|
1
|
Hima S, Aiswarya N, Remya C, Vasudevan DM, Dileep KV, Francis D. Deciphering protein aggregation: Insights into morphology, contributing factors, and molecular pathologies. ADVANCES IN PROTEIN CHEMISTRY AND STRUCTURAL BIOLOGY 2025; 145:23-71. [PMID: 40324848 DOI: 10.1016/bs.apcsb.2024.11.009] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/07/2025]
Abstract
Protein aggregation research stands at the cutting edge of biomedical science, offering crucial insights into the molecular underpinnings of neurodegenerative and amyloid-associated diseases. Significant advancements in deciphering the structural, biophysical, and molecular intricacies of protein misfolding are driving the development of innovative therapies. Emerging approaches, from small molecule inhibitors to sophisticated polymer-based therapeutics, hold great promise for alleviating the toxic impacts of aggregation with the potential to prevent, delay, or even reverse disease progression. Despite these advances, the field contends with substantial challenges. The polymorphic and complex nature of protein aggregates poses major obstacles to both research and therapeutic design. Yet, interdisciplinary methodologies-integrating advanced spectroscopic, imaging, and computational tools-are creating new pathways to address these complexities, effectively bridging molecular breakthroughs and practical therapeutic applications. The rapid shift of foundational discoveries to clinical trials marks a pivotal step forward, instilling new hope for patients with protein aggregation disorders. Each breakthrough propels us closer to life-changing therapies that may reshape the outlook for these patients. The promise of precise and effective treatments is driving a transformative shift in medical science, establishing protein aggregation research as a crucial pillar in combating these challenging diseases and offering a beacon of hope for the future of neurodegenerative care.
Collapse
Affiliation(s)
- Sree Hima
- Laboratory for Computational and Structural Biology, Jubilee Centre for Medical Research, Jubilee Mission Medical College and Research Institute, Thrissur, Kerala, India; Amrita Institute of Medical Sciences and Research Center, Amrita Vishwa Vidyapeetham, Kochi, India
| | - N Aiswarya
- Laboratory for Computational and Structural Biology, Jubilee Centre for Medical Research, Jubilee Mission Medical College and Research Institute, Thrissur, Kerala, India; Amrita Institute of Medical Sciences and Research Center, Amrita Vishwa Vidyapeetham, Kochi, India
| | - Chandran Remya
- Laboratory for Computational and Structural Biology, Jubilee Centre for Medical Research, Jubilee Mission Medical College and Research Institute, Thrissur, Kerala, India
| | - D M Vasudevan
- Laboratory for Computational and Structural Biology, Jubilee Centre for Medical Research, Jubilee Mission Medical College and Research Institute, Thrissur, Kerala, India; Amrita Institute of Medical Sciences and Research Center, Amrita Vishwa Vidyapeetham, Kochi, India
| | - K V Dileep
- Laboratory for Computational and Structural Biology, Jubilee Centre for Medical Research, Jubilee Mission Medical College and Research Institute, Thrissur, Kerala, India; Amrita Institute of Medical Sciences and Research Center, Amrita Vishwa Vidyapeetham, Kochi, India.
| | - Dileep Francis
- Department of Life sciences, Kristu Jayanti College, Autonomous, Bengaluru, Karnataka, India.
| |
Collapse
|
|
2
|
Kell DB, Pretorius E. Proteomic Evidence for Amyloidogenic Cross-Seeding in Fibrinaloid Microclots. Int J Mol Sci 2024; 25:10809. [PMID: 39409138 PMCID: PMC11476703 DOI: 10.3390/ijms251910809] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2024] [Revised: 10/01/2024] [Accepted: 10/03/2024] [Indexed: 10/20/2024] Open
Abstract
In classical amyloidoses, amyloid fibres form through the nucleation and accretion of protein monomers, with protofibrils and fibrils exhibiting a cross-β motif of parallel or antiparallel β-sheets oriented perpendicular to the fibre direction. These protofibrils and fibrils can intertwine to form mature amyloid fibres. Similar phenomena can occur in blood from individuals with circulating inflammatory molecules (and also some originating from viruses and bacteria). Such pathological clotting can result in an anomalous amyloid form termed fibrinaloid microclots. Previous proteomic analyses of these microclots have shown the presence of non-fibrin(ogen) proteins, suggesting a more complex mechanism than simple entrapment. We thus provide evidence against such a simple entrapment model, noting that clot pores are too large and centrifugation would have removed weakly bound proteins. Instead, we explore whether co-aggregation into amyloid fibres may involve axial (multiple proteins within the same fibril), lateral (single-protein fibrils contributing to a fibre), or both types of integration. Our analysis of proteomic data from fibrinaloid microclots in different diseases shows no significant quantitative overlap with the normal plasma proteome and no correlation between plasma protein abundance and their presence in fibrinaloid microclots. Notably, abundant plasma proteins like α-2-macroglobulin, fibronectin, and transthyretin are absent from microclots, while less abundant proteins such as adiponectin, periostin, and von Willebrand factor are well represented. Using bioinformatic tools, including AmyloGram and AnuPP, we found that proteins entrapped in fibrinaloid microclots exhibit high amyloidogenic tendencies, suggesting their integration as cross-β elements into amyloid structures. This integration likely contributes to the microclots' resistance to proteolysis. Our findings underscore the role of cross-seeding in fibrinaloid microclot formation and highlight the need for further investigation into their structural properties and implications in thrombotic and amyloid diseases. These insights provide a foundation for developing novel diagnostic and therapeutic strategies targeting amyloidogenic cross-seeding in blood clotting disorders.
Collapse
Affiliation(s)
- Douglas B. Kell
- Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, Faculty of Health and Life Sciences, University of Liverpool, Crown St., Liverpool L69 7ZB, UK
- The Novo Nordisk Foundation Centre for Biosustainability, Building 220, Søltofts Plads 200, Technical University of Denmark, 2800 Kongens Lyngby, Denmark
- Department of Physiological Sciences, Faculty of Science, Stellenbosch University, Private Bag X1 Matieland, Stellenbosch 7602, South Africa
| | - Etheresia Pretorius
- Department of Biochemistry, Cell and Systems Biology, Institute of Systems, Molecular and Integrative Biology, Faculty of Health and Life Sciences, University of Liverpool, Crown St., Liverpool L69 7ZB, UK
- Department of Physiological Sciences, Faculty of Science, Stellenbosch University, Private Bag X1 Matieland, Stellenbosch 7602, South Africa
| |
Collapse
|
|
3
|
Emmanouilidis L, Bartalucci E, Kan Y, Ijavi M, Pérez ME, Afanasyev P, Boehringer D, Zehnder J, Parekh SH, Bonn M, Michaels TCT, Wiegand T, Allain FHT. A solid beta-sheet structure is formed at the surface of FUS droplets during aging. Nat Chem Biol 2024; 20:1044-1052. [PMID: 38467846 PMCID: PMC11288893 DOI: 10.1038/s41589-024-01573-w] [Citation(s) in RCA: 22] [Impact Index Per Article: 22.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2023] [Accepted: 02/07/2024] [Indexed: 03/13/2024]
Abstract
Phase transitions are important to understand cell dynamics, and the maturation of liquid droplets is relevant to neurodegenerative disorders. We combined NMR and Raman spectroscopies with microscopy to follow, over a period of days to months, droplet maturation of the protein fused in sarcoma (FUS). Our study reveals that the surface of the droplets plays a critical role in this process, while RNA binding prevents it. The maturation kinetics are faster in an agarose-stabilized biphasic sample compared with a monophasic condensed sample, owing to the larger surface-to-volume ratio. In addition, Raman spectroscopy reports structural differences upon maturation between the inside and the surface of droplets, which is comprised of β-sheet content, as revealed by solid-state NMR. In agreement with these observations, a solid crust-like shell is observed at the surface using microaspiration. Ultimately, matured droplets were converted into fibrils involving the prion-like domain as well as the first RGG motif.
Collapse
Affiliation(s)
- Leonidas Emmanouilidis
- Department of Biology, Institute of Biochemistry, ETH Zurich, Zurich, Switzerland.
- Bringing Materials to Life Initiative, ETH Zurich, Zurich, Switzerland.
| | - Ettore Bartalucci
- Max Planck Institute for Chemical Energy Conversion, Mülheim/Ruhr, Germany
- Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Aachen, Germany
| | - Yelena Kan
- Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, USA
- Max Planck Institute for Polymer Research, Mainz, Germany
| | - Mahdiye Ijavi
- Department of Materials, ETH Zurich, Zurich, Switzerland
| | - Maria Escura Pérez
- Department of Biology, Institute of Biochemistry, ETH Zurich, Zurich, Switzerland
| | | | | | - Johannes Zehnder
- Laboratory of Physical Chemistry, ETH Zurich, Zurich, Switzerland
| | - Sapun H Parekh
- Department of Biomedical Engineering, University of Texas at Austin, Austin, TX, USA
- Max Planck Institute for Polymer Research, Mainz, Germany
| | - Mischa Bonn
- Max Planck Institute for Polymer Research, Mainz, Germany
| | - Thomas C T Michaels
- Department of Biology, Institute of Biochemistry, ETH Zurich, Zurich, Switzerland
- Bringing Materials to Life Initiative, ETH Zurich, Zurich, Switzerland
| | - Thomas Wiegand
- Max Planck Institute for Chemical Energy Conversion, Mülheim/Ruhr, Germany.
- Institute of Technical and Macromolecular Chemistry, RWTH Aachen University, Aachen, Germany.
- Laboratory of Physical Chemistry, ETH Zurich, Zurich, Switzerland.
| | - Frédéric H-T Allain
- Department of Biology, Institute of Biochemistry, ETH Zurich, Zurich, Switzerland.
