Structure-based design and protein engineering of intersubunit disulfide bonds in gonadotropins.
Pairs of cystine residues were introduced in the alpha- and beta-subunits of human choriogonadotropin at positions with optimal geometries for the formation of disulfide bonds. Using the homology with luteinizing hormone and follicle stimulating hormone, similar mutations were carried out in these glycoprotein hormones. In nearly all mutants the corresponding disulfide bonds were formed leading to a non-natural, covalent linkage between the alpha- and beta-subunits. The mutants typically display wild-type receptor binding and bioactivity. The mutants with non-natural intersubunit disulfide bonds display enhanced thermostabilities relative to the corresponding heterodimeric glycoprotein hormones, rendering them candidates for long acting gonadotropins with enhanced shelf lives.
Heikoop
J C
JC
N.V. Organon, Oss, The Netherlands.
van den Boogaart
P
P
Mulders
J W
JW
Grootenhuis
P D
PD
D000818:Animals; D001709:Biotechnology; D016466:CHO Cells; D006063:Chorionic Gonadotropin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006224:Cricetinae; D003553:Cystine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D015195:Drug Design; D004355:Drug Stability; D005640:Follicle Stimulating Hormone / Q000737:chemistry / Q000235:genetics; D006062:Gonadotropins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007986:Luteinizing Hormone / Q000737:chemistry / Q000235:genetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D015202:Protein Engineering; D011974:Receptors, LH / Q000378:metabolism; D015398:Signal Transduction; D013696:Temperature; D014162:Transfection
D006063:Chorionic Gonadotropin; D004220:Disulfides; D006062:Gonadotropins; D011974:Receptors, LH; D003553:Cystine; D007986:Luteinizing Hormone; D005640:Follicle Stimulating Hormone