An interactive web-tool for molecular analyses links naturally occurring mutation data with three-dimensional structures of the rhodopsin-like glycoprotein hormone receptors

Kleinau, Gunnar; Kreuchwig, Annika; Worth, Catherine L.; Krause, Gerd

doi:10.1002/humu.21265

Cited by 19 publications

(11 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Most activating GPHR mutations were found in the 7TM domain (46). Based on previous studies (5), we can rule out that the constitutive activity of those 7TM mutations is p10 sequence-mediated.…”

Section: Figure 7 P10 Activates Gphrs In Tissuesmentioning

confidence: 73%

The Activation Mechanism of Glycoprotein Hormone Receptors with Implications in the Cause and Therapy of Endocrine Diseases

Brüser¹,

Schulz²,

Rothemund

et al. 2016

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Glycoprotein hormones (GPHs) are the main regulators of the pituitary-thyroid and pituitary-gonadal axes. Selective interaction between GPHs and their cognate G protein-coupled receptors ensure specificity in GPH signaling. The mechanisms of how these hormones activate glycoprotein hormone receptors (GPHRs) or how mutations and autoantibodies can alter receptor function were unclear. Based on the hypothesis that GPHRs contain an internal agonist, we systematically screened peptide libraries derived from the ectodomain for agonistic activity on the receptors. We show that a peptide (p10) derived from a conserved sequence in the C-terminal part of the extracellular N terminus can activate all GPHRs in vitro and in GPHR-expressing tissues. Inactivating mutations in this conserved region or in p10 can inhibit activation of the thyroid-stimulating hormone receptor by autoantibodies. Our data suggest an activation mechanism where, upon extracellular ligand binding, this intramolecular agonist isomerizes and induces structural changes in the 7-transmembrane helix domain, triggering G protein activation. This mechanism can explain the pathophysiology of activating autoantibodies and several mutations causing endocrine dysfunctions such as Graves disease and hypo-and hyperthyroidism. Our findings highlight an evolutionarily conserved activation mechanism of GPHRs and will further promote the development of specific ligands useful to treat Graves disease and other dysfunctions of GPHRs. Glycoprotein hormone receptors (GPHRs)2 were cloned about 25 years ago (1-3) and, since then, the glycoprotein hormones (GPHs) TSH, LH/hCG, and FSH are considered as the agonists for their respective receptors. In contrast to other rhodopsin-like G protein-coupled receptors (GPCRs), they possess a large extracellular leucine-rich repeat (LRR) hormone-binding domain, which is linked via a hinge region (HR) to the 7-transmembrane helix domain (7TM) (see Fig. 1A). Based on the crystal structure of human FSH bound to the receptor's extracellular domain (ECD), it has been suggested that the FSHR grabs the FSH in a "hand-clasp" mode, orienting the hormone to interact with the extracellular loops and juxtamembrane regions of the 7TM to induce signaling (4). However, to date, all attempts to identify agonistic parts of GPHs have failed, and even extreme high concentrations of hCG did not activate the 7TM of LHR lacking the ECD (5).In addition to TSH and thyrostimulin (6), the TSHR can be activated by mutations in the ECD and by ECD-directed autoantibodies, which cause hyperthyroidism and Graves disease, respectively (7). The detailed mechanism of how the ECD integrates the activating actions of TSH, thyrostimulin, mutations, and autoantibodies to trigger GPHR signal transduction is unsolved, although there are numerous mutagenesis studies (5,8) and crystallographic data of the liganded GPHR ECD (4, 9 -11).Based on findings that proteolytic cleavage of the ECD by trypsin, or artificially generated ECD deletions and truncations, can increase TSHR ac...

show abstract

Section: Figure 7 P10 Activates Gphrs In Tissuesmentioning

confidence: 73%

The Activation Mechanism of Glycoprotein Hormone Receptors with Implications in the Cause and Therapy of Endocrine Diseases

Brüser¹,

Schulz²,

Rothemund

et al. 2016

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…This novel feature can be used to identify interacting residues as counterparts and to evaluate potential mechanisms of molecular malfunction caused by specific mutations. We recently described the interactive 3D-Search applet in detail, demonstrating its usefulness in the analysis of naturally occurring mutations (17).…”

Section: An Innovative New Tool: the Interactive Structure-based Searchmentioning

confidence: 99%

“…This provides a platform for investigation of molecular mechanisms and a better understanding of protein malfunctions. Recently, we described one of the extensions in detail, namely applications for the analysis of pathogenic mutations in three-dimensional (3D) structures (17). Here, we will focus on the complete renewal of the previous database version which includes the addition of several new features as extensions.…”

mentioning

confidence: 99%

Research Resource: Update and Extension of a Glycoprotein Hormone Receptors Web Application