- Bringing Materials to Life Initiative, ETH Zurich, Zurich, Switzerland.
| |
Collapse
|
|
4
|
Scheidt HA, Korn A, Schwarze B, Krueger M, Huster D. Conformation of Pyroglutamated Amyloid β (3-40) and (11-40) Fibrils - Extended or Hairpin? J Phys Chem B 2024; 128:1647-1655. [PMID: 38334278 PMCID: PMC10895672 DOI: 10.1021/acs.jpcb.3c07285] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2023] [Revised: 01/04/2024] [Accepted: 01/30/2024] [Indexed: 02/10/2024]
Abstract
Amyloid β (Aβ) is a hallmark protein of Alzheimer's disease. One physiologically important Aβ variant is formed by initial N-terminal truncation at a glutamic acid position (either E3 or E11), which is subsequently cyclized to a pyroglutamate (either pE3 or pE11). Both forms have been found in high concentrations in the core of amyloid plaques and are likely of high importance in the pathology of Alzheimer's disease. However, the molecular structure of the fibrils of these variants is not entirely clear. Solid-state NMR spectroscopy studies have reported a molecular contact between Gly25 and Ile31, which would disagree with the conventional hairpin model of wildtype (WT-)Aβ1-40 fibrils, most often described in the literature. We investigated the conformation of the monomeric unit of pE3-Aβ3-40 and pE11-Aβ11-40 (and for comparison also wildtype (WT)-Aβ1-40) fibrils to find out whether the hairpin or a newly suggested extended structure dominates the structure of the Aβ monomers in these fibrils. To this end, solid-state NMR spectroscopy was applied probing the inter-residual contacts between Phe19/Leu34, Ala21/Leu34, and especially Gly25/Ile31 using suitable isotopic labeling schemes. In the second part, the flexible turn of the Aβ40 peptides was replaced by a (3-(3-aminomethyl)phenylazo)phenylacetic acid (AMPP)-based photoswitch, which can predefine the peptide conformation to either an extended (trans) or hairpin (cis) conformation. This enables simultaneous spectroscopic assessment of the conformation of the AMPP-photoswitch, allowing in situ structural investigations during fibrillation in contrast to structural techniques such as NMR spectroscopy or cryo-EM, which can only be applied to stable conformers. Both methods confirm an extended structure for the peptidic monomers in fibrils of all investigated Aβ variants. Especially the Gly25/Ile31 contact is a decisive indicator for the extended structure along with the characteristic absorption spectra of trans-AMPP-Aβ.
Collapse
Affiliation(s)
- Holger A. Scheidt
- Institute
for Medical Physics and Biophysics, Leipzig
University Härtelstr. 16/18, D-04107 Leipzig, Germany
| | - Alexander Korn
- Institute
for Medical Physics and Biophysics, Leipzig
University Härtelstr. 16/18, D-04107 Leipzig, Germany
| | - Benedikt Schwarze
- Institute
for Medical Physics and Biophysics, Leipzig
University Härtelstr. 16/18, D-04107 Leipzig, Germany
| | - Martin Krueger
- Institute
of Anatomy, Leipzig University, Liebigstr. 13, 04103 Leipzig, Germany
| | - Daniel Huster
- Institute
for Medical Physics and Biophysics, Leipzig
University Härtelstr. 16/18, D-04107 Leipzig, Germany
| |
Collapse
|
|
5
|
Lends A, Birlirakis N, Cai X, Daskalov A, Shenoy J, Abdul-Shukkoor MB, Berbon M, Ferrage F, Liu Y, Loquet A, Tan KO. Efficient 18.8 T MAS-DNP NMR reveals hidden side chains in amyloid fibrils. JOURNAL OF BIOMOLECULAR NMR 2023:10.1007/s10858-023-00416-5. [PMID: 37289306 DOI: 10.1007/s10858-023-00416-5] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Subscribe] [Scholar Register] [Received: 01/04/2023] [Accepted: 05/02/2023] [Indexed: 06/09/2023]
Abstract
Amyloid fibrils are large and insoluble protein assemblies composed of a rigid core associated with a cross-β arrangement rich in β-sheet structural elements. It has been widely observed in solid-state NMR experiments that semi-rigid protein segments or side chains do not yield easily observable NMR signals at room temperature. The reasons for the missing peaks may be due to the presence of unfavorable dynamics that interfere with NMR experiments, which result in very weak or unobservable NMR signals. Therefore, for amyloid fibrils, semi-rigid and dynamically disordered segments flanking the amyloid core are very challenging to study. Here, we show that high-field dynamic nuclear polarization (DNP), an NMR hyperpolarization technique typically performed at low temperatures, can circumvent this issue because (i) the low-temperature environment (~ 100 K) slows down the protein dynamics to escape unfavorable detection regime, (ii) DNP improves the overall NMR sensitivity including those of flexible side chains, and (iii) efficient cross-effect DNP biradicals (SNAPol-1) optimized for high-field DNP (≥ 18.8 T) are employed to offer high sensitivity and resolution suitable for biomolecular NMR applications. By combining these factors, we have successfully established an impressive enhancement factor of ε ~ 50 on amyloid fibrils using an 18.8 T/ 800 MHz magnet. We have compared the DNP efficiencies of M-TinyPol, NATriPol-3, and SNAPol-1 biradicals on amyloid fibrils. We found that SNAPol-1 (with ε ~ 50) outperformed the other two radicals. The MAS DNP experiments revealed signals of flexible side chains previously inaccessible at conventional room-temperature experiments. These results demonstrate the potential of MAS-DNP NMR as a valuable tool for structural investigations of amyloid fibrils, particularly for side chains and dynamically disordered segments otherwise hidden at room temperature.
Collapse
Affiliation(s)
- Alons Lends
- CNRS, Chemistry and Biology of Membranes and Nanoobjects (CBMN), UMR 5348, Institut Europeen de Chimie et Biologie (IECB), University of Bordeaux, 33600, Pessac, France
| | - Nicolas Birlirakis
- Laboratoire des Biomolécules, LBM, Département de Chimie, École Normale Supérieure, PSL University, Sorbonne Université, CNRS, 75005, Paris, France
| | - Xinyi Cai
- Tianjin Key Laboratory on Technologies Enabling Development of Clinical Therapeutics and Diagnostics, School of Pharmacy, Tianjin Medical University, Tianjin, 300070, China
| | - Asen Daskalov
- CNRS, Chemistry and Biology of Membranes and Nanoobjects (CBMN), UMR 5348, Institut Europeen de Chimie et Biologie (IECB), University of Bordeaux, 33600, Pessac, France
| | - Jayakrishna Shenoy
- CNRS, Chemistry and Biology of Membranes and Nanoobjects (CBMN), UMR 5348, Institut Europeen de Chimie et Biologie (IECB), University of Bordeaux, 33600, Pessac, France
| | - Muhammed Bilal Abdul-Shukkoor
- CNRS, Chemistry and Biology of Membranes and Nanoobjects (CBMN), UMR 5348, Institut Europeen de Chimie et Biologie (IECB), University of Bordeaux, 33600, Pessac, France
| | - Mélanie Berbon
- CNRS, Chemistry and Biology of Membranes and Nanoobjects (CBMN), UMR 5348, Institut Europeen de Chimie et Biologie (IECB), University of Bordeaux, 33600, Pessac, France
| | - Fabien Ferrage
- Laboratoire des Biomolécules, LBM, Département de Chimie, École Normale Supérieure, PSL University, Sorbonne Université, CNRS, 75005, Paris, France
| | - Yangping Liu
- Tianjin Key Laboratory on Technologies Enabling Development of Clinical Therapeutics and Diagnostics, School of Pharmacy, Tianjin Medical University, Tianjin, 300070, China
| | - Antoine Loquet
- CNRS, Chemistry and Biology of Membranes and Nanoobjects (CBMN), UMR 5348, Institut Europeen de Chimie et Biologie (IECB), University of Bordeaux, 33600, Pessac, France.
| | - Kong Ooi Tan
- Laboratoire des Biomolécules, LBM, Département de Chimie, École Normale Supérieure, PSL University, Sorbonne Université, CNRS, 75005, Paris, France.
| |
Collapse
|
|
6
|
Deike S, Rothemund S, Voigt B, Samantray S, Strodel B, Binder WH. β-Turn mimetic synthetic peptides as amyloid-β aggregation inhibitors. Bioorg Chem 2020; 101:104012. [PMID: 32683138 DOI: 10.1016/j.bioorg.2020.104012] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2020] [Revised: 06/09/2020] [Accepted: 06/10/2020] [Indexed: 12/28/2022]
Abstract
Aggregation of amyloid peptides results in severe neurodegenerative diseases. While the fibril structures of Aβ40 and Aβ42 have been described recently, resolution of the aggregation pathway and evaluation of potent inhibitors still remains elusive, in particular in view of the hairpin-region of Aβ40. We here report the preparation of beta-turn mimetic conjugates containing synthetic turn mimetic structures in the turn region of Aβ40 and Aβ16-35, replacing 2 amino acids in the turn-region G25 - K28. The structure of the turn mimic induces both, acceleration of fibrillation and the complete inhibition of fibrillation, confirming the importance of the turn region on the aggregation. Replacing position G25-S26 provided the best inhibition effect for both beta-turn mimetics, the bicyclic BTD 1 and the aromatic TAA 2, while positions N27-K28 and V24-G25 showed only weaker or no inhibitory effects. When comparing different turn mimetics at the same position (G25-S26), conjugate 1a bearing the BTD turn showed the best inhibition of Aβ40 aggregation, while 5-amino-valeric acid 4a showed the weakest effect. Thus there is a pronounced impact on fibrillation with the chemical nature of the embedded beta-turn-mimic: the conformationally constrained turns 1 and 2 lead to a significantly reduced fibrillation, even inhibiting fibrillation of native Aβ40 when added in amounts down to 1/10, whereas the more flexible beta-turn-mimics 4-amino-benzoic acid 3a and 5-amino-valeric acid 4a lead to enhanced fibrillation. Toxicity-testing of the most successful conjugate showed only minor toxicity in cell-viability assays using the N2a cell line. Structural downsizing lead to the short fragment BTD/peptide Aβ16-35 as inhibitor of the aggregation of Aβ40, opening large potential for further small peptide based inhibitors.
