmacology; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D011994:Recombinant Proteins; D019000:Phosphotyrosine; D014443:Tyrosine; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone