I03-Macromolecular Crystallography
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Elisa
Peirano
,
Laura
O'Regan
,
Philip
Harrer
,
Ivana
Dzinovic
,
Magda S.
Chegkazi
,
Petra
Havrankova
,
Theresa
Brunet
,
Rossella
Capolino
,
Claudia
Cesario
,
Stefania
Ferro
,
Eva
Hammar
,
Russia
Hà-Vinh Leuchter
,
Elisabetta
Indelicato
,
Maureen
Jacob
,
Lukas
Kunc
,
Henri
Margot
,
Oriano
Marin
,
Maria
Mazurkiewicz-Bełdzińska
,
Niccolò E.
Mencacci
,
Antonio
Novelli
,
Laura
Orec
,
Michael
Poschmann
,
Alexandra
Sitzberger
,
Ugo
Sorrentino
,
Melanie
Spanjaard
,
Matias
Wagner
,
Magdalena
Krygier
,
Sylvia
Boesch
,
Jan
Necpal
,
Matej
Skorvanek
,
Donald L.
Gilbert
,
Robert
Jech
,
Mark
Dodding
,
Roberto A.
Steiner
,
Michael
Zech
Open Access
Abstract: Background: Although de novo causation in dystonia is widely acknowledged, there have been only a few trio-sequencing analyses in this field. We sought to prioritise de novo variants in dystonia and characterise the clinical and molecular features associated with the top gene candidate identified after genomic matchmaking.
Methods: We (re)assessed exome-sequencing data for de novo variants in genes with strong mutational constraint in a sample of 257 dystonia trios. Via data sharing, we collected information on individuals with variants in KLC1, encoding a subunit of the axonal-transport motor protein kinesin-1. Biophysical, biochemical, and functional studies, including differential scanning fluorimetry, X-ray crystallography, fluorescence-polarisation measurements, and immunoprecipitation from cells were performed for representative KLC1 variants. Findings: Missense and loss-of-function de novo variants in constrained genes without implication in autosomal dominant or X-linked conditions were found in 11.7% (30/257) of cases with dystonia. We then ascertained 7 unrelated patients with movement and neurodevelopmental disorders who harboured distinct, predicted deleterious de novo KLC1 missense variants. These variants clustered within the cargo adaptor-binding tetratricopeptide repeat domain and 3 variants mapped to an identical amino-acid position. Highly similar infantile-onset dystonic-spastic phenotypes were observed in the subjects with the recurrently affected residue. For all functionally tested variants, we observed changes in KLC1 stability and/or altered binding behaviour to known kinesin-1 interactors, such as JIP3, previously associated with dystonia and neurodevelopmental impairment. Interpretation: Our research supports the existence of a kinesinopathy linked to KLC1, featuring phenotypic overlap with diseases related to mutational defects of key interactors of KLC1. The full dystonia de-novo variant compendium is reported as a resource for additional disease-gene discovery.
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Jul 2026
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I04-Macromolecular Crystallography
I23-Long wavelength MX
I24-Microfocus Macromolecular Crystallography
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Diamond Proposal Number(s):
[23853]
Open Access
Abstract: Voltage-gated sodium channels (VGSCs) mediate neuronal excitability and synaptic transmission and are functionally relevant targets for volatile anaesthetic (VA) actions. Here, we show that multiple VAs at clinically relevant concentrations share binding sites on NavMs, a prokaryotic VGSC. Sevoflurane, a representative VA, interacts with NavMs and NaChBac with functional effects paralleling those on human VGSCs, including modulation of channel inactivation. X-ray crystallography of purified NavMs reveals an atomic-resolution VA binding site in a VGSC, in which sevoflurane displaces lipid to occupy a membrane-embedded hydrophobic pocket. Alanine substitution of an invariant tyrosine within this binding pocket abolishes sevoflurane binding and eliminates the sevoflurane-induced hyperpolarising shift of steady-state inactivation. Sevoflurane modulates both fast and slow inactivation of human Nav1.1, demonstrating VA modulation of steady-state slow inactivation in a neuronal VGSC. Supporting evidence shows that VAs interact with homologous sites in human VGSCs. These findings define a VA binding site in VGSCs that supports a membrane-assisted pathway for modulating channel gating and neuronal activity in general anaesthesia.