Kreuchwig

Kleinau

Kreuchwig

et al. 2011

Molecular Endocrinology

Self Cite

View full text Add to dashboard Cite

The SSFA-GPHR (Sequence-Structure-Function-Analysis of Glycoprotein Hormone Receptors) database provides a comprehensive set of mutation data for the glycoprotein hormone receptors (covering the lutropin, the FSH, and the TSH receptors). Moreover, it provides a platform for comparison and investigation of these homologous receptors and helps in understanding protein malfunctions associated with several diseases. Besides extending the data set (> 1100 mutations), the database has been completely redesigned and several novel features and analysis tools have been added to the web site. These tools allow the focused extraction of semiquantitative mutant data from the GPHR subtypes and different experimental approaches. Functional and structural data of the GPHRs are now linked interactively at the web interface, and new tools for data visualization (on three-dimensional protein structures) are provided. The interpretation of functional findings is supported by receptor morphings simulating intramolecular changes during the activation process, which thus help to trace the potential function of each amino acid and provide clues to the local structural environment, including potentially relocated spatial counterpart residues. Furthermore, double and triple mutations are newly included to allow the analysis of their functional effects related to their spatial interrelationship in structures or homology models. A new important feature is the search option and data visualization by interactive and user-defined snake-plots. These new tools allow fast and easy searches for specific functional data and thereby give deeper insights in the mechanisms of hormone binding, signal transduction, and signaling regulation. The web application "Sequence-Structure-Function-Analysis of GPHRs" is accessible on the internet at http://www.ssfa-gphr.de/.

show abstract

“…The high conservation of these amino acids in contrast to other hydrophobic residues in this region, like Leu 265 , Leu 267 , or Leu 270 , indicates their evolutionary importance for the TSHR. Screening of available mutagenesis data for the LRRDs of the GPHRs (see the SSFA-GPHR database on the World Wide Web) (57,58) revealed that mutations at specific hydrophobic key positions of the typical LRR profile always lead to an impaired receptor expression level. The hydrophobic positions that were identified here as fold-sensitive match exactly this repeat pattern of hydrophobic residues by pointing into the interior of the LRR domain model (Fig.…”

Section: Hydrophobic Amino Acids At the Junction Between The Leucine-mentioning

confidence: 99%

Defining Structural and Functional Dimensions of the Extracellular Thyrotropin Receptor Region

Kleinau

Mueller

Jaeschke

et al. 2011

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The extracellular region of the thyrotropin receptor (TSHR) can be subdivided into the leucine-rich repeat domain (LRRD) and the hinge region. Both the LRRD and the hinge region interact with thyrotropin (TSH) or autoantibodies. Structural data for the TSHR LRRD were previously determined by crystallization (amino acids Glu 30 -Thr 257 , 10 repeats), but the structure of the hinge region is still undefined. Of note, the amino acid sequence (Trp 258 -Tyr 279 ) following the crystallized LRRD comprises a pattern typical for leucine-rich repeats with conserved hydrophobic side chains stabilizing the repeat fold. Moreover, functional data for amino acids between the LRRD and the transmembrane domain were fragmentary. We therefore investigated systematically these TSHR regions by mutagenesis to reveal insights into their functional contribution and potential structural features. We found that mutations of conserved hydrophobic residues between Thr 257 and Tyr 279 cause TSHR misfold, which supports a structural fold of this peptide, probably as an additional leucine-rich repeat. Furthermore, we identified several new mutations of hydrophilic amino acids in the entire hinge region leading to partial TSHR inactivation, indicating that these positions are important for intramolecular signal transduction. In summary, we provide new information regarding the structural features and functionalities of extracellular TSHR regions. Based on these insights and in context with previous results, we suggest an extracellular activation mechanism that supports an intramolecular agonistic unit as a central switch for activating effects at the extracellular region toward the serpentine domain.

show abstract

An interactive web-tool for molecular analyses links naturally occurring mutation data with three-dimensional structures of the rhodopsin-like glycoprotein hormone receptors

Cited by 19 publications

References 34 publications

The Activation Mechanism of Glycoprotein Hormone Receptors with Implications in the Cause and Therapy of Endocrine Diseases

The Activation Mechanism of Glycoprotein Hormone Receptors with Implications in the Cause and Therapy of Endocrine Diseases

Research Resource: Update and Extension of a Glycoprotein Hormone Receptors Web Application

Defining Structural and Functional Dimensions of the Extracellular Thyrotropin Receptor Region

Contact Info

Product

Resources

About