Alternatively folded choriogonadotropin analogs. Implications for hormone folding and biological activity.
Most heterodimeric proteins are stabilized by intersubunit contacts or disulfide bonds. In contrast, human chorionic gonadotropin (hCG) and other glycoprotein hormones are secured by a strand of their beta-subunits that is wrapped around alpha-subunit loop 2 "like a seatbelt." During studies of hCG synthesis in COS-7 cells, we found that, when the seatbelt was prevented from forming the disulfide that normally "latches" it to the beta-subunit, its carboxyl-terminal end can "scan" the surface of the heterodimer and become latched by a disulfide to cysteines substituted for residues in the alpha-subunit. Analogs in which the seatbelt was latched to residues 35, 37, 41-43, and 56 of alpha-subunit loop 2 had similar lutropin activities to those of hCG; that in which it was latched to residue 92 at the carboxyl terminus had 10-20% the activity of hCG. Attachment of the seatbelt to alpha-subunit residues 45-51, 86, 88, 90, and 91 reduced lutropin activity substantially. These findings show that the heterodimer can form before the beta-subunit has folded completely and support the notions that the carboxyl-terminal end of the seatbelt, portions of alpha-subunit loop 2, and the end of the alpha-subunit carboxyl terminus do not participate in lutropin receptor interactions. They suggest also that several different architectures could have been sampled without disrupting hormone activity as the glycoprotein hormones diverged from other cysteine knot proteins.
Xing
Y
Y
Department of Obstetrics and Gynecology, Robert Wood Johnson (Rutgers) Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA.
Lin
W
W
Jiang
M
M
Myers
R V
RV
Cao
D
D
Bernard
M P
MP
Moyle
W R
WR
D000409:Alanine / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D016466:CHO Cells; D019556:COS Cells; D006063:Chorionic Gonadotropin / Q000096:biosynthesis / Q000737:chemistry* / Q000652:urine; D006224:Cricetinae; D003545:Cysteine / Q000737:chemistry; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D006728:Hormones / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007118:Immunoassay; D007986:Luteinizing Hormone / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D051381:Rats; D015398:Signal Transduction
D006063:Chorionic Gonadotropin; D004220:Disulfides; D006728:Hormones; D007986:Luteinizing Hormone; D003545:Cysteine; D000409:Alanine