Over-expression and characterization of recombinant beta subunit of the human chorionic gonadotropin hormone synthesized in insect cells infected with a genetically engineered baculovirus.
A recombinant baculovirus, vAc beta hCG, having a replacement of the viral polyhedrin gene with the cDNA encoding the beta subunit of hCG was used to express beta hCG, an extensively glycosylated hormone, in insect cells. Virus-infected cells, 72 hr pi, secreted approximately 8.02 micrograms beta hCG/2 x 10(6) cells/ml. The recombinant beta hCG purified from insect cells exhibited increased mobility on SDS-PAGE as compared to authentic urinary beta hCG, a reflection on differences in glycosylation between insect and mammalian systems. The insect derived beta hCG, however, was identical to the native hormonal peptide in terms of immunoreactivity and bioactivity on association with alpha-subunit, as evident by its binding to rat testicular receptors and induction of steroidogenesis in a mouse Leydig cell bioassay system. The implications of using the baculovirus system to study the importance of carbohydrates for biological activity are also discussed.
Nakhai
B
B
National Institute of Immunology, Shahid Jeet Singh Marg, New Delhi, India.
Sridhar
P
P
Pal
R
R
Talwar
G P
GP
Hasnain
S E
SE
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D015153:Blotting, Western; D006063:Chorionic Gonadotropin / Q000096:biosynthesis* / Q000302:isolation & purification; D018997:Chorionic Gonadotropin, beta Subunit, Human; D004247:DNA / Q000032:analysis; D015870:Gene Expression / Q000235:genetics*; D006801:Humans; D007313:Insecta / Q000235:genetics*; D010446:Peptide Fragments / Q000096:biosynthesis* / Q000302:isolation & purification; D010957:Plasmids; D011506:Proteins / Q000032:analysis; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection
D006063:Chorionic Gonadotropin; D018997:Chorionic Gonadotropin, beta Subunit, Human; D010446:Peptide Fragments; D011506:Proteins; D011994:Recombinant Proteins; D004247:DNA