Bridging biophysics and cell biology: The role of G protein-coupled receptor conformations in signalling

Informacje na temat projektu

signalling dynamics

Identyfikator umowy o grant: 844622

DOI

10.3030/844622

Projekt został zamknięty

Data podpisania przez KE 12 Kwietnia 2019

Data rozpoczęcia 1 Stycznia 2020

Data zakończenia 31 Grudnia 2022

Finansowanie w ramach

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Koszt całkowity

€ 271 732,80

Wkład UE

€ 271 732,80

Koordynowany przez

UNITED KINGDOM RESEARCH AND INNOVATION
United Kingdom

Periodic Reporting for period 2 - signalling dynamics (Bridging biophysics and cell biology: The role of G protein-coupled receptor conformations in signalling)

Okres sprawozdawczy: 2022-01-01 do 2022-12-31

Podsumowanie kontekstu i ogólnych celów projektu

G protein coupled receptors (GPCRs) are a class of membrane receptors that transmits extracellular signals into the cell. They can be activated by a diverse set of ligands including small molecules, hormones, neurotransmitters, or photons and are targeted by a third of currently marketed drugs. Endogenous ligands and drugs may exhibit different efficacy profiles, ranging from full activation to complete inactivation of a signalling pathway. The key to the selective interaction with signalling partners in response to ligand binding lies in the conformational flexibility of the membrane receptors. Previous research has extensively studied the three-dimensional structures of GPCRs and their signalling. However, the link between active conformations and signalling is still missing.

In this project, we are filling this gap by linking the three-dimensional structure of the receptor to signalling using a combination of biophysical experiments, computational analyses and high throughput signalling approaches. Our objectives are to further our understanding of the conformational changes in GPCRs and how they are linked to signalling. Which of the conformational changes observed are important for signalling and which ones are not? Which residues of the receptor are the most important ones for translating a ligand signal into an intracellular signal? Can this process be modified, either through changes in the ligand or through modulation of signal transduction at a later stage?

Better understanding the molecular basis behind signalling, including the conformational changes that the receptor undergoes upon activation, are important for the development of better drugs with fewer side effects. Understanding how exactly ligand binding is converted into an intracellular signalling event will help researchers modify existing ligands or help them design new ligands with better characteristics.

In conclusion, we have developed a framework for integrating pharmacological and structural data in this project. This allows us to determine which parts of the receptor are key for its functions, which parts are structurally important, pharmacologically important, or both. This improves our interpretation of both pharmacological and structural data. Our approach can be applied to other proteins where functional and structural data are available at similar resolution.

Prace wykonane od początku projektu do końca okresu sprawozdawczego oraz najważniejsze dotychczasowe rezultaty

To address these questions, we use cell-based signalling assays that allow us to follow how intracellular signalling partners are activated with increasing concentrations of extracellular ligands. By introducing single amino-acid changes in the receptor, we can then determine which of the many amino acids of the receptor are the most important ones for function. We have performed cell-based signalling assays and analysed the data for over 400 mutations in two medically relevant GPCRs. Selected mutations were then studied using biophysical methods to learn more about the reasons why certain effects were observed in our signalling assays. Finally, we combine the signalling data with computational analyses to better understand signal transduction through the receptor and how this may be modified.

By applying computational analyses to our signalling data, we were able to show which residues of a receptor contribute to signalling, how they are distributed and how conserved these positions are likely to be across different GPCRs. We found that highly conserved residues, ligand- and G protein-binding sites of the receptor could only explain a fraction of the residues that we found to be important using our mutagenesis approach. In addition, involvement in structural changes alone did not necessarily correlate with pharmacological importance of a residue. We found that the a subset of residues formed an allosteric network of connections between the ligand- and G- protein binding sites which is key for receptor activation and signalling. Our data can be used to explain the function of allosteric modulators at known binding sites and pinpoint additional sites that could be targeted in the future.

So far, we have presented the results of our work at multiple scientific conferences and have submitted a paper for publication.

Innowacyjność oraz oczekiwany potencjalny wpływ (w tym dotychczasowe znaczenie społeczno-gospodarcze i szersze implikacje społeczne projektu)

During this project we have developed new computational methods to maximise the insights gained from high throughput signalling data in GPCRs. Our insights will be helpful for medicinal and computational chemists focusing on GPCR drug design, promising to improve the design of novel drugs with less side effects.

Overview of approach: combining GPCR signalling and conformation data