Expression of two forms of carp gonadotropin alpha subunit in insect cells by recombinant baculovirus.
There are two types of cDNA clones (designated alpha 1 and alpha 2) encoding the alpha subunit of carp gonadotropin. These two cDNAs are derived from different genes and encode proteins that differ by seven amino acid residues (three in the signal peptide and four in the mature polypeptide). Expression of these two cDNAs in insect cells by recombinant baculovirus revealed that the alpha 1 subunit, after noncovalent association with the beta subunit, has the same potency as the native alpha subunit purified from the pituitary. In contrast, the alpha 2 subunit can associate with the beta subunit, but only to form an inactive gonadotropin. Competition of the alpha 2 subunit with the alpha 1 subunit for association with the beta subunit decreases the gonadotropin activity of the alpha/beta complex. In addition, both alpha 1 and alpha 2 subunits are secreted into the culture medium by insect cells and have an apparent molecular mass approximately 5 kDa higher than that of the native alpha subunit. These results indicate that the insect cell-derived alpha 1 subunit is biologically active and that those four amino acid changes in the mature of alpha 2 protein affect the biological activity and thus provide valuable clues for the study of the structure-function relationship of the alpha subunit of glycoprotein hormones.
Huang
C J
CJ
Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan.
Huang
F L
FL
Chang
G D
GD
Chang
Y S
YS
Lo
C F
CF
Fraser
M J
MJ
Lo
T B
TB
D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D001681:Biological Assay; D015153:Blotting, Western; D002347:Carps; D002460:Cell Line; D015870:Gene Expression; D005822:Genetic Vectors; D015292:Glycoprotein Hormones, alpha Subunit / Q000235:genetics* / Q000302:isolation & purification / Q000494:pharmacology; D007313:Insecta; D007700:Kinetics; D008297:Male; D011863:Radioimmunoassay; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000494:pharmacology; D015183:Restriction Mapping; D013737:Testis / Q000187:drug effects / Q000378:metabolism; D013739:Testosterone / Q000096:biosynthesis; D014162:Transfection
D015292:Glycoprotein Hormones, alpha Subunit; D011994:Recombinant Proteins; D013739:Testosterone