Award details

A dynamic view of GPCR-G protein complexes: insight into partial agonism and G protein selectivity

Reference	BB/W020718/1
Principal Investigator / Supervisor	Professor Daniel Nietlispach
Co-Investigators / Co-Supervisors	Dr Andrew Jones
Institution	University of Cambridge
Department	Biochemistry
Funding type	Research
Value (£)	553,243
Status	Current
Type	Research Grant
Start date	05/11/2022
End date	04/11/2025
Duration	36 months

Abstract

GPCRs are signaling proteins that, when activated by agonists change their conformation, enabling transducers such as heterotrimeric G proteins or arrestins to bind. Structural and biophysical data have given us extensive insight into how agonists affect the function of GPCRs. However, how conformational changes in the receptor affect coupling and activation of intracellular transducers remains poorly understood. Major questions remain e.g.: Why do certain agonists produce a less than full response (known as partial agonism)? Why does a given receptor bind selectively to one particular subtype of the G protein family over another? Currently, we are reliant on structures of immobilised proteins but to answer these questions we need additional insight on GPCR-G protein complexes that reveals their dynamic nature. We propose to investigate this using NMR, to provide a much-needed dynamic explanation of how GPCRs and G proteins interact. NMR provides a dynamic, atomistic-level description of structure and conformational variability and can detect both low and highly populated states. Preliminary NMR studies in our lab with b1AR revealed agonist-dependent conformational variations on the cytoplasmic side of b1AR-G protein complexes. We hypothesise that, depending on the agonist bound, these complexes can adopt a range of specific active conformational states that facilitate different levels of allosteric signal transmission. We will study a range of agonists and complexes with different G proteins to establish how conformational variability arises. We will relate our molecular observations to biophysical properties, such as binding affinity, kinetics of exchange and nucleotide exchange rates, that link to signaling function, and we will build 3D model structures of b1AR-G protein complexes. Overall, our work will provide a dynamic perspective of GPCR-G protein complexes and reveal key molecular features of partial agonism and G protein selectivity.

Summary

The human body is made of many cells, which work together in large numbers to sustain life. Each cell is surrounded by a lipid membrane that is impenetrable to most chemicals. However, in order to work properly together, individual cells need to communicate with each other and respond or adjust to external needs. To help with that, a host of sensor proteins, so-called G protein-coupled receptors (GPCRs) are embedded in the lipid cell envelope. There are over 850 different kinds of GPCRs in humans and their task is to sense the presence of a wide range of ligand molecules (agonists) on the outside of the cell and to communicate that information across the lipid membrane. GPCRs excel at this, as each kind of receptor is specialised to bind a particular set of ligands. Ligand binding changes the shape of the receptor, which in turn allows other proteins, so-called transducers, to bind to that part of the receptor on the inside of the cell membrane. Coupling of the transducer to the GPCR then activates a wide range of cellular signaling processes, which instruct the cell to respond and take necessary actions. The net effect is that the presence of a particular type of ligand on the outside of a cell has been communicated to the cell interior. Owing to their central role as sensors of the cell, GPCRs regulate a wide range of physiological processes and hence are central to human function in health and disease. Not surprisingly, a large proportion of prescription drugs target GPCRs. A defining feature of GPCRs is their mobility. They can easily change conformation and the effortlessness with which agonist-binding influences this strongly relates to the intrinsic function of these receptors. Similarly, GPCRs interact with their transducer G proteins in a dynamic fashion but how these GPCR-G protein complexes interact is currently poorly understood. Most structural investigations have relied on methods that reveal the GPCRs within static complexes, where the binding partners have usually been engineered into a stabilised form. This overlooks the fact, however, that the binding partners remain inherently mobile and can adopt a range of different conformations. We propose therefore to conduct studies using a method called NMR, which allows investigation of these proteins while retaining their natural mobility. We will investigate in greater detail how GPCRs interact with their transducers, in particular the family of heterotrimeric G proteins. Obtaining structural insight into the molecular details of these interactions, while preserving the dynamic nature of these proteins, is key for improving our understanding of how these receptors work. Through our investigations with the GPCR b1AR we will learn the molecular determinants of GPCR interactions with G proteins. We will discover how the dynamic nature of the complexes relate to the properties of the binding partners and how their conformations vary with the type of agonist bound and the type of G protein that interacts. Correlating our molecular observations from NMR with biophysical properties of these complexes, such as affinity of the binding partners, how rapidly the complexes are formed and how the rate of nucleotide exchange is affected, will inform us on how these conformational differences affect signaling function. While increasing our general knowledge of these receptor-transducer complexes and how GPCRs and G proteins dynamically interact, our work also will inform us on two major unresolved questions that are key for the function of these proteins: Why do certain ligands when bound to a receptor result in a moderated, less than maximal response? Why does a particular GPCR interact with a selected member of the G protein family preferably over another one? Both questions have direct implications for cellular signaling and our investigations will provide a dynamic perspective of the key molecular determinants.

Committee	Research Committee D (Molecules, cells and industrial biotechnology)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	X - not in an Initiative
Funding Scheme	X – not Funded via a specific Funding Scheme

I accept the terms and conditions of use (opens in new window)

export PDF file

back to list new search