Partial restoration of lutropin activity by an intersubunit disulfide bond: implications for structure/function studies.
Gonadal function is controlled by lutropins and follitropins, heterodimeric cystine knot proteins that have nearly identical alpha-subunits. These heterodimeric proteins are stabilized by a portion of the hormone-specific beta-subunit termed the "seatbelt" that is wrapped around alpha-subunit loop 2 (alpha 2). Here we show that replacing human chorionic gonadotropin (hCG) alpha 2 residue Lys51 with cysteine or alanine nearly abolished its lutropin activity, an observation that implies that alpha Lys51 has a key role in hormone activity. The activity of the heterodimer containing alpha K51C, but not that containing alpha K51A, was increased substantially when beta-subunit seatbelt residue beta Asp99 was converted to cysteine. As had been reported by others, heterodimers containing alpha K51C and beta D99C were crosslinked by a disulfide. The finding that an intersubunit disulfide restored some of the activity lost by replacing alpha Lys51 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.
Einstein
M
M
Department of Obstetrics and Gynecology, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Lin
W
W
MacDonald
G J
GJ
Moyle
W R
WR
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D018997:Chorionic Gonadotropin, beta Subunit, Human / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003553:Cystine; D004220:Disulfides / Q000378:metabolism*; D005640:Follicle Stimulating Hormone / Q000378:metabolism; D015292:Glycoprotein Hormones, alpha Subunit / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007986:Luteinizing Hormone / Q000378:metabolism*; D017433:Protein Structure, Secondary; D013329:Structure-Activity Relationship
D018997:Chorionic Gonadotropin, beta Subunit, Human; D004220:Disulfides; D015292:Glycoprotein Hormones, alpha Subunit; D003553:Cystine; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone