G-protein coupled receptors (GPCRs) are indispensable signalling
molecules that orchestrate a myriad of physiological processes, rendering
them pivotal therapeutic targets. Remarkably, approximately 35% of
approved drugs modulate GPCR activity, underscoring their profound
significance. Nonetheless, understanding the mechanisms underpinning
GPCR activation remains a challenge, particularly in the context of the GnRH1 receptor (GnRH1R). This receptor is dedicated to mediating the effects of gonadotropin-releasing hormone (GnRH) and the production of fertility hormones, exhibits unusual structural characteristics, such as the absence of a C-terminal helix and variations in what are otherwise highly conserved motifs amongst the GPCR superfamily. Unveiling the GnRH binding that triggers GnRH1R activation is imperative, and characterisation of the GnRH1R active conformation will address a critical knowledge gap.
The research methodology employed a combination of computational
docking and molecular dynamic (MD) simulations. Docking simulations were
performed using the Rosetta software and tailored protocols for membrane
proteins and flexible peptide ligands. The docking process resulted into
thousands of potential GnRH binding modes which were clustered and
filtered based on criteria such as energy metrics and contacts with
experimentally important receptor residues. The binding mode elimination
process yielded two promising binding modes representing the native-like
GnRH-GnRH1R complex. MD simulations of the selected best ranking
binding modes were performed to investigate their ability to activate the
GnRH1R. Receptor activation was monitored and evaluated by the increase of the distance between two key transmembrane helices (TM3 and TM6) - a widely accepted characteristic of GPCR activation. One of the predicted binding modes induced activation after 1.0 μs of simulation and displayed an
increased TM3-TM6 distance (~13 Å) compared to the inactive GnRH1R (~8
Å). GnRH binding displayed high stability and involved several ππ interactions, especially through tryptophan 3 (W3) and tyrosine 5 (Y5) of
GnRH and the CWxPY motif of the receptor. The crucial residue R8 of GnRH
was found to form cation-π interactions with W2806.48 of the CWxPY motif and in addition, it mediated interaction to the G-protein binding pocket (DRS
motif) and the DPxxY motif through interactions with residues of TM3 and
water mediated hydrogen bonds respectfully. The resulting active GnRH1R
conformation exhibited an open G-protein binding pocket, an enlarged ligand
binding pocket and key rearrangements of the DPxxY motif indicative of
activation. Communication between GnRH1R and lipid molecules was also
observed. The significance of this work transcends the confines of GnRH1R,
contributing essential knowledge to the broader field of GPCR biology and
computational studies. The identified binding mechanism and proposed
interhelical communication networks offer valuable insights for drug design
targeting GnRH1R, with implications for developing more precise
therapeutics for reproductive system-related disorders. Finally, the details of
the discovered communication pathways initiated by GnRH pave the way for
a deeper understanding of GPCR activation.
| Date of Award | 8 Oct 2024 |
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| Original language | English |
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| Awarding Institution | - University Of Strathclyde
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| Sponsors | University of Strathclyde |
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| Supervisor | Paul Mulheran (Supervisor) & Valerie Ferro (Supervisor) |
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