Biological activities of recombinant equine luteinizing hormone/chorionic gonadotropin (eLH/CG) expressed in Sf9 and Mimic insect cell lines.
Equine luteinizing hormone (eLH) and chorionic gonadotropin (eCG) are composed of identical alpha and beta polypeptide chains, but eCG subunits are much more heavily glycosylated and sialylated. Consequently, eCG exhibits a much longer half-life than eLH in blood. Recombinant eLH/CG, expressed in Sf9 and Mimic insect cells, were compared with one another and to the natural hormones eCG and eLH. Mimic cells are stably-transformed Sf9 cells, expressing five mammalian genes encoding glycosyltransferases involved in the synthesis of complex N-carbohydrate chains. Recombinant eLH/CG expressed in Mimic cells exhibited a higher apparent molecular weight (MW) than that expressed in Sf9 cells, suggesting that its N-glycosylation was, as expected, more complete. Nevertheless, the two recombinant eLH/CG exhibited lower MW than natural eCG from pregnant mare plasma. The two eLH/CG produced in Sf9 and Mimic cells were found to be active in in vitro LH and FSH bioassays, with potencies similar to those of eCG. By contrast, they exhibited no significant in vivo bioactivity, neither in the specific follicle-stimulating hormone (FSH) assay nor in the specific eCG assay. Although recombinant eLH/CG produced in Mimic cells bears more elaborate carbohydrate chains than recombinant eLH/CG from Sf9 cells, it exhibits no significant in vivo bioactivity, probably because of insufficient terminal sialylation of its carbohydrate chains, leading to its rapid removal from blood.
Legardinier
Sébastien
S
Unité de Physiologie de la Reproduction et des Comportements, Institut National de la Recherche Agronomique (INRA), Nouzilly, France. legardin@tours.inra.fr
Duonor-Cérutti
Martine
M
Devauchelle
Gérard
G
Combarnous
Yves
Y
Cahoreau
Claire
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001681:Biological Assay; D015153:Blotting, Western; D019556:COS Cells; D002522:Chlorocebus aethiops; D006063:Chorionic Gonadotropin / Q000235:genetics / Q000378:metabolism*; D005260:Female; D006736:Horses / Q000378:metabolism*; D007986:Luteinizing Hormone / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D018411:Spodoptera
D006063:Chorionic Gonadotropin; D011994:Recombinant Proteins; D007986:Luteinizing Hormone