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Jun 2026
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I04-Macromolecular Crystallography
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Rashi
Goel
,
Kristina
Jevdokimenko
,
Ronja
Rehm
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Jannik
Hentze
,
Paola
Agüi-Gonzalez
,
Momchil
Ninov
,
Erik
Maier
,
Yin
Wu
,
Felix
Lange
,
Agata
Witkowska
,
Svenja
Bolz
,
Francesca
Pennacchietti
,
Martina
Damenti
,
Natalie
Kaempf
,
Vladimir
Khayenko
,
Carles
Calatayud
,
Viveka Nand
Malviya
,
Natali L.
Chanaday
,
Emma
Scaletti Hutchinson
,
Hao
Liu
,
Kirsten
Weyand
,
Valentin
Schwarze
,
Daniela
Ivanova
,
Tristan P.
Wallis
,
Christopher
Small
,
Hans M.
Maric
,
Merja
Joensuu
,
Michael A.
Cousin
,
Frédéric A.
Meunier
,
Patrik
Verstreken
,
Ilaria
Testa
,
Ege T.
Kavalali
,
Volker
Haucke
,
Stefan
Jakobs
,
Henning
Urlaub
,
Nils
Brose
,
Benjamin H.
Cooper
,
Pal
Stenmark
,
Felipe
Opazo
,
Reinhard
Jahn
,
Silvio O.
Rizzoli
,
Eugenio F.
Fornasiero
Diamond Proposal Number(s):
[21625]
Open Access
Abstract: Synaptic neurotransmission is a critical hallmark of brain activity and one of the first processes affected in neural diseases. Monitoring this process, particularly synaptic vesicle recycling, in living cells has been instrumental in revealing the mechanisms responsible for neurotransmitter release. However, currently available reporters suffer from limitations, such as large probe sizes or limited compatibility for human neurons, hampering the quantitative analysis of synaptic pathophysiology. Here, we describe the NbLumSyt1 toolkit, a panel of nanobody-based affinity probes that target the luminal domain of the synaptic vesicle protein Synaptotagmin 1 (Syt1). These new tools enable quantitative, noninvasive imaging and functional interrogation of Syt1 exo-endocytosis and trafficking in human neurons, with unprecedented precision, versatility and cost efficiency, in technologies ranging from fixed- and live-cell super-resolution imaging to electron microscopy and mass spectrometry. Overall, NbLumSyt1 nanobinders provide a valuable platform for studying synaptic physiology and pathophysiology, benefiting fundamental neuroscience and translational efforts to study and develop treatments for brain-related disorders.
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May 2026
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I02-Macromolecular Crystallography
I04-Macromolecular Crystallography
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Jakob
Bouton
,
Alexis
Bretteville
,
Gary
Tresadern
,
Paul
Shaffer
,
Nigel
Austin
,
Peter
Buijnsters
,
E. Peder
Cedervall
,
Nicolas
Darville
,
Ineke
Fonteyn
,
Joseph
Leenaerts
,
Carolina Martínez
Lamenca
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Liesbeth
Mertens
,
Daniele
Peeters
,
Adriana I.
Velter
,
Yves Van
Roosbroeck
,
Andreas
Ebneth
,
Jose Manuel
Bartolomé
,
Andrés A.
Trabanco
,
Daniel
Oehlrich
Abstract: Alzheimer’s disease (AD) is a neurodegenerative disorder characterized by extracellular amyloid-β plaque accumulation and intracellular tau neurofibrillary tangles, with tau pathology correlating more closely with cognitive decline. Modulation of tau phosphorylation through the regulation of O-GlcNAcylation, a post-translational modification controlled by O-GlcNAcase (OGA), represents a promising therapeutic strategy. In this study, we report the optimization of a pyrimidine hit identified by high-throughput screening, leading to the discovery and optimization of a novel series of 5-azaindole-based OGA inhibitors. From this series, compound 24 was identified as an in vivo tool candidate that demonstrated a favorable pharmacokinetic profile and measurable brain exposure. Pharmacodynamic studies in murine models demonstrated that compound 24 induced a significant and transient elevation of brain O-GlcNAcylation levels, confirming the in vivo target engagement. These findings underscore the potential of 5-azaindole-based OGA inhibitors as a novel validated chemotype for modulation of O-GlcNAcylation.
