N-terminal-truncated recombinant analogs of bovine placental lactogen: interaction with human and rat growth hormone receptors and insulin-like growth factor-I secretion mediated by somatogenic receptors in rat hepatocytes.
Bovine placental lactogen (bPL) was found to be as potent as human GH (hGH) in its ability to bind to soluble full-size recombinant hGH-binding protein (hGHBP) and to membrane-embedded hGH receptor in intact IM-9 human lymphocytes. bPL was also capable of forming a 1:2 complex with hGHBP, although the structure of this complex was probably more compact than that with hGH. Removal of 13 amino acids from the N-terminus of bPL did not affect its ability to bind to hGHBP or hGH receptors in intact IM-9 cells. Its ability to form a 1:2 complex with hGHBP was, however, impaired, unlike that of a corresponding analog in which an L28F mutation has been simultaneously introduced. Truncation of 17 amino acids decreased its affinity toward both hGHBP and GH receptors on intact IM-9 lymphocytes and in liver rat microsomal fraction and inhibited the formation of 1:2 complexes with hGHBP. Simultaneous L28F mutation did not affect affinity toward hGHBP, but increased affinity toward rat liver GH receptors and restored affinity toward membrane-embedded hGH receptors in IM-9 lymphocytes and the ability to form a 1:2 complex with hGHBP. Truncation of 20 amino acids further decreased affinity toward both hGHBP and receptors in intact IM-9 lymphocytes and completely abolished formation of a 1:2 complex with hGHBP. Both des-13-bPLs and bPL-des-17 (L28F) retained their full ability to stimulate insulin-like growth factor-I secretion by rat hepatocytes compared to that of bPL. The insulin-like growth factor-I stimulatory activities of bPL-des-17 and bPL-des-20, however, were decreased to 1-5%. These results indicate that the stoichiometry of 1:2 complex formation with hGHBP may be preserved despite decreased receptor binding affinity, but the lower affinity of the putative site 1 or site 2 of the analog may account for the decrease in biological activity. Furthermore, the ability or inability of bPL or its truncated analogs to form 1:2 complexes with soluble hGHBP cannot predict their somatogen receptor-mediated biological activity in rat hepatocytes.
Vashdi-Elberg
D
D
Department of Biochemistry, Food Science, and Nutrition, Faculty of Agriculture, Hebrew University of Jerusalem, Rehovot, Israel.
Staten
N R
NR
Sakal
E
E
Krivi
G G
GG
Gertler
A
A
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D013006:Growth Hormone / Q000378:metabolism; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000378:metabolism*; D008099:Liver / Q000166:cytology / Q000378:metabolism*; D008214:Lymphocytes / Q000378:metabolism; D010928:Placental Lactogen / Q000031:analogs & derivatives / Q000737:chemistry*; D051381:Rats; D011986:Receptors, Somatotropin / Q000378:metabolism*; D011994:Recombinant Proteins
D002352:Carrier Proteins; D011986:Receptors, Somatotropin; D011994:Recombinant Proteins; D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone; D010928:Placental Lactogen; C051815:somatotropin-binding protein