Collapse
Affiliation(s)
- Stefanie Deike
- Department of Chemistry, Martin Luther University Halle-Wittenberg, Von-Danckelmann-Platz 4, 06120 Halle, Germany
| | - Sven Rothemund
- Core Unit Peptid-Technologien, University Leipzig, Liebigstr. 21, 04103 Leipzig, Germany
| | - Bruno Voigt
- Department of Physics, Martin Luther University Halle-Wittenberg, Betty-Heimannstrasse 7 4, 06120 Halle, Germany
| | - Suman Samantray
- Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Forschungszentrum Jülich, 52428 Jülich, Germany
| | - Birgit Strodel
- Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Forschungszentrum Jülich, 52428 Jülich, Germany; Institute of Theoretical and Computational Chemistry, Heinrich Heine University Düsseldorf, 40225 Düsseldorf, Germany
| | - Wolfgang H Binder
- Department of Chemistry, Martin Luther University Halle-Wittenberg, Von-Danckelmann-Platz 4, 06120 Halle, Germany.
| |
Collapse
|
|
7
|
Perissinotto F, Stani C, De Cecco E, Vaccari L, Rondelli V, Posocco P, Parisse P, Scaini D, Legname G, Casalis L. Iron-mediated interaction of alpha synuclein with lipid raft model membranes. NANOSCALE 2020; 12:7631-7640. [PMID: 32104855 DOI: 10.1039/d0nr00287a] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/10/2023]
Abstract
The aberrant misfolding and aggregation of alpha synuclein (αS) into toxic oligomeric species is one of the key features associated with the pathogenesis of Parkinson's disease (PD). It involves different biochemical and biophysical factors as plasma membrane binding and interaction with heavy metal ions. In the present work, atomic force microscopy (AFM) is combined with Fourier Transform Infrared Spectroscopy (FTIR) measurements to investigate the interaction of wild-type (WT) and A53T mutated alpha synuclein with artificial lipid bilayers mimicking lipid raft (LR) domains, before and after ferrous cations (Fe2+) treatment. In the absence of iron, protein monomers produce a thinning of the membrane, targeting the non-raft phase of the bilayer preferentially. On the contrary, iron actively promotes the formation of globular protein aggregates, resembling oligomers, targeted to LR domains. In both aggregation states, monomer and oligomer, the mutated A53T protein exhibits a greater and faster membrane-interaction. These results underlie a new mechanism of membrane-protein interaction in PD. The targeting of Fe2+-promoted αS oligomers to LRs might be functional for the disease and be helpful for the development of new therapeutic strategies.
Collapse
|
|
8
|
Guerrero-Ferreira R, Taylor NMI, Arteni AA, Kumari P, Mona D, Ringler P, Britschgi M, Lauer ME, Makky A, Verasdonck J, Riek R, Melki R, Meier BH, Böckmann A, Bousset L, Stahlberg H. Two new polymorphic structures of human full-length alpha-synuclein fibrils solved by cryo-electron microscopy. eLife 2019; 8:e48907. [PMID: 31815671 PMCID: PMC6957273 DOI: 10.7554/elife.48907] [Citation(s) in RCA: 209] [Impact Index Per Article: 34.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2019] [Accepted: 11/30/2019] [Indexed: 12/19/2022] Open
Abstract
Intracellular inclusions rich in alpha-synuclein are a hallmark of several neuropathological diseases including Parkinson's disease (PD). Previously, we reported the structure of alpha-synuclein fibrils (residues 1-121), composed of two protofibrils that are connected via a densely-packed interface formed by residues 50-57 (Guerrero-Ferreira, eLife 218;7:e36402). We here report two new polymorphic atomic structures of alpha-synuclein fibrils termed polymorphs 2a and 2b, at 3.0 Å and 3.4 Å resolution, respectively. These polymorphs show a radically different structure compared to previously reported polymorphs. The new structures have a 10 nm fibril diameter and are composed of two protofilaments which interact via intermolecular salt-bridges between amino acids K45, E57 (polymorph 2a) or E46 (polymorph 2b). The non-amyloid component (NAC) region of alpha-synuclein is fully buried by previously non-described interactions with the N-terminus. A hydrophobic cleft, the location of familial PD mutation sites, and the nature of the protofilament interface now invite to formulate hypotheses about fibril formation, growth and stability.
Collapse
Affiliation(s)
- Ricardo Guerrero-Ferreira
- Center for Cellular Imaging and NanoAnalytics (C-CINA), BiozentrumUniversity of BaselBaselSwitzerland
| | - Nicholas MI Taylor
- Structural Biology of Molecular Machines Group, Protein Structure & Function Programme, Novo Nordisk Foundation Center for Protein Research, Faculty of Health and Medical SciencesUniversity of CopenhagenCopenhagenDenmark
| | - Ana-Andreea Arteni
- Institut de Biologie Intégrative de la Cellule (I2BC), CEA, CNRS, Université Paris Sud, Université Paris-SaclayGif-sur-YvetteFrance
- Institut Fancois Jacob (MIRCen), CEA and Laboratory of Neurodegenerative Diseases, CNRSFontenay-Aux-RosesFrance
| | | | - Daniel Mona
- Roche Pharma Research and Early Development, Neuroscience and Rare Diseases Discovery and Translational Medicine Area, Neuroscience DiscoveryRoche Innovation Center BaselBaselSwitzerland
| | - Philippe Ringler
- Center for Cellular Imaging and NanoAnalytics (C-CINA), BiozentrumUniversity of BaselBaselSwitzerland
| | - Markus Britschgi
- Roche Pharma Research and Early Development, Neuroscience and Rare Diseases Discovery and Translational Medicine Area, Neuroscience DiscoveryRoche Innovation Center BaselBaselSwitzerland
| | - Matthias E Lauer
- Roche Pharma Research and Early Development, Therapeutic ModalitiesRoche Innovation Center BaselBaselSwitzerland
| | - Ali Makky
- Institut Galien Paris-Sud, CNRS, Université Paris-Sud, Université Paris-SaclayChâtenay-MalabryFrance
| | | | - Roland Riek
- Laboratory of Physical ChemistryETH ZurichZurichSwitzerland
| | - Ronald Melki
- Institut Fancois Jacob (MIRCen), CEA and Laboratory of Neurodegenerative Diseases, CNRSFontenay-Aux-RosesFrance
| | - Beat H Meier
- Laboratory of Physical ChemistryETH ZurichZurichSwitzerland
| | - Anja Böckmann
- Molecular Microbiology and Structural BiochemistryLabex Ecofect, UMR 5086 CNRS, Université de LyonLyonFrance
| | - Luc Bousset
- Institut Fancois Jacob (MIRCen), CEA and Laboratory of Neurodegenerative Diseases, CNRSFontenay-Aux-RosesFrance
| | - Henning Stahlberg
- Center for Cellular Imaging and NanoAnalytics (C-CINA), BiozentrumUniversity of BaselBaselSwitzerland
| |
Collapse
|
|
9
|
Huang YW, King CY. A complete catalog of wild-type Sup35 prion variants and their protein-only propagation. Curr Genet 2019; 66:97-122. [DOI: 10.1007/s00294-019-01003-8] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2019] [Revised: 05/28/2019] [Accepted: 06/01/2019] [Indexed: 10/26/2022]
|
|
10
|
Perez A, Gaalswyk K, Jaroniec CP, MacCallum JL. High Accuracy Protein Structures from Minimal Sparse Paramagnetic Solid‐State NMR Restraints. Angew Chem Int Ed Engl 2019. [DOI: 10.1002/ange.201811895] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Affiliation(s)
- Alberto Perez
- Department of Chemistry University of Florida Gainesville FL USA
| | - Kari Gaalswyk
- Department of Chemistry University of Calgary Calgary Alberta Canada
| | | | | |
Collapse
|
|
11
|
Götzke L, Schaper G, März J, Kaden P, Huittinen N, Stumpf T, Kammerlander KK, Brunner E, Hahn P, Mehnert A, Kersting B, Henle T, Lindoy LF, Zanoni G, Weigand JJ. Coordination chemistry of f-block metal ions with ligands bearing bio-relevant functional groups. Coord Chem Rev 2019. [DOI: 10.1016/j.ccr.2019.01.006] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
|
|
12
|
Perez A, Gaalswyk K, Jaroniec CP, MacCallum JL. High Accuracy Protein Structures from Minimal Sparse Paramagnetic Solid-State NMR Restraints. Angew Chem Int Ed Engl 2019; 58:6564-6568. [PMID: 30913341 DOI: 10.1002/anie.201811895] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2018] [Revised: 02/01/2019] [Indexed: 11/08/2022]
Abstract
There is a pressing need for new computational tools to integrate data from diverse experimental approaches in structural biology. We present a strategy that combines sparse paramagnetic solid-state NMR restraints with physics-based atomistic simulations. Our approach explicitly accounts for uncertainty in the interpretation of experimental data through the use of a semi-quantitative mapping between the data and the restraint energy that is calibrated by extensive simulations. We apply our approach to solid-state NMR data for the model protein GB1 labeled with Cu2+ -EDTA at six different sites. We are able to determine the structure to 0.9 Å accuracy within a single day of computation on a GPU cluster. We further show that in some cases, the data from only a single paramagnetic tag are sufficient for accurate folding.
Collapse
Affiliation(s)
- Alberto Perez
- Department of Chemistry, University of Florida, Gainesville, FL, USA
| | - Kari Gaalswyk
- Department of Chemistry, University of Calgary, Calgary, Alberta, Canada
| | | | - Justin L MacCallum
- Department of Chemistry, University of Calgary, Calgary, Alberta, Canada
| |
Collapse
|
|
13
|
Conservation of the Amyloid Interactome Across Diverse Fibrillar Structures. Sci Rep 2019; 9:3863. [PMID: 30846764 PMCID: PMC6405930 DOI: 10.1038/s41598-019-40483-z] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2018] [Accepted: 11/23/2018] [Indexed: 12/30/2022] Open
Abstract
Several human proteins cause disease by misfolding and aggregating into amyloid fibril deposits affecting the surrounding tissues. Multiple other proteins co-associate with the diseased deposits but little is known about how this association is influenced by the nature of the amyloid aggregate and the properties of the amyloid-forming protein. In this study, we investigated the co-aggregation of plasma and cerebrospinal proteins in the presence of pre-formed amyloid fibrils. We evaluated the fibril-associated proteome across multiple amyloid fibril types that differ in their amino acid sequences, ultrastructural morphologies, and recognition by amyloid-binding dyes. The fibril types included aggregates formed by Amyloid β, α-synuclein, and FAS4 that are associated with pathological disorders, and aggregates formed by the glucagon and C-36 peptides, currently not linked to any human disease. Our results highlighted a highly similar response to the amyloid fold within the body fluid of interest. Fibrils with diverse primary sequences and ultrastructural morphologies only differed slightly in the composition of the co-aggregated proteins but were clearly distinct from less fibrillar and amorphous aggregates. The type of body fluid greatly affected the resulting amyloid interactome, underlining the role of the in vivo environment. We conclude that protein fibrils lead to a specific response in protein co-aggregation and discuss the effects hereof in the context of amyloid deposition.
