Site-directed mutations of human growth hormone that selectively modify its lactogenic activity and binding properties.
Two novel analogs of human (h) GH, 1) Des-7-hGH (Arg8Met, Asp11Ala) in which the Arg8 was substituted by Met and Asp11 by Ala, and 2) bovine (b) GH/hGH hybrid II (MetAla 1-13/14-191, Ala11Asp) composed of 13 N-terminal amino acid of bGH and elongated by two amino acids (Met-Ala-1-13) and 14-191 amino acids of hGH, were constructed and expressed in Escherichia coli. CD spectra indicated that the alpha-helix content of the purified proteins was similar to that of the native hormone. Both analogs retained their full ability to stimulate the proliferation of Nb2 lymphoma cells, and their binding to the lactogen receptors in homogenate of Nb2 cells and in microsomal fraction from bovine lactating mammary gland was only slightly reduced. However, their ability to bind to the somatogen receptors in intact IM-9 lymphocytes and bovine liver was reduced by 7- to 11-fold (bGH/hGH hybrid II) and 20- to 30-fold (Des-7-hGH). Both analogs were able to down-regulate the respective lactogen and somatogen receptors in intact Nb2 and IM-9 cells. The galactopoietic activity of both analogs in the lactating bovine mammary explants bioassay was almost completely abolished, and the bGH/hGH hybrid II exhibited a remarkable antagonistic activity. These results further indicate that the lactogen receptors in different species or organs are not identical. We have shown that the new recombinant analogs of hGH that recognize both somatogen and lactogen receptors but have modified postreceptor effects are helpful in elucidating these differences.
Binder
L
L
Department of Biochemistry and Human Nutrition, Faculty of Agriculture, Hebrew University of Jerusalem, Rehovot, Israel.
Gertler
A
A
Elberg
G
G
Guy
R
R
Vogel
T
T
D001483:Base Sequence; D002478:Cells, Cultured; D002942:Circular Dichroism; D003001:Cloning, Molecular; D015536:Down-Regulation; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010928:Placental Lactogen / Q000235:genetics* / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism*; D018000:Receptors, Peptide
D011956:Receptors, Cell Surface; D018000:Receptors, Peptide; C028202:placental lactogen receptor; D010928:Placental Lactogen