title: Identification of novel TSH interaction sites by systematic binding analysis of the TSHR hinge region.
abstract: In which ways the binding of the thyroid stimulating hormone to the extracellular domain of its receptor leads to activation of the thyroid-stimulating hormone receptor (TSHR) is currently only incompletely understood. It is known that TSH binding to the TSHR depends on the interaction with the leucine-rich repeat and sulfation at Y385 of the hinge region. Recently it was also shown that electrostatic interactions between positive charges of bovine (b) TSH and the residues E297, E303, and D382 of the hinge region contribute to hormone-TSHR binding. After the identification of these first TSH binding sites in the hinge region, it was apparent that multiple positions in this region remained to be characterized for their roles in hormone binding. The goal of this study was therefore to clarify whether further contact points of TSH exist in the structurally undefined hinge region. Therefore, we systematically analyzed 41 uncharacterized residues of the TSHR hinge region as single mutants regarding differences between cell surface expression and bTSH binding. Indeed, we identified further amino acids of the hinge region with influence on bTSH binding. Some of these contribute to a new binding domain from human TSHR position F381 to D386. These hinge mutants with influence on bTSH binding were also analyzed for binding of the superagonistic human TSH analog TR1401 demonstrating that these positions also have an impact on TR1401 binding. Moreover, side chain variations revealed that different amino acid properties like the negative charge, aromatic as well as hydrophilic characteristics, contribute to maintain the hormone-TSHR hinge interaction.
authors.affiliation: Division of Endocrinology and Nephrology, University of Leipzig, Liebigstraße 20, D-04103 Leipzig, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Mueller
authors.forename: Mariusz W
authors.initials: MW
authors.lastname: Szkudlinski
authors.forename: Jörg
authors.initials: J
authors.lastname: Schaarschmidt
authors.forename: Robert
authors.initials: R
authors.lastname: Günther
authors.forename: Ralf
authors.initials: R
authors.lastname: Paschke
authors.forename: Holger
authors.initials: H
authors.lastname: Jaeschke
mesh_terms: D000818:Animals; D001665:Binding Sites; D002417:Cattle; D006801:Humans; D011989:Receptors, Thyrotropin / Q000737:chemistry* / Q000378:metabolism; D013972:Thyrotropin / Q000737:chemistry / Q000378:metabolism*
chemical_list: D011989:Receptors, Thyrotropin; D013972:Thyrotropin