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May 2026
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B21-High Throughput SAXS
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Thomas
Lundbäck
,
Vijay
Chandrasekar
,
Chendi
Gu
,
Hyoungseok
Ju
,
Robyn
Mcadam
,
Maria
Palomero
,
Kasim
Sader
,
Bradley
Peter
,
Lisa
Wissler
,
Philip
Nevin
,
Edmund
Foster
,
Tanguy
Jamier
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Pravallika
Manjappa
,
Carina
Johansson
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Jenny
Sandmark
,
Mei
Ding
,
Anette
Persson-Kry
,
Sanhita
Mitra
,
Tugce
Munise Satir
,
Bilada
Bilican
,
Mirko
Messa
,
Graham
Fraser
,
John
Linley
,
Helen
Plant
,
Rachel
Moore
,
Tina
Seifert
,
Michael
Lerche
,
Carina
Raynochek
,
Ewa
Nilsson
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Nour
Majbour
,
Richard
Lucey
,
Taiana
Maia De Oliveira
,
Qi
Wang
,
Iain
Chessell
,
Perla
Breccia
,
Rebecca
Jarvis
Open Access
Abstract: The sterile alpha and TIR motif containing 1 (SARM1) enzyme is a key driver of axonal degeneration in response to injury, making it an attractive target for treating chemotherapy-induced peripheral neuropathy (CIPN) and other nervous system diseases. In this study, we identified and optimised a class of base-exchange inhibitors (BEXi) targeting human SARM1 and explored their molecular interactions and conformational effects using cryo-EM, HDX-MS and SAXS. Although BEXi produced robust inhibition across all biochemical and cellular assay formats, application at sub-inhibitory concentrations consistently led to paradoxical SARM1 activation, and in neuronal assays, accelerated neurite degeneration. Further analysis showed that BEXi only delayed, rather than prevented, neurite degeneration when applied to primary neuronal cells, even at exceedingly high inhibitor concentrations. These results prompted us to discontinue BEXi development in favour of alternative strategies, underscoring the complexity of SARM1 as a therapeutic target and the need for comprehensive, mechanistically informed screening cascades.
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May 2026
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Miguel
Berbeira-Santana
,
Claudia
Peregrina
,
Kosuke
Okuda
,
Jin Chuan
Zhou
,
Maria
Carrasquero-Ordaz
,
Amy V.
Roberts
,
Anne E.
Thomas
,
Evert
Haanappel
,
Matthieu
Chavent
,
Kamel
El Omari
,
Lindsay A.
Baker
,
Daniel T.
Pederick
,
Els
Pardon
,
Jan
Steyaert
,
U. Valentin
Nägerl
,
Daniel
Del Toro
,
Elena
Seiradake
Open Access
Abstract: Cortical migration is a complex process in which neurons migrate along radial glial cells (RGC) to form functional layers. Teneurins (Ten1-4) play a role by interacting with Latrophilins (Lphn/ADGRL1-3). Teneurins are also known as cell adhesion molecules, but how homophilic and heterophilic Teneurin interactions are integrated is unknown. Here, single-particle-cryo-EM data of Ten2 shows that canonical Latrophilin-binding is sterically incompatible with Ten2-dimerisation, making these interactions exclusive. We engineered surface mutations that specifically disrupt Ten2-Ten2 or Ten2-Latrophilin interactions. These are transferrable to Ten4, suggesting conserved binding mechanisms. Proteomics, in-vivo-gene-editing and super-resolution-microscopy show that Ten4 is expressed along RGC fibres and that migrating neurons switch from low-to-high Ten4-expression. Ten4 expression is highest in the cortical plate where Ten4-Ten4 interactions reduce RGC-attachment. In the intermediate zone, Ten4-Latrophilin interactions are required to promote neuron-RGC association. The results show how Ten4 orchestrates different stages of cortical migration by using a structural/functional switch between high-affinity Lphn interactions and low-affinity homophilic interactions, underpinning the integration of distinct migration programmes.
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Apr 2026
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Krios I-Titan Krios I at Diamond
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Brajabandhu
Pradhan
,
Senthil T.
Kumar
,
Jessica
Wagner
,
Rodrigo
Gallardo
,
Gabriele
Orlando
,
Matthias
De Vleeschouwer
,
Valentina
Zorzini
,
Jillian
Madine
,
Nikolaos
Louros
,
Jonas J.
