Stable isotope labeling by amino acids in cell culture based proteomics reveals differences in protein abundances between spiral and coccoid forms of the gastric pathogen Helicobacter pylori

Müller, Stephan; Pernitzsch, Sandy R.; Haange, Sven-Bastiaan; Uetz, Peter; Bergen, Martin von; Sharma, Cynthia M.; Kalkhof, Stefan

doi:10.1016/j.jprot.2015.05.011

Cited by 19 publications

(16 citation statements)

References 111 publications

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“…MS Analysis of Cross-linked Soluble TSHR-ECD-Chemical cross-linking of the soluble TSHR-ECD with bTSH was performed as previously described (12,22) 5 and DST were freshly prepared in dry DMSO to a final concentration of 0.25 mol/liter and 0.1 mol/liter, respectively.…”

Section: Methodsmentioning

confidence: 99%

Rearrangement of the Extracellular Domain/Extracellular Loop 1 Interface Is Critical for Thyrotropin Receptor Activation

Schaarschmidt

Nagel

Huth

et al. 2016

Journal of Biological Chemistry

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The thyroid stimulating hormone receptor (TSHR) is a G protein-coupled receptor (GPCR) with a characteristic large extracellular domain (ECD). TSHR activation is initiated by binding of the hormone ligand TSH to the ECD. How the extracellular binding event triggers the conformational changes in the transmembrane domain (TMD) necessary for intracellular G protein activation is poorly understood. To gain insight in this process, the knowledge on the relative positioning of ECD and TMD and the conformation of the linker region at the interface of ECD and TMD are of particular importance. To generate a structural model for the TSHR we applied an integrated structural biology approach combining computational techniques with experimental data. Chemical cross-linking followed by mass spectrometry yielded 17 unique distance restraints within the ECD of the TSHR, its ligand TSH, and the hormone-receptor complex. These structural restraints generally confirm the expected binding mode of TSH to the ECD as well as the general fold of the domains and were used to guide homology modeling of the ECD. Functional characterization of TSHR mutants confirms the previously suggested close proximity of Ser-281 and Ile-486 within the TSHR. Rigidifying this contact permanently with a disulfide bridge disrupts ligand-induced receptor activation and indicates that rearrangement of the ECD/extracellular loop 1 (ECL1) interface is a critical step in receptor activation. The experimentally verified contact of Ser-281 (ECD) and Ile-486 (TMD) was subsequently utilized in docking homology models of the ECD and the TMD to create a full-length model of a glycoprotein hormone receptor. Glycoprotein hormones (GPHs)3 normally regulate crucial processes in metabolism and reproduction by activating GPHRs. This is especially true for TSHR, which can cause several clinically relevant conditions like hypo-and hyperthyroidism when it malfunctions. Yet the mechanism of how extracellular ligand binding induces the structural changes required for intracellular G protein activation is unknown. We pursued an integrated structural biology approach using modeling guided by experimental data to generate experimentally supported full-length TSHR models. It is expected that some insights gleaned from a TSHR model can be generalized to other GPHRs. These models in turn create testable hypotheses on the mechanism of GPHR activation and can promote drug development to treat GPHR-associated diseases.GPHs bind to the ECD of their respective receptors (Fig. 1A) and consequently initiate activation, which is presumably propagated by induction of conformational changes within the ECD's hinge region (HR) (1-4). Interestingly, GPHRs still possess a binding site within the TMD not associated with physiological receptor activation but accessible to low molecular weight agonists and allosteric modulators (5-7). Another important aspect of GPHR function and physiology is posttranslational modification, including disulfide bond formation, glycosylation, tyrosine sulfation, and p...

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Section: Methodsmentioning

confidence: 99%

Rearrangement of the Extracellular Domain/Extracellular Loop 1 Interface Is Critical for Thyrotropin Receptor Activation

Schaarschmidt

Nagel

Huth

et al. 2016

Journal of Biological Chemistry

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“…Moreover, proteins controlling host colonization, including arginase RocF and CagA, were also downregulated. Various outer membrane proteins were detected with elevated expression, suggesting a role for host cell adherence . Another recent study revealed that H. pylori infection supports glutamine and glutathione deficiency of the host, which is connected to tissue damage and inflammation .…”

Section: Orientation and Colonization Of H Pylori In The Stomachmentioning

confidence: 97%

“…A novel study investigated these H. pylori forms by quantitative proteomics. Interestingly, enzymes involved in cell division as well as transcription and translation exhibited a lower abundance in coccoid cells . Moreover, proteins controlling host colonization, including arginase RocF and CagA, were also downregulated.…”

Section: Orientation and Colonization Of H Pylori In The Stomachmentioning

confidence: 99%

“…Interestingly, enzymes involved in cell division as well as transcription and translation exhibited a lower abundance in coccoid cells. 3 Moreover, proteins controlling host colonization, including arginase RocF and CagA, were also downregulated. Various outer membrane proteins were detected with elevated expression, suggesting a role for host cell adherence.…”

mentioning

confidence: 99%

“…Various outer membrane proteins were detected with elevated expression, suggesting a role for host cell adherence. 3 Another recent study revealed that H. pylori infection supports glutamine and glutathione deficiency of the host, which is connected to tissue damage and inflammation. 4 In a related study, it was demonstrated that H. pylori represses H,K-ATPase in a NF-κB-dependent and interleukin-8 (IL-8)-independent manner to decrease acid secretion in the stomach, enhancing bacterial colonization.…”

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confidence: 99%

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Pathogenesis of Helicobacter pylori infection

et al. 2016

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Helicobacter pylori is estimated to infect more than half of the worlds human population and represents a major risk factor for chronic gastritis, peptic ulcer disease, MALT lymphoma, and gastric adenocarcinoma. H. pylori infection and clinical consequences are controlled by highly complex interactions between the host, colonizing bacteria, and environmental parameters. Important bacterial determinants linked with gastric disease development include the cag pathogenicity island encoding a type IV secretion system (T4SS), the translocated effector protein CagA, vacuolating cytotoxin VacA, adhesin BabA, urease, serine protease HtrA, secreted outer membrane vesicles, and many others. The high quantity of these factors and allelic changes in the corresponding genes reveals a sophisticated picture and problems in evaluating the impact of each distinct component. Extensive work has been performed to pinpoint molecular processes related to H. pylori-triggered pathogenesis using Mongolian gerbils, mice, primary tissues, as well as novel in vitro model systems such as gastroids. The manipulation of host signaling cascades by the bacterium appears to be crucial for inducing pathogenic downstream activities and gastric disease progression. Here, we review the most recent advances in this important research area.

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Sulfur-34S and 36S Stable Isotope Labeling of Amino Acids for Quantification (SULAQ34/36) of Proteome Analyses

Herbst

Jehmlich

Bergen

et al. 2018

Methods in Molecular Biology

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Stable isotope labeling by amino acids in cell culture based proteomics reveals differences in protein abundances between spiral and coccoid forms of the gastric pathogen Helicobacter pylori

Cited by 19 publications

References 111 publications

Rearrangement of the Extracellular Domain/Extracellular Loop 1 Interface Is Critical for Thyrotropin Receptor Activation

Rearrangement of the Extracellular Domain/Extracellular Loop 1 Interface Is Critical for Thyrotropin Receptor Activation

Pathogenesis of Helicobacter pylori infection

Sulfur-34S and 36S Stable Isotope Labeling of Amino Acids for Quantification (SULAQ34/36) of Proteome Analyses

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