Characterization of glycosylated bovine placental lactogen and the effect of enzymatic deglycosylation on receptor binding and biological activity.
Bovine placental lactogen (bPL) is a glycoprotein hormone that has both somatogenic and lactogenic properties. Purified preparations of the hormone contain many isoforms that are separated by isoelectric focusing. The sequence for bPL contains one consensus site for an N-linked oligosaccharide and many potential sites for O-linked sugars. To determine whether the isoforms are the result of differences in glycosylation, the oligosaccharide portion of bPL was partially characterized. In addition, a number of the isoforms were isolated and enzymatically deglycosylated to determine the effect of O- and N-linked glycosylation on biological activity. Biological activity was assessed in a somatotropin radioreceptor assay and also in the Nb2 lymphoma lactogenic bioassay. The structure of N-linked oligosaccharide was found to be sialylated and triantennary and appeared to be the same for all of the different charge isomers. Compositional analysis suggested that O-linked oligosaccharides were also present. Treatment of the intact hormone with neuraminidase resulted in the loss of some, but not all, of the isoforms, suggesting that a large degree of the charge heterogeneity is due to posttranslational modifications unrelated to glycosylation. Enzymatic removal of N-linked oligosaccharides from native bPL resulted in a 1.2-2.3-fold increase in binding to the somatotropin receptor, whereas receptor binding was unaffected by enzymatic removal of O-linked oligosaccharide. Lactogenic activity was affected very little by the removal of either type of oligosaccharide. The data suggests that glycosylation of bPL may have a small effect on receptor specificity, but that overall its presence does not dramatically affect receptor binding or biological activity.
Byatt
J C
JC
Animal Sciences Division, Monsanto Company, St. Louis, Missouri 63198.
Welply
J K
JK
Leimgruber
R M
RM
Collier
R J
RJ
D000818:Animals; D001681:Biological Assay; D002236:Carbohydrate Conformation; D002417:Cattle; D002455:Cell Division / Q000187:drug effects; D005260:Female; D006026:Glycoside Hydrolases / Q000378:metabolism; D006031:Glycosylation; D007526:Isoelectric Point; D008223:Lymphoma; D008970:Molecular Weight; D009005:Monosaccharides / Q000032:analysis; D009844:Oligosaccharides / Q000032:analysis; D010928:Placental Lactogen / Q000032:analysis / Q000378:metabolism* / Q000494:pharmacology; D011869:Radioligand Assay; D011956:Receptors, Cell Surface / Q000378:metabolism*; D018000:Receptors, Peptide; D013329:Structure-Activity Relationship; D014407:Tumor Cells, Cultured
D009005:Monosaccharides; D009844:Oligosaccharides; D011956:Receptors, Cell Surface; D018000:Receptors, Peptide; C028202:placental lactogen receptor; D010928:Placental Lactogen; D006026:Glycoside Hydrolases