Neher
,
Frederic
Rousseau
,
Joost
Schymkowitz
Diamond Proposal Number(s):
[41018]
Open Access
Abstract: Amyloid assembly in vivo occurs in complex environments where multiple aggregation-prone species coexist. Aβ and medin are prevalent amyloids in ageing humans that co-localize in cerebral amyloid angiopathy (CAA), yet their structural interactions remain poorly understood. Here, using cryo-electron microscopy, we determine high-resolution fibril structures from in vitro mixtures of Aβ40 and medin. From the same reaction, we resolve three distinct fibril populations: (i) a previously characterized Aβ40 polymorph that also forms in isolation, (ii) a Aβ40 polymorph with Aβ42-like features, including ordered N- and C-terminal regions, and (iii) the atomic structure of full-length medin fibrils. Biochemical and immunogold analyses demonstrate Aβ–medin association within mixed assemblies, though medin is not resolved within the ordered Aβ core. These findings support two non-exclusive mechanisms: transient heterotypic interactions redirecting Aβ folding, or partial medin incorporation into fibril architecture. Our data reveal how coexisting amyloids remodel each other’s polymorphic landscapes.
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Apr 2026
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Open Access
Abstract: DISC1 is a pleiotropic protein with essential roles in neuronal proliferation and migration, intracellular signalling and cargo transport. It associates with a diverse array of partner molecules in these contexts. Mutations at the DISC1 locus are strongly associated with a spectrum of mental illnesses such as schizophrenia and depression. Despite its clinical relevance, the molecular architecture and function of DISC1 have remained largely elusive. We present a cryo-EM structure of the entire conserved core region of DISC1. The structure reveals an intricate homotetrameric assembly that harbours conserved bacteria-derived UVR domains. Four of these domains, one from each monomer, mediate extensive contacts forming two asymmetric dimer units. The dimers in turn interface with each other at two distinct coiled coil domains to achieve a two-fold symmetric tetramer. Mutational analysis shows that this tetrameric architecture enables DISC1 to simultaneously bind multiple copies of NDE1 client protein. Importantly, tetramerization and partner binding are structurally independent functions of DISC1. Altogether, our study provides a compelling molecular model of an ancient bacteria protein fold participating in the assembly of a multivalent mammalian scaffold hub that can coordinate multiple partner molecules.
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Apr 2026
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I18-Microfocus Spectroscopy
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Abstract: The results obtained in this thesis demonstrate that NCLX plays a relevant role in the pathophysiology of ischemic stroke, particularly during the reperfusion phase, by modulating ROS production, lipid peroxidation, and iron metabolism. In the murine model, NCLX inhibition effectively reduced infarct volume and cerebral iron overload, although its effects did not extend to other critical aspects of functional recovery, such as oedema, neurodegeneration or glial inflammation. These findings indicate that ferroptosis represents one of the multiple pathological mechanisms involved in ischemic stroke and highlight the relevance of the mitochondrial exchanger NCLX as a key factor at the intersection of mitochondrial metabolism, oxidative stress, and iron homeostasis.
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Mar 2026
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Open Access
Abstract: Perspectives surrounding Cannabis use have transformed over the past decade. This shift in perspective has been noted in pregnant populations in Canada and the US, where various investigations report that the use of Cannabis in pregnancy is increasingly commonplace. There is some evidence indicating that Δ9-tetrahydrocannabinol (THC), the main intoxicating phytocannabinoid found within Cannabis flower, may influence the biochemical composition of lipids within the developing fetal brain. The aim of this study was to apply multimodal biospectroscopic imaging techniques, X-ray fluorescence imaging (XFI) and Fourier transform mid-infrared spectromicroscopy (FTIR), to investigate the biochemical and biomolecular changes underlying the distinct behavioral phenotypes identified previously. XFI was used to investigate the presence of metal, nonmetal, and alkali dysregulation, while FTIR provided information on neurochemical dysbiosis within the brains of offspring exposed to THC (3 mg/kg; i.p.) or vehicle (VEH). The THC offspring exhibited decreased copper (Cu) concentrations within the perimeter of their corpus callosum, as identified by XFI. FTIR hyperspectral data from the brain revealed noteworthy changes in peaks associated with lipid methylene (CH2as), carbohydrates, and peak ratios identifying changes in the lipid structure and the relative content of lipids, cholesterol esters, and cholesterols to saturated fatty acids. These changes were particularly evident in the hippocampus, where THC offspring exhibited increased CH2as, lipid esters, phosphate, protein, and unsaturation levels of lipids. The biochemical changes seen in the FTIR spectra were modest, with THC offspring showing an increase in the number of structural changes of lipids in the corpus callosum and an increase in protein in the lateral ventricle. This study supports the usefulness of these techniques to detect subtle changes in biomolecular composition within brain tissues exposed to gestational THC. These results contribute to the growing body of knowledge unraveling the complex effects of THC on fetal neurodevelopmental trajectories.
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Feb 2026
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