The α-subunit of human choriogonadotropin interacts with the exodomain of the luteinizing hormone choriogonadotropin receptor

The LH/CG receptor, a G protein-coupled receptor, consists of two parts, the N-terminal extracellular segment (exodomain) and the membrane-associated C-terminal segment (endodomain). hCG initially binds the exodomain of the receptor and then, the hormone/ exodomain complex is thought to make the secondary contact with the endodomain of the receptor and generate a hormone signal. However, little direct evidence is available about which hormone subunits (alpha or beta) interact with which domains of the receptor. To determine whether the alpha-subunit contacts the exodomain of its receptor, hCG containing [I-125]alpha and truncated exodomain lacking the endodomain were prepared. They were chemically cross-linked, and the resulting cross-linked complexes were solubilized and electrophoresed. The results indicate that the alpha-subunit of hCG was directly and specifically cross-linked to the exodomain. To verify the cross-linked exodomain by the independent method, the Flag epitope was inserted between the signal sequence and the mature exodomain, hCG containing [I-126]alpha was cross-linked to the Flag exodomain, and the resulting cross-linked hCG/Flag exodomain complexes were immunoprecipitated with anti-Flag antibody. The results show that the material cross-linked to hCG containing [I-125]alpha is indeed the exodomain. In conclusion, our results show the direct interaction of the alpha-subunit with the exodomain and, therefore, its crucial role in the hormone-receptor interaction in addition to its involvement in signal generation.