Collapse
|
|
14
|
Mukhopadhyay D, Gupta C, Theint T, Jaroniec CP. Peptide bond conformation in peptides and proteins probed by dipolar coupling-chemical shift tensor correlation solid-state NMR. JOURNAL OF MAGNETIC RESONANCE (SAN DIEGO, CALIF. : 1997) 2018; 297:152-160. [PMID: 30396157 PMCID: PMC6289736 DOI: 10.1016/j.jmr.2018.10.015] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/29/2018] [Revised: 10/25/2018] [Accepted: 10/26/2018] [Indexed: 05/30/2023]
Abstract
Multidimensional magic-angle spinning solid-state NMR experiments are described that permit cis and trans peptide bonds in uniformly 13C,15N-labeled peptides and proteins to be unambiguously distinguished in residue-specific manner by determining the relative orientations of the amide 13C' CSA and 1H-15N dipolar coupling tensors. The experiments are demonstrated for model peptides glycylglycine and 2,5-diketopiperazine containing trans and cis peptide bonds, respectively. Subsequently, the measurements are extended to two representative proteins that contain exclusively trans peptide bonds, microcrystalline B3 immunoglobulin domain of protein G and Y145Stop human prion protein amyloid fibrils, to illustrate their applicability to a wide range of protein systems.
Collapse
Affiliation(s)
- Dwaipayan Mukhopadhyay
- Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, United States
| | - Chitrak Gupta
- Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, United States
| | - Theint Theint
- Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, United States
| | - Christopher P Jaroniec
- Department of Chemistry and Biochemistry, The Ohio State University, Columbus, OH 43210, United States.
| |
Collapse
|
|
15
|
Ghosh C, Harmouche N, Bechinger B, Haldar J. Aryl-Alkyl-Lysines Interact with Anionic Lipid Components of Bacterial Cell Envelope Eliciting Anti-Inflammatory and Antibiofilm Properties. ACS OMEGA 2018; 3:9182-9190. [PMID: 31459052 PMCID: PMC6645134 DOI: 10.1021/acsomega.8b01052] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/18/2018] [Accepted: 07/30/2018] [Indexed: 05/05/2023]
Abstract
The emergence of bacterial resistance and hesitance in approving new drugs has bolstered research on membrane-active agents such as antimicrobial peptides and their synthetic derivatives as therapeutic alternatives against bacterial infections. Herein, we document the action of aryl-alkyl-lysines on liposomes mimicking bacterial membranes using solid-state nuclear magnetic resonance spectroscopy. A significant perturbation of the lipid thickness and order parameter of the lipid membrane was observed upon treatment with this class of compounds. Encouraged by these results, the ability of the most active compound (NCK-10) to interact with aggregates of lipopolysaccharides (LPSs) was studied. In vitro experiments showed that NCK-10 was able to prevent the LPS-induced stimulation of proinflammatory cytokines such as tumor necrosis factor-α and interleukin-6. The compound could also disrupt the biofilms of Pseudomonas aeruginosa in vitro and bring down the bacterial burden by more than 99% in a mice model of burn infections caused by the biofilms of P. aeruginosa.
Collapse
Affiliation(s)
- Chandradhish Ghosh
- Antimicrobial Research
Laboratory, New Chemistry Unit, Jawaharlal
Nehru Centre for Advanced Scientific Research, Jakkur, Bengaluru 560064, Karnataka, India
| | - Nicole Harmouche
- Université
de Strasbourg/CNRS, UMR7177, Institut de Chimie, 67008 Strasbourg, France
| | - Burkhard Bechinger
- Université
de Strasbourg/CNRS, UMR7177, Institut de Chimie, 67008 Strasbourg, France
| | - Jayanta Haldar
- Antimicrobial Research
Laboratory, New Chemistry Unit, Jawaharlal
Nehru Centre for Advanced Scientific Research, Jakkur, Bengaluru 560064, Karnataka, India
- E-mail: (J.H.)
| |
Collapse
|
|
16
|
Kell DB, Pretorius E. No effects without causes: the Iron Dysregulation and Dormant Microbes hypothesis for chronic, inflammatory diseases. Biol Rev Camb Philos Soc 2018; 93:1518-1557. [PMID: 29575574 PMCID: PMC6055827 DOI: 10.1111/brv.12407] [Citation(s) in RCA: 70] [Impact Index Per Article: 10.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2017] [Revised: 02/12/2018] [Accepted: 02/15/2018] [Indexed: 12/11/2022]
Abstract
Since the successful conquest of many acute, communicable (infectious) diseases through the use of vaccines and antibiotics, the currently most prevalent diseases are chronic and progressive in nature, and are all accompanied by inflammation. These diseases include neurodegenerative (e.g. Alzheimer's, Parkinson's), vascular (e.g. atherosclerosis, pre-eclampsia, type 2 diabetes) and autoimmune (e.g. rheumatoid arthritis and multiple sclerosis) diseases that may appear to have little in common. In fact they all share significant features, in particular chronic inflammation and its attendant inflammatory cytokines. Such effects do not happen without underlying and initially 'external' causes, and it is of interest to seek these causes. Taking a systems approach, we argue that these causes include (i) stress-induced iron dysregulation, and (ii) its ability to awaken dormant, non-replicating microbes with which the host has become infected. Other external causes may be dietary. Such microbes are capable of shedding small, but functionally significant amounts of highly inflammagenic molecules such as lipopolysaccharide and lipoteichoic acid. Sequelae include significant coagulopathies, not least the recently discovered amyloidogenic clotting of blood, leading to cell death and the release of further inflammagens. The extensive evidence discussed here implies, as was found with ulcers, that almost all chronic, infectious diseases do in fact harbour a microbial component. What differs is simply the microbes and the anatomical location from and at which they exert damage. This analysis offers novel avenues for diagnosis and treatment.
Collapse
Affiliation(s)
- Douglas B. Kell
- School of ChemistryThe University of Manchester, 131 Princess StreetManchesterLancsM1 7DNU.K.
- The Manchester Institute of BiotechnologyThe University of Manchester, 131 Princess StreetManchesterLancsM1 7DNU.K.
- Department of Physiological SciencesStellenbosch University, Stellenbosch Private Bag X1Matieland7602South Africa
| | - Etheresia Pretorius
- Department of Physiological SciencesStellenbosch University, Stellenbosch Private Bag X1Matieland7602South Africa
| |
Collapse
|
|
17
|
Hung NB, Le DM, Hoang TX. Sequence dependent aggregation of peptides and fibril formation. J Chem Phys 2018; 147:105102. [PMID: 28915764 DOI: 10.1063/1.5001517] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/25/2023] Open
Abstract
Deciphering the links between amino acid sequence and amyloid fibril formation is key for understanding protein misfolding diseases. Here we use Monte Carlo simulations to study the aggregation of short peptides in a coarse-grained model with hydrophobic-polar (HP) amino acid sequences and correlated side chain orientations for hydrophobic contacts. A significant heterogeneity is observed in the aggregate structures and in the thermodynamics of aggregation for systems of different HP sequences and different numbers of peptides. Fibril-like ordered aggregates are found for several sequences that contain the common HPH pattern, while other sequences may form helix bundles or disordered aggregates. A wide variation of the aggregation transition temperatures among sequences, even among those of the same hydrophobic fraction, indicates that not all sequences undergo aggregation at a presumable physiological temperature. The transition is found to be the most cooperative for sequences forming fibril-like structures. For a fibril-prone sequence, it is shown that fibril formation follows the nucleation and growth mechanism. Interestingly, a binary mixture of peptides of an aggregation-prone and a non-aggregation-prone sequence shows the association and conversion of the latter to the fibrillar structure. Our study highlights the role of a sequence in selecting fibril-like aggregates and also the impact of a structural template on fibril formation by peptides of unrelated sequences.
Collapse
Affiliation(s)
- Nguyen Ba Hung
- Institute of Physics, Vietnam Academy of Science and Technology, 10 Dao Tan, Ba Dinh, Hanoi, Vietnam
| | - Duy-Manh Le
- Institute of Research and Development, Duy Tan University, K7/25 Quang Trung, Da Nang, Vietnam
| | - Trinh X Hoang
- Institute of Physics, Vietnam Academy of Science and Technology, 10 Dao Tan, Ba Dinh, Hanoi, Vietnam
| |
Collapse
|
|
18
|
Femtosecond X-ray coherent diffraction of aligned amyloid fibrils on low background graphene. Nat Commun 2018; 9:1836. [PMID: 29743480 PMCID: PMC5943278 DOI: 10.1038/s41467-018-04116-9] [Citation(s) in RCA: 25] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2017] [Accepted: 04/03/2018] [Indexed: 11/09/2022] Open
Abstract
Here we present a new approach to diffraction imaging of amyloid fibrils, combining a free-standing graphene support and single nanofocused X-ray pulses of femtosecond duration from an X-ray free-electron laser. Due to the very low background scattering from the graphene support and mutual alignment of filaments, diffraction from tobacco mosaic virus (TMV) filaments and amyloid protofibrils is obtained to 2.7 Å and 2.4 Å resolution in single diffraction patterns, respectively. Some TMV diffraction patterns exhibit asymmetry that indicates the presence of a limited number of axial rotations in the XFEL focus. Signal-to-noise levels from individual diffraction patterns are enhanced using computational alignment and merging, giving patterns that are superior to those obtainable from synchrotron radiation sources. We anticipate that our approach will be a starting point for further investigations into unsolved structures of filaments and other weakly scattering objects.
Collapse
|
|
19
|
Guo ZH, Yang CI, Ho CI, Huang SJ, Chen YC, Tai HC, Chan JCC. Fibrillization of β-Amyloid Peptides via Chemically Modulated Pathway. Chemistry 2018; 24:4939-4943. [PMID: 29380450 DOI: 10.1002/chem.201706001] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2017] [Indexed: 11/11/2022]
Abstract
The aggregation of β-amyloid peptides is closely associated with Alzheimer's disease. We have used liposomes to modulate the early aggregation events of 40-residue β-amyloid peptides. The spatial confinement provided by liposomes leads to the formation of nonfibrillar aggregates of β-amyloid peptides. These on-pathway β-sheet intermediates were used to seed the fibrillization of the monomer peptides. Solid-state NMR spectroscopy revealed that the resultant fibrils have a more uniform structure than those formed in liposome-free solution.
