title: The superagonistic activity of bovine thyroid-stimulating hormone (TSH) and the human TR1401 TSH analog is determined by specific amino acids in the hinge region of the human TSH receptor.
abstract: Bovine TSH (bTSH) has a higher affinity to the human TSHR (hTSHR) and a higher signaling activity than human TSH (hTSH). The molecular reasons for these phenomena are unknown. Distinct negatively charged residues (Glu297, Glu303, and Asp382) in the hinge region of the hTSHR are known to be important for bTSH binding and signaling. To investigate the potential relevance of these positions for differences between bTSH and hTSH in the interaction to the hTSHR, we determined bTSH- and hTSH-mediated cAMP production of several substitutions at these three hinge residues. To examine specific variations of hTSH, we also investigated the superagonistic hTSH analog TR1401 (TR1401), whose sequence differs from hTSH by four additional positively charged amino acids that are also present in bTSH. To characterize possible interactions between the acidic hTSHR positions Glu297, Glu303, or Asp382 and the additional basic residues of TR1401, we investigated TR1401 binding and signaling properties. Our data reveal increased cAMP signaling of the hTSHR using TR1401 and bTSH compared with hTSH. Whereas Asp382 seems to be important for bTSH- and TR1401-mediated but not for hTSH-mediated signaling, the substitution E297K exhibits a decreased signaling for all three TSH variants. Interestingly, bTSH and TR1401 showed only a slightly different binding pattern. These observations imply that specific residues of the hinge region are mediators of the superagonistic activity of bTSH and TR1401 in contrast to hTSH. Moreover, the simultaneous localization of binding components in the glycoprotein hormone molecule and the receptor hinge region permits important reevaluation of interacting hormone receptor domains.
authors.affiliation: From the III Medical Department, University of Leipzig, Philipp-Rosenthal-Strasse 27, D-04103 Leipzig, Germany.
authors.forename: Sandra
authors.initials: S
authors.lastname: Mueller
authors.affiliation: Leibniz-Institut für Molekulare Pharmakologie, Robert-Roessle-Strasse 10, D-13125 Berlin, Germany.
authors.forename: Gunnar
authors.initials: G
authors.lastname: Kleinau
authors.affiliation: Trophogen, Incorporated, Rockville, Maryland 20850.
authors.forename: Mariusz W
authors.initials: MW
authors.lastname: Szkudlinski
authors.affiliation: From the III Medical Department, University of Leipzig, Philipp-Rosenthal-Strasse 27, D-04103 Leipzig, Germany.
authors.forename: Holger
authors.initials: H
authors.lastname: Jaeschke
authors.affiliation: Leibniz-Institut für Molekulare Pharmakologie, Robert-Roessle-Strasse 10, D-13125 Berlin, Germany.
authors.forename: Gerd
authors.initials: G
authors.lastname: Krause
authors.affiliation: From the III Medical Department, University of Leipzig, Philipp-Rosenthal-Strasse 27, D-04103 Leipzig, Germany. Electronic address: pasr@medizin.uni-leipzig.de.
authors.forename: Ralf
authors.initials: R
authors.lastname: Paschke
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism; D019556:COS Cells; D002417:Cattle; D002522:Chlorocebus aethiops; D000242:Cyclic AMP / Q000378:metabolism; D005973:Glutamine / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary; D011989:Receptors, Thyrotropin / Q000737:chemistry* / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology; D013045:Species Specificity; D013972:Thyrotropin / Q000031:analogs & derivatives / Q000235:genetics* / Q000378:metabolism*
chemical_list: D011989:Receptors, Thyrotropin; D005973:Glutamine; D001224:Aspartic Acid; D013972:Thyrotropin; D000242:Cyclic AMP