Partial Restoration of Lutropin Activity by an Intersubunit Disulfide Bond: Implications for Structure/Function Studies

Abstract

Gonadal function is controlled by lutropins and follitropins, heterodimeric cystine knot proteins that have nearly identical α-subunits. These heterodimeric proteins are stabilized by a portion of the hormone-specific β-subunit termed the “seatbelt” that is wrapped around α-subunit loop 2 (α2). Here we show that replacing human chorionic gonadotropin (hCG) α2 residue Lys51 with cysteine or alanine nearly abolished its lutropin activity, an observation that implies that αLys51 has a key role in hormone activity. The activity of the heterodimer containing αK51C, but not that containing αK51 A, was increased substantially when β-subunit seatbelt residue pAsp99 was converted to cysteine. As had been reported by others, heterodimers containing αK51C and βD99C were crosslinked by a disulfide. The finding that an intersubunit disulfide restored some of the activity lost by replacing αLys51 suggests that this residue is not crucial for receptor binding or signaling and also that hCG and related hormones may be particularly sensitive to mutations that alter interactions between their subunits. We propose the unique structures of hCG and related family members may permit some subunit movement in the heterodimer, making it difficult to deduce key residues involved in receptor contacts simply by correlating the activities of hormone analogs with their amino acid sequences.