Collapse
Affiliation(s)
- Zhong-Hong Guo
- Department of Chemistry, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| | - Chien-I Yang
- Department of Chemistry, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| | - Cheng-I Ho
- Department of Chemistry, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| | - Shing-Jong Huang
- Instrumentation Center, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| | - Yin-Chung Chen
- Department of Chemistry, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| | - Hwan-Ching Tai
- Department of Chemistry, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| | - Jerry Chun Chung Chan
- Department of Chemistry, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei, 10617, Taiwan
| |
Collapse
|
|
20
|
3D structure determination of amyloid fibrils using solid-state NMR spectroscopy. Methods 2018; 138-139:26-38. [DOI: 10.1016/j.ymeth.2018.03.014] [Citation(s) in RCA: 53] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2018] [Revised: 03/28/2018] [Accepted: 03/30/2018] [Indexed: 01/08/2023] Open
|
|
21
|
Schütz AK, Hornemann S, Wälti MA, Greuter L, Tiberi C, Cadalbert R, Gantner M, Riek R, Hammarström P, Nilsson KPR, Böckmann A, Aguzzi A, Meier BH. Binding of Polythiophenes to Amyloids: Structural Mapping of the Pharmacophore. ACS Chem Neurosci 2018; 9:475-481. [PMID: 29178774 DOI: 10.1021/acschemneuro.7b00397] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
Luminescent conjugated polythiophenes bind to amyloid proteins with high affinity. Their fluorescence properties, which are modulated by the detailed conformation in the bound state, are highly sensitive to structural features of the amyloid. Polythiophenes therefore represent diagnostic markers for the detection and differentiation of pathological amyloid aggregates. We clarify the binding site and mode of two different polythiophenes to fibrils of the prion domain of the HET-s protein by solid-state NMR and correlate these findings with their fluorescence properties. We demonstrate how amyloid dyes recognize distinct binding sites with specific topological features. Regularly spaced surface charge patterns and well-accessible grooves on the fibril surface define the pharmacophore of the amyloid, which in turn determines the binding mode and fluorescence wavelength of the polythiophene.
Collapse
Affiliation(s)
- Anne K. Schütz
- Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland
| | - Simone Hornemann
- Institute of Neuropathology, University Hospital of Zurich, University of Zürich, Schmelzbergstrasse 12, 8091 Zürich, Switzerland
| | - Marielle A. Wälti
- Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland
| | - Ladina Greuter
- Institute of Neuropathology, University Hospital of Zurich, University of Zürich, Schmelzbergstrasse 12, 8091 Zürich, Switzerland
| | - Cinzia Tiberi
- Institute of Neuropathology, University Hospital of Zurich, University of Zürich, Schmelzbergstrasse 12, 8091 Zürich, Switzerland
| | - Riccardo Cadalbert
- Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland
| | - Matthias Gantner
- Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland
| | - Roland Riek
- Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland
| | - Per Hammarström
- Department of Physics, Chemistry and Biology (IFM), Linköping University, 58183 Linköping, Sweden
| | - K. Peter R. Nilsson
- Department of Physics, Chemistry and Biology (IFM), Linköping University, 58183 Linköping, Sweden
| | - Anja Böckmann
- Molecular Microbiology and Structural Biochemistry, UMR 5086 CNRS/Université de Lyon 1, 7 passage du Vercors, 69367 Lyon, France
| | - Adriano Aguzzi
- Institute of Neuropathology, University Hospital of Zurich, University of Zürich, Schmelzbergstrasse 12, 8091 Zürich, Switzerland
| | - Beat H. Meier
- Physical Chemistry, ETH Zurich, Vladimir-Prelog-Weg 2, 8093 Zurich, Switzerland
| |
Collapse
|
|
22
|
Jiang Z, Flynn JD, Teague WE, Gawrisch K, Lee JC. Stimulation of α-synuclein amyloid formation by phosphatidylglycerol micellar tubules. BIOCHIMICA ET BIOPHYSICA ACTA-BIOMEMBRANES 2018; 1860:1840-1847. [PMID: 29501608 DOI: 10.1016/j.bbamem.2018.02.025] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/03/2018] [Revised: 02/23/2018] [Accepted: 02/25/2018] [Indexed: 10/25/2022]
Abstract
α-Synuclein (α-Syn) is a presynaptic protein that is accumulated in its amyloid form in the brains of Parkinson's patients. Although its biological function remains unclear, α-syn has been suggested to bind to synaptic vesicles and facilitate neurotransmitter release. Recently, studies have found that α-syn induces membrane tubulation, highlighting a potential mechanism for α-syn to stabilize highly curved membrane structures which could have both functional and dysfunctional consequences. To understand how membrane remodeling by α-syn affects amyloid formation, we have studied the α-syn aggregation process in the presence of phosphatidylglycerol (PG) micellar tubules, which were the first reported example of membrane tubulation by α-syn. Aggregation kinetics, β-sheet content, and macroscopic protein-lipid structures were observed by Thioflavin T fluorescence, circular dichroism spectroscopy and transmission electron microscopy, respectively. Collectively, the presence of PG micellar tubules formed at a stochiometric (L/P = 1) ratio was found to stimulate α-syn fibril formation. Moreover, transmission electron microscopy and solid-state nuclear magnetic resonance spectroscopy revealed the co-assembly of PG and α-syn into fibril structures. However, isolated micellar tubules do not form fibrils by themselves, suggesting an important role of free α-syn monomers during amyloid formation. In contrast, fibrils did not form in the presence of excess PG lipids (≥L/P = 50), where most of the α-syn molecules are in a membrane-bound α-helical form. Our results provide new mechanistic insights into how membrane tubules modulate α-syn amyloid formation and support a pivotal role of protein-lipid interaction in the dysfunction of α-syn.
Collapse
Affiliation(s)
- Zhiping Jiang
- Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
| | - Jessica D Flynn
- Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA
| | - Walter E Teague
- Laboratory of Membrane Biochemistry and Biophysics, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892, USA
| | - Klaus Gawrisch
- Laboratory of Membrane Biochemistry and Biophysics, National Institute on Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, MD 20892, USA
| | - Jennifer C Lee
- Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD 20892, USA.
| |
Collapse
|
|
23
|
Tolchard J, Pandey MK, Berbon M, Noubhani A, Saupe SJ, Nishiyama Y, Habenstein B, Loquet A. Detection of side-chain proton resonances of fully protonated biosolids in nano-litre volumes by magic angle spinning solid-state NMR. JOURNAL OF BIOMOLECULAR NMR 2018; 70:177-185. [PMID: 29502224 DOI: 10.1007/s10858-018-0168-3] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/20/2018] [Accepted: 02/16/2018] [Indexed: 06/08/2023]
Abstract
We present a new solid-state NMR proton-detected three-dimensional experiment dedicated to the observation of protein proton side chain resonances in nano-liter volumes. The experiment takes advantage of very fast magic angle spinning and double quantum 13C-13C transfer to establish efficient (H)CCH correlations detected on side chain protons. Our approach is demonstrated on the HET-s prion domain in its functional amyloid fibrillar form, fully protonated, with a sample amount of less than 500 µg using a MAS frequency of 70 kHz. The majority of aliphatic and aromatic side chain protons (70%) are observable, in addition to Hα resonances, in a single experiment providing a complementary approach to the established proton-detected amide-based multidimensional solid-state NMR experiments for the study and resonance assignment of biosolid samples, in particular for aromatic side chain resonances.
Collapse
Affiliation(s)
- James Tolchard
- Institute of Chemistry & Biology of Membranes & Nanoobjects, (UMR5248 CBMN), CNRS, Université Bordeaux, Institut Européen de Chimie et Biologie, 33600, Pessac, France
| | - Manoj Kumar Pandey
- JEOL RESONANCE Inc., Musashino, Akishima, Tokyo, 196-8558, Japan
- RIKEN CLST-JEOL Collaboration Center, Tsurumi, Yokohama, Kanagawa, 230-0045, Japan
- Department of Chemistry, Indian Institute of Technology Ropar, Rupnagar, India
| | - Mélanie Berbon
- Institute of Chemistry & Biology of Membranes & Nanoobjects, (UMR5248 CBMN), CNRS, Université Bordeaux, Institut Européen de Chimie et Biologie, 33600, Pessac, France
| | - Abdelmajid Noubhani
- Institute of Chemistry & Biology of Membranes & Nanoobjects, (UMR5248 CBMN), CNRS, Université Bordeaux, Institut Européen de Chimie et Biologie, 33600, Pessac, France
| | - Sven J Saupe
- Institut de Biochimie et de Génétique Cellulaire, (UMR 5095 IBGC), CNRS, Université Bordeaux, 33077, Bordeaux, France
| | - Yusuke Nishiyama
- JEOL RESONANCE Inc., Musashino, Akishima, Tokyo, 196-8558, Japan.
- RIKEN CLST-JEOL Collaboration Center, Tsurumi, Yokohama, Kanagawa, 230-0045, Japan.
| | - Birgit Habenstein
- Institute of Chemistry & Biology of Membranes & Nanoobjects, (UMR5248 CBMN), CNRS, Université Bordeaux, Institut Européen de Chimie et Biologie, 33600, Pessac, France.
| | - Antoine Loquet
- Institute of Chemistry & Biology of Membranes & Nanoobjects, (UMR5248 CBMN), CNRS, Université Bordeaux, Institut Européen de Chimie et Biologie, 33600, Pessac, France.
| |
Collapse
|
|
24
|
Martinez D, Legrand A, Gronnier J, Decossas M, Gouguet P, Lambert O, Berbon M, Verron L, Grélard A, Germain V, Loquet A, Mongrand S, Habenstein B. Coiled-coil oligomerization controls localization of the plasma membrane REMORINs. J Struct Biol 2018; 206:12-19. [PMID: 29481850 DOI: 10.1016/j.jsb.2018.02.003] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2017] [Revised: 01/25/2018] [Accepted: 02/12/2018] [Indexed: 12/20/2022]
Abstract
REMORINs are nanodomain-organized proteins located in the plasma membrane and involved in cellular responses in plants. The dynamic assembly of the membrane nanodomains represents an essential tool of the versatile membrane barriers to control and modulate cellular functions. Nevertheless, the assembly mechanisms and protein organization strategies of nanodomains are poorly understood and many structural aspects are difficult to visualize. Using an ensemble of biophysical approaches, including solid-state nuclear magnetic resonance, cryo-electron microscopy and in vivo confocal imaging, we provide first insights on the role and the structural mechanisms of REMORIN trimerization. Our results suggest that the formation of REMORIN coiled-coil trimers is essential for membrane recruitment and promotes REMORIN assembly in vitro into long filaments by trimer-trimer interactions that might participate in nanoclustering into membrane domains in vivo.
