The groove between the α- and β-subunits of hormones with lutropin (LH) activity appears to contact the LH receptor, and its conformation is changed during hormone binding

Abstract

Gonadotropins are heterodimeric glycoprotein hormones that control vertebrate fertility through their actions on gonadal lutropin (luteinizing hormone, LH) and follitropin (follicle-stimulating hormone, FSH) receptors. The β-subunits of these hormones control receptor binding specificity; however, the region of the β-subunit that contacts the receptor has not been identified. By a process of elimination we show this contact to be the portions of β-subunit loops one and three found in a hormone groove created by the juxtaposition of the α- and β-subunits. Most other regions of the β-subunit can be recognized by antibodies that bind to human chorionic hormone (hCG)-receptor complexes or replaced without disrupting hormone function. Using a series of bovine LH/hCG and human FSH/hCG β-subunit chimeras we identified key hCG β-subunit residues in the epitopes of two antibodies that bind to hCG-receptor complexes. These epitopes include the surfaces of β-subunit loops one and three near residue 74 on the outside of the hormone groove and parts of the C-terminal end of the 'seat belt' that holds the two subunits together. The antibody that recognized residue 74 bound to receptor complexes containing most mammalian lutropins better than to the free hormones, an indication that the outside surface of the β- subunit groove is altered during hormone binding. This region of the β- subunit is furthest from the α-subunit and is recognized equally well in the free β-subunit and in the heterodimer. Thus, the receptor associated increase in antibody binding appears due to an interaction of this portion of the β-subunit with the receptor and not to an effect of the receptor on the relative positions of the α- and β-subunits. Unlike most previous studies designed to identify portions of the β-subunit likely to contact the LH receptor, this indirect approach provides data that are more easily interpreted because it does not rely on the use of mutations that disrupt hormone function. The approach described here should be valuable for studying the receptor interactions of other complex ligands.