Molecular Details of Membrane Protein Receptor Dynamics

Información del proyecto

GPCR-MS

Identificador del acuerdo de subvención: 836073

DOI

10.3030/836073

Proyecto cerrado

Fecha de la firma de la CE 20 Marzo 2019

Fecha de inicio 1 Agosto 2019

Fecha de finalización 31 Julio 2021

Financiado con arreglo a

• EXCELLENT SCIENCE - Marie Skłodowska-Curie Actions

Coste total

€ 224 933,76

Aportación de la UE

€ 224 933,76

Coordinado por

THE CHANCELLOR, MASTERS AND SCHOLARS OF THE UNIVERSITY OF OXFORD
United Kingdom

Periodic Reporting for period 1 - GPCR-MS (Molecular Details of Membrane Protein Receptor Dynamics)

Período documentado: 2019-08-01 hasta 2021-07-31

Resumen del contexto y de los objetivos generales del proyecto

G protein-coupled receptors (GPCRs) are the proteins responsible for transducing “outside” signals into an intracellular response and their dysfunction is implicated in many diseases. As a result, they are the target of ~40% of drugs on the market. However, many GPCR drugs elicit off-target effects and can produce unwanted effects such as receptor desensitization and/or drug dependence. Currently, the mechanism of action of many GPCR drug targets is poorly understood.

GPCRs don’t act in isolation; proteins, metabolites, and lipids, and other molecules all interact synergistically to generate distinct signaling outputs. However, conventional ‘omics’ approaches will never be able to connect the status of critical proteins with metabolites or other effectors since the links between them are broken during sample preparation. It is therefore imperative to maintain molecular interactions to understand the molecular network that contributes to a biological signaling output.

This project successfully developed state-of-the-art methods in native mass spectrometry (MS) to interrogate membrane proteins in complex with lipids, ligands, and other molecular interactors intact in the mass spectrometer. By maintaining complexes from the cell to the mass spectrometer, and by dissecting protein-effector interactions in a controlled manner, we are now better equipped to understand mechanisms of membrane protein signal transduction. Specific knowledge of how protein receptors translate “outside” signals into a biological response will change how we approach the treatment of human disease, pain, and other conditions.

Trabajo realizado desde el comienzo del proyecto hasta el final del período abarcado por el informe y los principales resultados hasta la fecha

The project required interdisciplinary training in molecular biology and physical analytical chemistry. Mammalian expression systems were used to produce and isolate membrane protein receptors which provided several advantages toward achieving the goals of the project such as (i) realistic native environments with proper lipid environment, (ii) proper post-translational modifications, and were (iii) easily manipulated by drugs or other stimulants in vitro to induce a desired pathophysiological response (e.g. specific signalling outputs). In addition, novel mass spectrometry workflows were developed to interrogate membrane proteins and their molecular interactors. The use of novel mass spectrometry workflows on proteins involved in human health and disease resulted in three peer-reviewed publications, two conference presentations, and were highlighted in three workshop presentations.

Avances que van más allá del estado de la técnica e impacto potencial esperado (incluida la repercusión socioeconómica y las implicaciones sociales más amplias del proyecto hasta la fecha)

This project successfully developed state-of-the-art methods in native mass spectrometry (MS) to interrogate membrane proteins in complex with lipids, ligands, and other molecular interactors intact in the mass spectrometer. By maintaining complexes from the cell to the MS, and by dissecting protein-effector interactions in a controlled manner, we are now better equipped to understand mechanisms of membrane protein signal transduction. Specific knowledge of how protein receptors translate “outside” signals into a biological response will change how we approach the treatment of human disease, pain, and other conditions.

The fellow using the newest state-of-the-art mass spectrometer to look at membrane protein complexes