Expression and characterization of recombinant beta-subunit hCG homodimer.
We have linked two human chorionic gonadotropin (hCG) beta-subunit cDNAs in tandem such that the expressed fusion protein consists of two mature beta-subunits joined through the carboxy terminal peptide of the first beta-subunit. A single glycine residue is inserted between the two subunits in the fusion protein. Chinese hamster ovary (CHO) cells transformed with a clone that contains the fused cDNAs express and secrete a protein that is consistent with it being a beta-hCG homodimer protein. These beta-homodimer molecules can recombine with two free alpha-subunits indicating that both beta-subunits within the homodimer are likely folded in their native conformation. Our data also suggest that the two beta-subunits fold upon each other as a globular protein and do not appear to exist as a simple fusion of two linear beta-subunits. Furthermore, the two beta-monomer subunits in the fusion protein form a stable homodimer that can bind and activate the hLH/CG receptor specifically. Recombination of the fusion protein with alpha-subunits appears to favor an arrangement where two alpha-subunits combine with a single molecule of the fusion protein. The recombined molecule consists of four subunits and is comparable to two tethered hCG moieties, which constitutes a hCG dimer. This hormone dimer can bind and activate the hLH/CG receptor with an activity approximating that of native hCG.
Lobel
L
L
Department of Obstetrics and Gynecology, Center for Reproductive Sciences, Columbia University, New York, NY 10032, USA.
Pollak
S
S
Wang
S
S
Chaney
M
M
Lustbader
J W
JW
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D018997:Chorionic Gonadotropin, beta Subunit, Human / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006224:Cricetinae; D019281:Dimerization; D015870:Gene Expression; D015292:Glycoprotein Hormones, alpha Subunit / Q000737:chemistry; D006801:Humans; D008958:Models, Molecular; D011487:Protein Conformation; D011974:Receptors, LH / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D018997:Chorionic Gonadotropin, beta Subunit, Human; D015292:Glycoprotein Hormones, alpha Subunit; D011974:Receptors, LH; D011993:Recombinant Fusion Proteins