Periodic Reporting for period 4 - PentaBrain (Structural studies of mammalian Cys-loop receptors)
Reporting period: 2019-12-01 to 2021-11-30
In terms of methods, we have optimised ways of obtaining large quantities of pure and stable receptors, which enables their imaging by cryo-electron microscopy. We also participated in making that imaging easier when the receptors are embedded into a tiny discs made of lipids.
In terms of applications, we have shown how serotonin receptors activate and how a panel of antiemetics (including tropisetron and palonosetron) or an antidepressant (vortioxetine) inhibit them. We have also shown how agonists, and in particular nicotine, binds to the muscle-type acetylcholine receptor. We have identified that agonist-bound receptors (both the serotonin and the acetylcholine ones) can exist in intermediate states. We have described how the receptors block or allow the passage of ions (the signalling) through a gate, using a mechanism of wetting/dewetting.
Taken together, the project has shed light onto the molecular mechanism of action and the structural pharmacology of pentameric-ligand channels and has helped the development of techniques for their study.
A large emphasis was put on the serotonin 5-HT3 receptor. Using electron microscopy, we have obtained four structures of the serotonin 5-HT3 receptor. With an inhibited state, a putative pre-active state, and two putative distinct open states, these structures have extended our understanding of the gating mechanisms. They have also provided unprecedented details of serotonin binding and of orthosteric and allosteric site geometries. Those key results were published in an article lead by Dr. Polovinkin, the first PhD student of the team, in the journal Nature in 2018. We have also provided a more exhaustive description of the binding of anti-emetics to the serotonin receptor, using a combination of electron microscopy and molecular dynamics.
We have also studied the acetylcholine receptor that sits at the neuromuscular junction, using the homologous receptor from the Torpedo fish. The Torpedo fish receptor is of key historical importance : it was the first receptor ever discovered. We have shown how agonists, and in particular nicotine at high concentration, bind to and activate the acetylcholine receptor. We have described a possible intermediate state of the receptor in the presence of an agonist, a result that questions the assumption that agonist-bound receptors rather exist in a state called desensitised, and is therefore of broad relevance to the research field.
Those results were reported in scientific publications, and presented at European and international conferences by the team members.