Collapse
Affiliation(s)
- Denis Martinez
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Anthony Legrand
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Julien Gronnier
- Laboratoire de Biogènese Membranaire - UMR 5200 - CNRS, Université de Bordeaux, 71 Avenue Edouard Bourlaux, 33883 Villenave d'Ornon Cédex, France
| | - Marion Decossas
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, 14 All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Paul Gouguet
- Laboratoire de Biogènese Membranaire - UMR 5200 - CNRS, Université de Bordeaux, 71 Avenue Edouard Bourlaux, 33883 Villenave d'Ornon Cédex, France
| | - Olivier Lambert
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, 14 All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Mélanie Berbon
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Loris Verron
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Axelle Grélard
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France
| | - Veronique Germain
- Laboratoire de Biogènese Membranaire - UMR 5200 - CNRS, Université de Bordeaux, 71 Avenue Edouard Bourlaux, 33883 Villenave d'Ornon Cédex, France
| | - Antoine Loquet
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France.
| | - Sébastien Mongrand
- Laboratoire de Biogènese Membranaire - UMR 5200 - CNRS, Université de Bordeaux, 71 Avenue Edouard Bourlaux, 33883 Villenave d'Ornon Cédex, France.
| | - Birgit Habenstein
- Institute of Chemistry & Biology of Membranes & Nanoobjects (UMR5248 CBMN), IECB, CNRS, Universite Bordeaux, Institut Polytechnique Bordeaux, All. Geoffroy Saint-Hilaire, 33600 Pessac, France.
| |
Collapse
|
|
25
|
Kulawik A, Heise H, Zafiu C, Willbold D, Bannach O. Advancements of the
sFIDA
method for oligomer‐based diagnostics of neurodegenerative diseases. FEBS Lett 2018; 592:516-534. [DOI: 10.1002/1873-3468.12983] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2017] [Revised: 01/11/2018] [Accepted: 01/16/2018] [Indexed: 01/11/2023]
Affiliation(s)
- Andreas Kulawik
- Institute of Complex Systems (ICS‐6: Structural Biochemistry) Forschungszentrum Jülich Germany
- Institut für Physikalische Biologie Heinrich‐Heine‐Universität Düsseldorf Germany
| | - Henrike Heise
- Institute of Complex Systems (ICS‐6: Structural Biochemistry) Forschungszentrum Jülich Germany
- Institut für Physikalische Biologie Heinrich‐Heine‐Universität Düsseldorf Germany
| | - Christian Zafiu
- Institute of Complex Systems (ICS‐6: Structural Biochemistry) Forschungszentrum Jülich Germany
| | - Dieter Willbold
- Institute of Complex Systems (ICS‐6: Structural Biochemistry) Forschungszentrum Jülich Germany
- Institut für Physikalische Biologie Heinrich‐Heine‐Universität Düsseldorf Germany
| | - Oliver Bannach
- Institute of Complex Systems (ICS‐6: Structural Biochemistry) Forschungszentrum Jülich Germany
- Institut für Physikalische Biologie Heinrich‐Heine‐Universität Düsseldorf Germany
| |
Collapse
|
|
26
|
Mukhopadhyay D, Nadaud PS, Shannon MD, Jaroniec CP. Rapid Quantitative Measurements of Paramagnetic Relaxation Enhancements in Cu(II)-Tagged Proteins by Proton-Detected Solid-State NMR Spectroscopy. J Phys Chem Lett 2017; 8:5871-5877. [PMID: 29148785 PMCID: PMC5720925 DOI: 10.1021/acs.jpclett.7b02709] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 05/24/2023]
Abstract
We demonstrate rapid quantitative measurements of site-resolved paramagnetic relaxation enhancements (PREs), which are a source of valuable structural restraints corresponding to electron-nucleus distances in the ∼10-20 Å regime, in solid-state nuclear magnetic resonance (NMR) spectra of proteins containing covalent Cu2+-binding tags. Specifically, using protein GB1 K28C-EDTA-Cu2+ mutant as a model, we show the determination of backbone amide 15N longitudinal and 1H transverse PREs within a few hours of experiment time based on proton-detected 2D or 3D correlation spectra recorded with magic-angle spinning frequencies ≥ ∼ 60 kHz for samples containing ∼10-50 nanomoles of 2H,13C,15N-labeled protein back-exchanged in H2O. Additionally, we show that the electron relaxation time for the Cu2+ center, needed to convert PREs into distances, can be estimated directly from the experimental data. Altogether, these results are important for establishing solid-state NMR based on paramagnetic-tagging as a routine tool for structure determination of natively diamagnetic proteins.
Collapse
|
|
27
|
Species-dependent structural polymorphism of Y145Stop prion protein amyloid revealed by solid-state NMR spectroscopy. Nat Commun 2017; 8:753. [PMID: 28963458 PMCID: PMC5622040 DOI: 10.1038/s41467-017-00794-z] [Citation(s) in RCA: 56] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2016] [Accepted: 07/28/2017] [Indexed: 11/12/2022] Open
Abstract
One of the most puzzling aspects of the prion diseases is the intricate relationship between prion strains and interspecies transmissibility barriers. Previously we have shown that certain fundamental aspects of mammalian prion propagation, including the strain phenomenon and species barriers, can be reproduced in vitro in seeded fibrillization of the Y145Stop prion protein variant. Here, we use solid-state nuclear magnetic resonance spectroscopy to gain atomic level insight into the structural differences between Y145Stop prion protein amyloids from three species: human, mouse, and Syrian hamster. Remarkably, we find that these structural differences are largely controlled by only two amino acids at positions 112 and 139, and that the same residues appear to be key to the emergence of structurally distinct amyloid strains within the same protein sequence. The role of these residues as conformational switches can be rationalized based on a model for human Y145Stop prion protein amyloid, providing a foundation for understanding cross-seeding specificity. Prion diseases can be transmitted across species. Here the authors use solid-state NMR to study prion protein (PrP) amyloids from human, mouse and Syrian hamster and show that their structural differences are mainly governed by two residues, which helps to understand interspecies PrP propagation on a molecular level.
Collapse
|
|
28
|
Loquet A, Tolchard J, Berbon M, Martinez D, Habenstein B. Atomic Scale Structural Studies of Macromolecular Assemblies by Solid-state Nuclear Magnetic Resonance Spectroscopy. J Vis Exp 2017:55779. [PMID: 28994783 PMCID: PMC5752270 DOI: 10.3791/55779] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023] Open
Abstract
Supramolecular protein assemblies play fundamental roles in biological processes ranging from host-pathogen interaction, viral infection to the propagation of neurodegenerative disorders. Such assemblies consist in multiple protein subunits organized in a non-covalent way to form large macromolecular objects that can execute a variety of cellular functions or cause detrimental consequences. Atomic insights into the assembly mechanisms and the functioning of those macromolecular assemblies remain often scarce since their inherent insolubility and non-crystallinity often drastically reduces the quality of the data obtained from most techniques used in structural biology, such as X-ray crystallography and solution Nuclear Magnetic Resonance (NMR). We here present magic-angle spinning solid-state NMR spectroscopy (SSNMR) as a powerful method to investigate structures of macromolecular assemblies at atomic resolution. SSNMR can reveal atomic details on the assembled complex without size and solubility limitations. The protocol presented here describes the essential steps from the production of 13C/15N isotope-labeled macromolecular protein assemblies to the acquisition of standard SSNMR spectra and their analysis and interpretation. As an example, we show the pipeline of a SSNMR structural analysis of a filamentous protein assembly.
Collapse
Affiliation(s)
- Antoine Loquet
- Institute of Chemistry, Biology of Membranes, Nanoobjects, UMR5248 CNRS, Université de Bordeaux;
| | - James Tolchard
- Institute of Chemistry, Biology of Membranes, Nanoobjects, UMR5248 CNRS, Université de Bordeaux
| | - Melanie Berbon
- Institute of Chemistry, Biology of Membranes, Nanoobjects, UMR5248 CNRS, Université de Bordeaux
| | - Denis Martinez
- Institute of Chemistry, Biology of Membranes, Nanoobjects, UMR5248 CNRS, Université de Bordeaux
| | - Birgit Habenstein
- Institute of Chemistry, Biology of Membranes, Nanoobjects, UMR5248 CNRS, Université de Bordeaux;
| |
Collapse
|
|
29
|
Bacterial Filamentous Appendages Investigated by Solid-State NMR Spectroscopy. Methods Mol Biol 2017. [PMID: 28667627 DOI: 10.1007/978-1-4939-7033-9_29] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register]
Abstract
The assembly of filamentous appendages at the surface of bacteria is essential in many infection mechanisms. The extent of mechanical, dynamical, and functional properties of such appendages is very diverse, ranging from a structural scaffold of the pathogen-host cell interaction to cell motility, surface adhesion, or the export of virulence effectors. In particular, the architectures of several bacterial secretion systems have revealed the presence of filamentous architectures, known as pili, fimbriae, andneedles. At the macroscopic level, filamentous bacterial appendages appear as thin extracellular filaments of several nanometers in diameter and up to several microns in length. The structural characterization of these appendages at atomic-scale resolution represents an extremely challenging task because of their inherent noncrystallinity and very poor solubility. Here, we describe protocols based on recent advances in solid-state NMR spectroscopy to investigate the secondary structure, subunit-subunit protein interactions, symmetry parameters, and atomic architecture of bacterial filaments.
Collapse
|
|
30
|
Hewetson A, Do HQ, Myers C, Muthusubramanian A, Sutton RB, Wylie BJ, Cornwall GA. Functional Amyloids in Reproduction. Biomolecules 2017; 7:biom7030046. [PMID: 28661450 PMCID: PMC5618227 DOI: 10.3390/biom7030046] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/06/2017] [Revised: 06/20/2017] [Accepted: 06/23/2017] [Indexed: 11/16/2022] Open
Abstract
Amyloids are traditionally considered pathological protein aggregates that play causative roles in neurodegenerative disease, diabetes and prionopathies. However, increasing evidence indicates that in many biological systems nonpathological amyloids are formed for functional purposes. In this review, we will specifically describe amyloids that carry out biological roles in sexual reproduction including the processes of gametogenesis, germline specification, sperm maturation and fertilization. Several of these functional amyloids are evolutionarily conserved across several taxa, including human, emphasizing the critical role amyloids perform in reproduction. Evidence will also be presented suggesting that, if altered, some functional amyloids may become pathological.
