title: Structure determination of helical filaments by solid-state NMR spectroscopy.
abstract: The controlled formation of filamentous protein complexes plays a crucial role in many biological systems and represents an emerging paradigm in signal transduction. The mitochondrial antiviral signaling protein (MAVS) is a central signal transduction hub in innate immunity that is activated by a receptor-induced conversion into helical superstructures (filaments) assembled from its globular caspase activation and recruitment domain. Solid-state NMR (ssNMR) spectroscopy has become one of the most powerful techniques for atomic resolution structures of protein fibrils. However, for helical filaments, the determination of the correct symmetry parameters has remained a significant hurdle for any structural technique and could thus far not be precisely derived from ssNMR data. Here, we solved the atomic resolution structure of helical MAVS(CARD) filaments exclusively from ssNMR data. We present a generally applicable approach that systematically explores the helical symmetry space by efficient modeling of the helical structure restrained by interprotomer ssNMR distance restraints. Together with classical automated NMR structure calculation, this allowed us to faithfully determine the symmetry that defines the entire assembly. To validate our structure, we probed the protomer arrangement by solvent paramagnetic resonance enhancement, analysis of chemical shift differences relative to the solution NMR structure of the monomer, and mutagenesis. We provide detailed information on the atomic contacts that determine filament stability and describe mechanistic details on the formation of signaling-competent MAVS filaments from inactive monomers.
authors.affiliation: Laboratory for Macromolecular Interactions, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany;
authors.forename: Lichun
authors.identifier: http://orcid.org/0000-0002-0772-0452
authors.initials: L
authors.lastname: He
authors.affiliation: Unité de Bioinformatique Structurale, CNRS UMR 3528, Institut Pasteur, 75015 Paris, France;
authors.forename: Benjamin
authors.identifier: http://orcid.org/0000-0003-4014-9195
authors.initials: B
authors.lastname: Bardiaux
authors.affiliation: Laboratory for Macromolecular Interactions, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany; Department of Physics, Faculty of Science, Suez University, Suez, 43533, Egypt;
authors.forename: Mumdooh
authors.initials: M
authors.lastname: Ahmed
authors.affiliation: Laboratory for Macromolecular Interactions, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany;
authors.forename: Johannes
authors.initials: J
authors.lastname: Spehr
authors.affiliation: Research Group "Host-Pathogen Interactions", Paul-Ehrlich-Institut, 63225 Langen, Germany;
authors.forename: Renate
authors.initials: R
authors.lastname: König
authors.affiliation: Central Unit for Microscopy, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany;
authors.forename: Heinrich
authors.initials: H
authors.lastname: Lünsdorf
authors.affiliation: Immune Aging and Chronic Infection, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany;
authors.forename: Ulfert
authors.initials: U
authors.lastname: Rand
authors.affiliation: SeNostic GmbH, 38124 Braunschweig, Germany.
authors.forename: Thorsten
authors.initials: T
authors.lastname: Lührs
authors.affiliation: Laboratory for Macromolecular Interactions, Helmholtz Centre for Infection Research, 38124 Braunschweig, Germany; Christiane.Ritter@helmholtz-hzi.de.
authors.forename: Christiane
authors.initials: C
authors.lastname: Ritter
mesh_terms: D048868:Adaptor Proteins, Signal Transducing / Q000737:chemistry*; D057809:HEK293 Cells; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D016296:Mutagenesis; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D015203:Reproducibility of Results; D012997:Solvents
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D012997:Solvents
keywords: MAVS; grid search; innate immunity; protein structure; solid-state NMR