The Cyclic Imine Core Common to the Marine Macrocyclic Toxins Is Sufficient to Dictate Nicotinic Acetylcholine Receptor Antagonism.

Macrocyclic imine phycotoxins are an emerging class of chemical compounds associated with harmful algal blooms and shellfish toxicity. Earlier binding and electrophysiology experiments on nAChR subtypes and their soluble AChBP surrogates evidenced common trends for substantial antagonism, binding affinities, and receptor-subtype selectivity. Earlier, complementary crystal structures of AChBP complexes showed that common determinants within the binding nest at each subunit interface confer high-affinity toxin binding, while distinctive determinants from the flexible loop C, and either capping the nest or extending toward peripheral subsites, dictate broad versus narrow receptor subtype selectivity.

Report from the 29th Meeting on Toxinology, "Toxins: From the Wild to the Lab", Organized by the French Society of Toxinology on 30 November-1 December 2023.

The French Society of Toxinology (SFET), which celebrated its 30th anniversary this year, organized its 29th annual Meeting (RT29), shared by 87 participants, on 30 November-1 December 2023. The RT29 main theme, "Toxins: From the Wild to the Lab", focused on research in the field of animal venoms and animal, bacterial, fungal, or plant toxins, from their discovery in nature to their study in the laboratory. The exploration of the functions of toxins, their structures, their molecular or cellular ligands, their mode of action, and their potential therapeutic applications were emphasized during oral communications and posters through three sessions, of which each was dedicated to a secondary theme.

The ESTHER database on alpha/beta hydrolase fold proteins - An overview of recent developments.

The ESTHER database, dedicated to ESTerases and alpha/beta-Hydrolase Enzymes and Relatives (https://bioweb.supagro.inra.

Report from the 28th Meeting on Toxinology, "Toxins: What's up, Doc?", Organized by the French Society of Toxinology on 28-29 November 2022.

The French Society of Toxinology (SFET) organized its 28th annual meeting on 28-29 November 2022 (RT28). The central theme of this meeting was "Toxins: What's up, Doc?", emphasizing the latest findings on animal, bacterial, algal, plant and fungal toxins through sessions dedicated to deep learning, toxin tracking and toxinomic advances, shared by ca. 80 participants.

Patient-derived antibodies reveal the subcellular distribution and heterogeneous interactome of LGI1.

Autoantibodies against leucine-rich glioma-inactivated 1 (LGI1) occur in patients with encephalitis who present with frequent focal seizures and a pattern of amnesia consistent with focal hippocampal damage. To investigate whether the cellular and subcellular distribution of LGI1 may explain the localization of these features, and hence gain broader insights into LGI1's neurobiology, we analysed the detailed localization of LGI1 and the diversity of its protein interactome, in mouse brains using patient-derived recombinant monoclonal LGI1 antibodies. Combined immunofluorescence and mass spectrometry analyses showed that LGI1 is enriched in excitatory and inhibitory synaptic contact sites, most densely within CA3 regions of the hippocampus.

Report from the 27th (Virtual) Meeting on Toxinology, "Toxins: Mr Hyde or Dr Jekyll?", Organized by the French Society of Toxinology, 9-10 December 2021.

The French Society of Toxinology (SFET) organized its 27th annual meeting on 9-10 December 2021 as a virtual meeting (e-RT27). The central theme of this meeting was "Toxins: Mr Hyde or Dr Jekyll?", emphasizing the latest findings on plant, fungal, algal, animal and bacterial toxins during 10 lectures, 15 oral communications (shorter lectures) and 20 posters shared by ca. 80 participants.

Comparative mapping of selected structural determinants on the extracellular domains of cholinesterase-like cell-adhesion molecules.

Cell adhesion generally involves formation of homophilic or heterophilic protein complexes between two cells to form transcellular junctions. Neural cell-adhesion members of the α/β-hydrolase fold superfamily of proteins use their extracellular or soluble cholinesterase-like domain to bind cognate partners across cell membranes, as illustrated by the neuroligins. These cell-adhesion molecules currently comprise the synaptic organizers neuroligins found in all animal phyla, along with three proteins found only in invertebrates: the guidance molecule neurotactin, the glia-specific gliotactin, and the basement membrane protein glutactin.

The neuroligins and the synaptic pathway in Autism Spectrum Disorder.

The genetics underlying autism spectrum disorder (ASD) is complex and heterogeneous, and de novo variants are found in genes converging in functional biological processes. Neuronal communication, including trans-synaptic signaling involving two families of cell-adhesion proteins, the presynaptic neurexins and the postsynaptic neuroligins, is one of the most recurrently affected pathways in ASD. Given the role of these proteins in determining synaptic function, abnormal synaptic plasticity and failure to establish proper synaptic contacts might represent mechanisms underlying risk of ASD.