Collapse
Affiliation(s)
- Aveline Hewetson
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
| | - Hoa Quynh Do
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
| | - Caitlyn Myers
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
| | - Archana Muthusubramanian
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
| | - Roger Bryan Sutton
- Department of Cell Physiology and Molecular Biophysics, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
| | - Benjamin J Wylie
- Department of Chemistry, Texas Tech University, Lubbock, TX 79409, USA.
| | - Gail A Cornwall
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Center, Lubbock, TX 79430, USA.
| |
Collapse
|
|
31
|
McGlinchey RP, Dominah GA, Lee JC. Taking a Bite Out of Amyloid: Mechanistic Insights into α-Synuclein Degradation by Cathepsin L. Biochemistry 2017; 56:3881-3884. [PMID: 28614652 DOI: 10.1021/acs.biochem.7b00360] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
A common hallmark of amyloids is their resistance to an array of proteases, highlighting the difficulty in degrading these disease-related aggregated proteinaceous materials. Here, we report on the potent activity of cathepsin L (CtsL), a lysosomal protease that proteolyzes the Parkinson's disease-related amyloid formed by α-synuclein (α-syn). Using liquid chromatography with mass spectrometry and transmission electron microscopy, an elegant mechanism is revealed on the residue and ultrastructural level, respectively. Specifically, CtsL always truncates α-syn fibrils first at the C-terminus before attacking the internal β-sheet-rich region between residues 30 and 100. This suggests that only upon removal of the α-syn C-terminus can CtsL gain access to residues within the amyloid core. Interestingly, three of the four mapped sites contain a glycine residue (G36, G41, and G51) that is likely to be involved in a β-turn in the fibril, whereupon cutting would lead to solvent exposure of internal residues and allow further proteolysis. Via close inspection of the fibril morphology, products resulting from CtsL degradation show imperfections along the fibril axis, with missing protein density as though they have been cannibalized. The ability of CtsL to degrade α-syn amyloid fibrils offers a promising strategy for improving the cellular clearance of aggregated α-syn through the modulation of protease levels and activity.
Collapse
Affiliation(s)
- Ryan P McGlinchey
- Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health , Bethesda, Maryland 20892, United States
| | - Gifty A Dominah
- Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health , Bethesda, Maryland 20892, United States
| | - Jennifer C Lee
- Laboratory of Protein Conformation and Dynamics, Biochemistry and Biophysics Center, National Heart, Lung, and Blood Institute, National Institutes of Health , Bethesda, Maryland 20892, United States
| |
Collapse
|
|
32
|
Wolff M, Zhang-Haagen B, Decker C, Barz B, Schneider M, Biehl R, Radulescu A, Strodel B, Willbold D, Nagel-Steger L. Aβ42 pentamers/hexamers are the smallest detectable oligomers in solution. Sci Rep 2017; 7:2493. [PMID: 28559586 PMCID: PMC5449387 DOI: 10.1038/s41598-017-02370-3] [Citation(s) in RCA: 40] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2016] [Accepted: 04/11/2017] [Indexed: 12/14/2022] Open
Abstract
Amyloid β (Aβ) oligomers may play a decisive role in Alzheimer's disease related neurodegeneration, but their structural properties are poorly understood. In this report, sedimentation velocity centrifugation, small angle neutron scattering (SANS) and molecular modelling were used to identify the small oligomeric species formed by the 42 amino acid residue long isoform of Aβ (Aβ42) in solution, characterized by a sedimentation coefficient of 2.56 S, and a radius of gyration between 2 and 4 nm. The measured sedimentation coefficient is in close agreement with the sedimentation coefficient calculated for Aβ42 hexamers using MD simulations at µM concentration. To the best of our knowledge this is the first report detailing the Aβ42 oligomeric species by SANS measurements. Our results demonstrate that the smallest detectable species in solution are penta- to hexamers. No evidences for the presence of dimers, trimers or tetramers were found, although the existence of those Aβ42 oligomers at measurable quantities had been reported frequently.
Collapse
Affiliation(s)
- Martin Wolff
- Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, 40225, Düsseldorf, Germany
- Institute of Complex Systems, Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany
- Physikalische Biochemie, University Potsdam, 14476, Golm, Germany
| | - Bo Zhang-Haagen
- Institute of Complex Systems, Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany
- Jülich Centre for Neutron Science & Institute of Complex Systems, Neutron Scattering (JCNS-1&ICS-1), Forschungszentrum Jülich, 52425, Jülich, Germany
| | - Christina Decker
- Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, 40225, Düsseldorf, Germany
| | - Bogdan Barz
- Institute of Complex Systems, Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany
| | - Mario Schneider
- Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, 40225, Düsseldorf, Germany
| | - Ralf Biehl
- Jülich Centre for Neutron Science & Institute of Complex Systems, Neutron Scattering (JCNS-1&ICS-1), Forschungszentrum Jülich, 52425, Jülich, Germany
- Jülich Centre for Neutron Science, Outstation at MLZ (JCNS-MLZ), Forschungszentrum Jülich, 85747, Garching, Germany
| | - Aurel Radulescu
- Jülich Centre for Neutron Science, Outstation at MLZ (JCNS-MLZ), Forschungszentrum Jülich, 85747, Garching, Germany
| | - Birgit Strodel
- Institute of Complex Systems, Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany
| | - Dieter Willbold
- Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, 40225, Düsseldorf, Germany
- Institute of Complex Systems, Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany
| | - Luitgard Nagel-Steger
- Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, 40225, Düsseldorf, Germany.
- Institute of Complex Systems, Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany.
| |
Collapse
|
|
33
|
Fibril polymorphism affects immobilized non-amyloid flanking domains of huntingtin exon1 rather than its polyglutamine core. Nat Commun 2017; 8:15462. [PMID: 28537272 PMCID: PMC5458082 DOI: 10.1038/ncomms15462] [Citation(s) in RCA: 72] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2017] [Accepted: 03/31/2017] [Indexed: 02/07/2023] Open
Abstract
Polyglutamine expansion in the huntingtin protein is the primary genetic cause of Huntington's disease (HD). Fragments coinciding with mutant huntingtin exon1 aggregate in vivo and induce HD-like pathology in mouse models. The resulting aggregates can have different structures that affect their biochemical behaviour and cytotoxic activity. Here we report our studies of the structure and functional characteristics of multiple mutant htt exon1 fibrils by complementary techniques, including infrared and solid-state NMR spectroscopies. Magic-angle-spinning NMR reveals that fibrillar exon1 has a partly mobile α-helix in its aggregation-accelerating N terminus, and semi-rigid polyproline II helices in the proline-rich flanking domain (PRD). The polyglutamine-proximal portions of these domains are immobilized and clustered, limiting access to aggregation-modulating antibodies. The polymorphic fibrils differ in their flanking domains rather than the polyglutamine amyloid structure. They are effective at seeding polyglutamine aggregation and exhibit cytotoxic effects when applied to neuronal cells. Huntington's disease is caused by a polyglutamine stretch expansion in the first exon of huntingtin. Here, the authors use infrared spectroscopy and solid-state NMR and show that polymorphic huntingtin exon1 fibres differ in their flanking regions but not their core polyglutamine amyloid structures.
Collapse
|
|
34
|
Affiliation(s)
- Markus Zweckstetter
- Deutsches Zentrum für Neurodegenerative Erkrankungen, Göttingen, Germany
- Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
- University Medical Center Göttingen, University of Göttingen, Göttingen, Germany
| | - Jesús R. Requena
- CIMUS Biomedical Research Institute, University of Santiago de Compostela-IDIS, Santiago de Compostela, Spain
| | - Holger Wille
- Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada
- Centre for Prions and Protein Folding Diseases, University of Alberta, Edmonton, Alberta, Canada
- * E-mail:
| |
Collapse
|
|
35
|
Brinet D, Gaie-Levrel F, Delatour V, Kaffy J, Ongeri S, Taverna M. In vitro monitoring of amyloid β-peptide oligomerization by Electrospray differential mobility analysis: An alternative tool to evaluate Alzheimer's disease drug candidates. Talanta 2017; 165:84-91. [DOI: 10.1016/j.talanta.2016.12.011] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2016] [Revised: 12/04/2016] [Accepted: 12/06/2016] [Indexed: 11/27/2022]
|
|
36
|
Melki R. How the shapes of seeds can influence pathology. Neurobiol Dis 2017; 109:201-208. [PMID: 28363800 DOI: 10.1016/j.nbd.2017.03.011] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2016] [Revised: 03/16/2017] [Accepted: 03/26/2017] [Indexed: 10/19/2022] Open
Abstract
It is widely accepted that the loss of function of different cellular proteins following their aggregation into highly stable aggregates or the gain of pathologic function of the resulting macromolecular assemblies or both processes are tightly associated to distinct debilitating neurodegenerative diseases such as Alzheimer's, Parkinson's, Creutzfeldt-Jacob, Amyotrophic Lateral Sclerosis and Huntington's diseases. How the aggregation of one given protein leads to distinct diseases is unclear. Here, a structural-molecular explanation based on the ability of proteins such as α-synuclein or tau to form assemblies that differ by their intrinsic architecture, stability, seeding capacity, and surfaces is proposed to account for distinct synucleinopathies and tauopathies. The shape and surfaces of the seeds is proposed to define at the same time their seeding capacity, interactome and tropism for defined neuronal cells within the central nervous system.
Collapse
Affiliation(s)
- Ronald Melki
- Paris Saclay Institute of Neurosciences, CNRS, Bâtiment 32-33, 1 Avenue de la Terrasse, 91190 Gif-sur-Yvette, France.
| |
Collapse
|
|
37
|
Seuring C, Verasdonck J, Ringler P, Cadalbert R, Stahlberg H, Böckmann A, Meier BH, Riek R. Amyloid Fibril Polymorphism: Almost Identical on the Atomic Level, Mesoscopically Very Different. J Phys Chem B 2017; 121:1783-1792. [PMID: 28075583 DOI: 10.1021/acs.jpcb.6b10624] [Citation(s) in RCA: 36] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Amyloid polymorphism of twisted and straight β-endorphin fibrils was studied by negative-stain transmission electron microscopy, scanning transmission electron microscopy, and solid-state nuclear magnetic resonance spectroscopy. Whereas fibrils assembled in the presence of salt formed flat, striated ribbons, in the absence of salt they formed mainly twisted filaments. To get insights into their structural differences at the atomic level, 3D solid-state NMR spectra of both fibril types were acquired, allowing the detection of the differences in chemical shifts of 13C and 15N atoms in both preparations. The spectral fingerprints and therefore the chemical shifts are very similar for both fibril types. This indicates that the monomer structure and the molecular interfaces are almost the same but that these small differences do propagate to produce flat and twisted morphologies at the mesoscopic scale. This finding is in agreement with both experimental and theoretical considerations on the assembly of polymers (including amyloids) under different salt conditions, which attribute the mesoscopic difference of flat versus twisted fibrils to electrostatic intermolecular repulsions.
