Functional Reconstitution of the 5-HT3 Receptor

Abstract

The 5-HT3 receptor is a pentameric ligand-gated ion channel (pLGIC) embedded in the membrane of neurons, which plays an important role during fast neurotransmission in the brain and the gut. Mutations in this receptor have been associated to neurological and psychiatric disorders such as epilepsy, depression and schizophrenia; as a result, 5-HT3 receptor, among other pLGICs, is the target of a legion of therapeutic compounds. Therefore, understanding the molecular mechanisms of the 5-HT3 receptor can provide both fundamental biological insights and potential benefits to health.Our laboratory has previously obtained several structures of the 5-HT3 receptor in different conformations: inhibited, active open-pore and pre-active using both crystallography and Cryo-EM. All these structures, that shed light on conformational transitions of the 5-HT3 receptor, have been obtained in the presence of detergent. Detergents are useful biochemical tools; however, they might distort protein structure. Their putative deleterious influence should be assessed on case-by-case basis. The aim of the project is to achieve the functional reconstitution of the 5-HT3 receptor by evaluating several membrane mimetics. The most interesting results so far have been obtained with saposin A nanoparticles, with which we could obtain cryo-EM datasets of reconstituted 5-HT3 receptor. Surprisingly, while the conformation of the extracellular domain is similar to that in detergent, the structure shows an important level of distortion in the transmembrane α-helices. We are now investigating more extensively how various receptor to lipids to saposin A ratios lead to transmembrane domain distortions.