The Ig-like domain of Punctin/MADD-4 is the primary determinant for interaction with the ectodomain of neuroligin NLG-1.

Punctin/MADD-4, a member of the ADAMTSL extracellular matrix protein family, was identified as an anterograde synaptic organizer in the nematode At GABAergic neuromuscular junctions, the short isoform MADD-4B binds the ectodomain of neuroligin NLG-1, itself a postsynaptic organizer of inhibitory synapses. To identify the molecular bases of their partnership, we generated recombinant forms of the two proteins and carried out a comprehensive biochemical and biophysical study of their interaction, complemented by an localization study. We show that spontaneous proteolysis of MADD-4B first generates a shorter N-MADD-4B form, which comprises four thrombospondin (TSP) domains and one Ig-like domain and binds NLG-1.

Report from the 26th Meeting on Toxinology, "Bioengineering of Toxins", Organized by the French Society of Toxinology (SFET) and Held in Paris, France, 4-5 December 2019.

This 26th edition of the annual Meeting on Toxinology (RT26) of the SFET (http://sfet.asso.fr/international) was held at the Institut Pasteur of Paris on 4-5 December 2019 [.

An evolutionary perspective on the first disulfide bond in members of the cholinesterase-carboxylesterase (COesterase) family: Possible outcomes for cholinesterase expression in prokaryotes.

Within the alpha/beta hydrolase fold superfamily of proteins, the COesterase group (carboxylesterase type B, block C, cholinesterases …) diverged from the other groups through simultaneous integration of an N-terminal, first disulfide bond and a significant increase in the protein mean size. This first disulfide bond ties a large Cys loop, which in the cholinesterases is named the omega loop and forms the upper part of the active center gorge, essential for the high catalytic activity of these enzymes. In some non-catalytic members of the family, the loop may be necessary for heterologous partner recognition.

Hot Spots for Protein Partnerships at the Surface of Cholinesterases and Related α/β Hydrolase Fold Proteins or Domains-A Structural Perspective.

The hydrolytic enzymes acetyl- and butyryl-cholinesterase, the cell adhesion molecules neuroligins, and the hormonogenic macromolecule thyroglobulin are a few of the many members of the α/β hydrolase fold superfamily of proteins. Despite their distinctive functions, their canonical subunits, with a molecular surface area of ~20,000 Ų, they share binding patches and determinants for forming homodimers and for accommodating structural subunits or protein partners. Several of these surface regions of high functional relevance have been mapped through structural or mutational studies, while others have been proposed based on biochemical data or molecular docking studies.

A Triad of Crystals Sheds Light on MDGA Interference with Neuroligation.

Neurexins and neuroligins form trans-synaptic complexes that promote synapse development. In this issue of Neuron, Aricescu and colleagues (Elegheert et al., 2017) complement and strengthen two recent reports by the Kim and Rudenko teams (Kim et al.

Preface: Cholinergic Mechanisms.

This special issue is a companion to the meeting 'XVth International Symposium on Cholinergic Mechanisms', and is edited by Israel Silman, Marco Prado and Pascale Marchot. In the review articles, renowned researchers in the field capture key mechanisms of cholinergic neurotransmission, from genomic amplification of cholinesterase genes, splicing and post-translational modifications; features of the neuromuscular junction, implications of cholinergic circuitry that are relevant to addiction, anxiety and mood, to preclinical models, protein biomarkers, and clinical findings that are relevant to pathology, for example, developmental neurotoxicity. The broad variety of features reflects the impact of cholinergic mechanisms on many physiological events and emphasizes the importance of research in this area.

Natural genomic amplification of cholinesterase genes in animals.

Tight control of the concentration of acetylcholine at cholinergic synapses requires precise regulation of the number and state of the acetylcholine receptors, and of the synthesis and degradation of the neurotransmitter. In particular, the cholinesterase activity has to be controlled exquisitely. In the genome of the first experimental models used (man, mouse, zebrafish and drosophila), there are only one or two genes coding for cholinesterases, whereas there are more genes for their closest relatives the carboxylesterases.

Cyclic imine toxins from dinoflagellates: a growing family of potent antagonists of the nicotinic acetylcholine receptors.

We present an overview of the toxicological profile of the fast-acting, lipophilic macrocyclic imine toxins, an emerging family of organic compounds associated with algal blooms, shellfish contamination and neurotoxicity. Worldwide, shellfish contamination incidents are expanding; therefore, the significance of these toxins for the shellfish food industry deserves further study. Emphasis is directed to the dinoflagellate species involved in their production, their chemical structures, and their specific mode of interaction with their principal natural molecular targets, the nicotinic acetylcholine receptors, or with the soluble acetylcholine-binding protein, used as a surrogate receptor model.

