Please Note: The main page lists projects via BBSRC Research Theme(s) quoted and then relevant Topic(s).
Targeting novel molecular mechanisms of family B G-protein-coupled receptors for drug discovery
Secondary Supervisor(s): Dr Giuseppe Deganutti
University of Registration: Coventry University
BBSRC Research Themes:
Project Outline
G-protein-coupled receptors (GPCRs) are the largest family of membrane proteins and a major target for drug discovery due to their regulation of a plethora of biological responses.
A major limitation in the drug discovery pipeline is a lack of dynamic structural and functional information available for new GPCR targets. Extracting target receptors from their native cell membrane environment is highly detrimental to their stability. To circumvent this problem and enable GPCR studies in a more native-like setting, we employ styrene-co-maleic acid (SMA) copolymers (a synthetic polymer) to encapsulate receptors in SMA lipid particles (SMALPs) which preserve the native nano-scale lipid bilayer environment (Ayub et al, 2024, 10.1016/j.abb.2024.109946). This technique is well-established in our lab and novel bespoke polymer tools are being synthesised in collaboration with Prof Bert Klumperman (Stellenbosch University, South Africa).
This project will focus on the calcitonin receptor-like receptor (CLR), an important family B GPCR that is significantly under-explored, despite its huge therapeutic potential. A further complication is the obligation of CLR to dimerise with one of three receptor-activity modifying proteins (RAMP1, 2 & 3) to form a functional receptor family. CLR regulates the immune, cardiovascular and central nervous systems and is associated with neuro-inflammation, heart failure and cancer (Pioszak et al, 2020, 10.1016/bs.apha.2020.01.001).
It is known that CLR’s functional properties are dictated by interactions with RAMPs, ligands, G-proteins and lipids, however, the underlying molecular mechanisms dictating the receptors’ dynamic conformation and structure-function are poorly understood. The project will address this knowledge gap by focusing on three distinct CLR-based macromolecules: calcitonin gene-related peptide receptor (CGRPR; CLR-RAMP1), adrenomedullin 1 receptor (AM1R; CLR-RAMP2) and adrenomedullin 2 receptor (AM2R; CLR-RAMP3). Novel genetically engineered mammalian cell system will be generated to study CLR-RAMPs molecular mechanisms within SMA nanodiscs. This will aid rational design of chemical probes and potential therapeutics targeting CLR-RAMPs.
Objective
The primary objective of this project is to generate mechanistic insight into CLR structural dynamics and molecular interactions and the contributions of RAMPs, diverse ligands and natural lipids, that are known to modulate receptor functions. This will be achieved through three parallel research strands:
- Pharmacological investigation will be performed on CLR-RAMP complexes using fluorescence/bioluminescence and second messenger assays. Engineered CGRPR, AM1R and AM2R constructs and fluorescent ligands will be used to determine molecular interactions in mammalian cells.
- Computational modelling will be used to capture the main structural changes of CLR at the atomic level upon activation with ligands and in the presence of lipids.
- CLR structural dynamics will be examined experimentally in SMALP nanodiscs with diverse binding partners. This includes a wide range of ligands, RAMPs, and signalling proteins that will be studied through multitude of biophysical and structural techniques.
Method
The PGR will learn state-of-the-art technologies at Coventry University. The project will employ in vitro pharmacology assays, SMA nanotechnology, biophysical methodology and in silico molecular dynamic simulations to study CLR complexes (Deganutti et al, 2022, 10.1038/s41467-021-27760-0).
Impact
This multi-disciplinary project will impact drug design to improve patient benefit through understanding the structure-function intricacy of family B GPCRs influenced by RAMPs and CGRP family of ligands. Out of ~826 diverse GPCRs in humans, only ~12% are currently used as approved medicine targets, where family B receptors are even less studied but highly valuable targets. Therefore, this work has therapeutic relevance and will generate pharmaceutical interest.