The three-finger toxin fold: a multifunctional structural scaffold able to modulate cholinergic functions.

Three-finger fold toxins are miniproteins frequently found in Elapidae snake venoms. This fold is characterized by three distinct loops rich in β-strands and emerging from a dense, globular core reticulated by four highly conserved disulfide bridges. The number and diversity of receptors, channels, and enzymes identified as targets of three-finger fold toxins is increasing continuously.

Relationships of human α/β hydrolase fold proteins and other organophosphate-interacting proteins.

Organophosphates (OPs) are either found in nature or synthetized for use as pesticides, flame retardants, neurotoxic warfare agents or drugs (cholinergic enhancers in Alzheimer's disease and myasthenia gravis, or inhibitors of lipases in metabolic diseases). Because of the central role of acetylcholinesterase cholinergic neurotransmission in humans, one of the main purposes for using OPs is inactivation of the enzyme by phosphorylation of the nucleophilic serine residue in the active center. However, hundreds of serine hydrolases are expressed in the human proteome, and many of them are potential targets for OP adduction.

Steric and Dynamic Parameters Influencing In Situ Cycloadditions to Form Triazole Inhibitors with Crystalline Acetylcholinesterase.

Ligand binding sites on acetylcholinesterase (AChE) comprise an active center, at the base of a deep and narrow gorge lined by aromatic residues, and a peripheral site at the gorge entry. These features launched AChE as a reaction vessel for in situ click-chemistry synthesis of high-affinity TZ2PA6 and TZ2PA5 inhibitors, forming a syn-triazole upon cycloaddition within the gorge from alkyne and azide reactants bound at the two sites, respectively. Subsequent crystallographic analyses of AChE complexes with the TZ2PA6 regioisomers demonstrated that syn product association is accompanied by side chain reorganization within the gorge, freezing-in-frame a conformation distinct from an unbound state or anti complex.

Marine Macrocyclic Imines, Pinnatoxins A and G: Structural Determinants and Functional Properties to Distinguish Neuronal α7 from Muscle α1(2)βγδ nAChRs.

Pinnatoxins are macrocyclic imine phycotoxins associated with algal blooms and shellfish toxicity. Functional analysis of pinnatoxin A and pinnatoxin G by binding and voltage-clamp electrophysiology on membrane-embedded neuronal α7, α4β2, α3β2, and muscle-type α12βγδ nicotinic acetylcholine receptors (nAChRs) reveals high-affinity binding and potent antagonism for the α7 and α12βγδ subtypes. The toxins also bind to the nAChR surrogate, acetylcholine-binding protein (AChBP), with low Kd values reflecting slow dissociation.

Crystal structure of snake venom acetylcholinesterase in complex with inhibitory antibody fragment Fab410 bound at the peripheral site: evidence for open and closed states of a back door channel.

The acetylcholinesterase found in the venom of Bungarus fasciatus (BfAChE) is produced as a soluble, non-amphiphilic monomer with a canonical catalytic domain but a distinct C terminus compared with the other vertebrate enzymes. Moreover, the peripheral anionic site of BfAChE, a surface site located at the active site gorge entrance, bears two substitutions altering sensitivity to cationic inhibitors. Antibody Elec410, generated against Electrophorus electricus acetylcholinesterase (EeAChE), inhibits EeAChE and BfAChE by binding to their peripheral sites.

The neuroligins and their ligands: from structure to function at the synapse.

The neuroligins are cell adhesion proteins whose extracellular domain belongs to the α/β-hydrolase fold family of proteins, mainly containing enzymes and exemplified by acetylcholinesterase. The ectodomain of postsynaptic neuroligins interacts through a calcium ion with the ectodomain of presynaptic neurexins to form flexible trans-synaptic associations characterized by selectivity for neuroligin or neurexin subtypes. This heterophilic interaction, essential for synaptic differentiation, maturation, and maintenance, is regulated by gene selection, alternative mRNA splicing, and posttranslational modifications.

Tracking the origin and divergence of cholinesterases and neuroligins: the evolution of synaptic proteins.

A cholinesterase activity can be found in all kingdoms of living organism, yet cholinesterases involved in cholinergic transmission appeared only recently in the animal phylum. Among various proteins homologous to cholinesterases, one finds neuroligins. These proteins, with an altered catalytic triad and no known hydrolytic activity, display well-identified cell adhesion properties.