Collapse
Affiliation(s)
- Carolin Seuring
- Laboratory of Physical Chemistry, ETH Zürich , Vladimir-Prelog-Weg 2, 8093 Zürich, Switzerland
| | - Joeri Verasdonck
- Laboratory of Physical Chemistry, ETH Zürich , Vladimir-Prelog-Weg 2, 8093 Zürich, Switzerland
| | - Philippe Ringler
- Center for Cellular Imaging and Nano Analytics (C-CINA), Biozentrum University of Basel , 4085 Basel, Switzerland
| | - Riccardo Cadalbert
- Laboratory of Physical Chemistry, ETH Zürich , Vladimir-Prelog-Weg 2, 8093 Zürich, Switzerland
| | - Henning Stahlberg
- Center for Cellular Imaging and Nano Analytics (C-CINA), Biozentrum University of Basel , 4085 Basel, Switzerland
| | - Anja Böckmann
- Molecular Microbiology and Structural Biochemistry, UMR 5086 CNRS, Université de Lyon 1 , 7 passage du Vercors, 69367 Lyon, France
| | - Beat H Meier
- Laboratory of Physical Chemistry, ETH Zürich , Vladimir-Prelog-Weg 2, 8093 Zürich, Switzerland
| | - Roland Riek
- Laboratory of Physical Chemistry, ETH Zürich , Vladimir-Prelog-Weg 2, 8093 Zürich, Switzerland.,Structural Biology Laboratory, The Salk Institute , 10010 N Torrey Pines Road, 92037 La Jolla, California, United States
| |
Collapse
|
|
38
|
Smith AA, Ravotti F, Testori E, Cadalbert R, Ernst M, Böckmann A, Meier BH. Partially-deuterated samples of HET-s(218-289) fibrils: assignment and deuterium isotope effect. JOURNAL OF BIOMOLECULAR NMR 2017; 67:109-119. [PMID: 28074361 DOI: 10.1007/s10858-016-0087-0] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/01/2016] [Accepted: 12/25/2016] [Indexed: 05/26/2023]
Abstract
Fast magic-angle spinning and partial sample deuteration allows direct detection of 1H in solid-state NMR, yielding significant gains in mass sensitivity. In order to further analyze the spectra, 1H detection requires assignment of the 1H resonances. In this work, resonance assignments of backbone HN and Hα are presented for HET-s(218-289) fibrils, based on the existing assignment of Cα, Cβ, C', and N resonances. The samples used are partially deuterated for higher spectral resolution, and the shifts in resonance frequencies of Cα and Cβ due to the deuterium isotope effect are investigated. It is shown that the deuterium isotope effect can be estimated and used for assigning resonances of deuterated samples in solid-state NMR, based on known resonances of the protonated protein.
Collapse
Affiliation(s)
- Albert A Smith
- ETH Zürich, Physical Chemistry, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland
| | - Francesco Ravotti
- ETH Zürich, Physical Chemistry, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland
| | - Emilie Testori
- ETH Zürich, Physical Chemistry, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland
| | - Riccardo Cadalbert
- ETH Zürich, Physical Chemistry, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland
| | - Matthias Ernst
- ETH Zürich, Physical Chemistry, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland.
| | - Anja Böckmann
- Institut de Biologie et Chimie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France.
| | - Beat H Meier
- ETH Zürich, Physical Chemistry, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland.
| |
Collapse
|
|
39
|
Lendel C, Sparrman T, Mayzel M, Andersson CE, Karlsson G, Härd T. Combined Solution- and Magic Angle Spinning NMR Reveals Regions of Distinct Dynamics in Amyloid β Protofibrils. ChemistrySelect 2016. [DOI: 10.1002/slct.201601468] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022]
Affiliation(s)
- Christofer Lendel
- Dept. of Chemistry and Biotechnology; Swedish University of Agricultural Sciences (SLU); Uppsala Sweden
- Dept. of Chemistry - present address; KTH Royal Institute of Technology; Stockholm Sweden
| | | | - Maxim Mayzel
- Swedish NMR Centre; University of Gothenburg; Gothenburg Sweden
| | - C. Evalena Andersson
- Dept. of Chemistry and Biotechnology; Swedish University of Agricultural Sciences (SLU); Uppsala Sweden
- Dept. of Cell and Molecular Biology - present address; Uppsala University; Uppsala Sweden
| | - Göran Karlsson
- Swedish NMR Centre; University of Gothenburg; Gothenburg Sweden
| | - Torleif Härd
- Dept. of Chemistry and Biotechnology; Swedish University of Agricultural Sciences (SLU); Uppsala Sweden
| |
Collapse
|
|
40
|
Ravotti F, Wälti MA, Güntert P, Riek R, Böckmann A, Meier BH. Solid-state NMR sequential assignment of an Amyloid-β(1-42) fibril polymorph. BIOMOLECULAR NMR ASSIGNMENTS 2016; 10:269-76. [PMID: 27165577 DOI: 10.1007/s12104-016-9682-y] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/25/2016] [Accepted: 04/30/2016] [Indexed: 05/10/2023]
Abstract
The formation of fibrils of the amyloid-β (Aβ) peptide is considered to be a key event in the pathology of Alzheimer's disease (AD). The determination of a high-resolution structure of these fibrils is relevant for the understanding of the molecular basis of AD. In this work, we present the sequential resonance assignment of one of the polymorphs of Aβ(1-42) fibrils. We show that most of the protein is rigid, while a stretch of 4 residues (11-14) is not visible by solid-state NMR spectroscopy due to dynamics.
Collapse
Affiliation(s)
- Francesco Ravotti
- Physical Chemistry, ETH Zürich, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland
| | | | - Peter Güntert
- Institute of Biophysical Chemistry, Center for Biomolecular Magnetic Resonance, Goethe University Frankfurt am Main, Max-von-Laue-Str. 9, 60438, Frankfurt am Main, Germany.
- Department of Chemistry, Graduate School of Science and Engineering, Tokyo Metropolitan University, 1-1 Minami-Ohsawa, Hachioji, Tokyo, 192-0397, Japan.
| | - Roland Riek
- Physical Chemistry, ETH Zürich, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland.
| | - Anja Böckmann
- Institut de Biologie et Chemie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69007, Lyon, France.
| | - Beat H Meier
- Physical Chemistry, ETH Zürich, Vladimir-Prelog-Weg 2, 8093, Zürich, Switzerland.
| |
Collapse
|
|
41
|
Tyson T, Steiner JA, Brundin P. Sorting out release, uptake and processing of alpha-synuclein during prion-like spread of pathology. J Neurochem 2016; 139 Suppl 1:275-289. [PMID: 26617280 PMCID: PMC4958606 DOI: 10.1111/jnc.13449] [Citation(s) in RCA: 72] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2015] [Revised: 11/23/2015] [Accepted: 11/25/2015] [Indexed: 12/17/2022]
Abstract
Parkinson's disease is a progressive neurological disorder that is characterized by the formation of intracellular protein inclusion bodies composed primarily of a misfolded and aggregated form of the protein α-synuclein. There is growing evidence that supports the prion-like hypothesis of α-synuclein progression. This hypothesis postulates that α-synuclein is a prion-like pathological agent and is responsible for the progression of Parkinson pathology in the brain. Potential misfolding or aggregation of α-synuclein that might occur in the peripheral nervous system as a result of some insult, environmental or genetic (or more likely a combination of both) that might spread into the midbrain, eventually causing degeneration of the neurons in the substantia nigra. As the disease progresses further, it is likely that α-synuclein pathology continues to spread throughout the brain, including the cortex, leading to deterioration of cognition and higher brain functions. While it is unknown why α-synuclein initially misfolds and aggregates, a great deal has been learned about how the cell handles aberrant α-synuclein assemblies. In this review, we focus on these mechanisms and discuss them in an attempt to define the role that they might play in the propagation of misfolded α-synuclein from cell-to-cell. The prion-like hypothesis of α-synuclein pathology suggests a method for the transmission of misfolded α-synuclein from one neuron to another. This hypothesis postulates that misfolded α-synuclein becomes aggregation prone and when released and taken up by neighboring cells, seeds further misfolding and aggregation. In this review we examine the cellular mechanisms that are involved in the processing of α-synuclein and how these may contribute to the prion-like propagation of α-synuclein pathology. This article is part of a special issue on Parkinson disease.
Collapse
Affiliation(s)
- Trevor Tyson
- Center for Neurodegenerative Science, Van Andel Research Institute, Grand Rapids, Michigan, USA
| | - Jennifer A Steiner
- Center for Neurodegenerative Science, Van Andel Research Institute, Grand Rapids, Michigan, USA
| | - Patrik Brundin
- Center for Neurodegenerative Science, Van Andel Research Institute, Grand Rapids, Michigan, USA.
| |
Collapse
|
|
42
|
Elkins MR, Wang T, Nick M, Jo H, Lemmin T, Prusiner SB, DeGrado WF, Stöhr J, Hong M. Structural Polymorphism of Alzheimer's β-Amyloid Fibrils as Controlled by an E22 Switch: A Solid-State NMR Study. J Am Chem Soc 2016; 138:9840-52. [PMID: 27414264 PMCID: PMC5149419 DOI: 10.1021/jacs.6b03715] [Citation(s) in RCA: 72] [Impact Index Per Article: 8.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
The amyloid-β (Aβ) peptide of Alzheimer's disease (AD) forms polymorphic fibrils on the micrometer and molecular scales. Various fibril growth conditions have been identified to cause polymorphism, but the intrinsic amino acid sequence basis for this polymorphism has been unclear. Several single-site mutations in the center of the Aβ sequence cause different disease phenotypes and fibrillization properties. The E22G (Arctic) mutant is found in familial AD and forms protofibrils more rapidly than wild-type Aβ. Here, we use solid-state NMR spectroscopy to investigate the structure, dynamics, hydration and morphology of Arctic E22G Aβ40 fibrils. (13)C, (15)N-labeled synthetic E22G Aβ40 peptides are studied and compared with wild-type and Osaka E22Δ Aβ40 fibrils. Under the same fibrillization conditions, Arctic Aβ40 exhibits a high degree of polymorphism, showing at least four sets of NMR chemical shifts for various residues, while the Osaka and wild-type Aβ40 fibrils show a single or a predominant set of chemical shifts. Thus, structural polymorphism is intrinsic to the Arctic E22G Aβ40 sequence. Chemical shifts and inter-residue contacts obtained from 2D correlation spectra indicate that one of the major Arctic conformers has surprisingly high structural similarity with wild-type Aβ42. (13)C-(1)H dipolar order parameters, (1)H rotating-frame spin-lattice relaxation times and water-to-protein spin diffusion experiments reveal substantial differences in the dynamics and hydration of Arctic, Osaka and wild-type Aβ40 fibrils. Together, these results strongly suggest that electrostatic interactions in the center of the Aβ peptide sequence play a crucial role in the three-dimensional fold of the fibrils, and by inference, fibril-induced neuronal toxicity and AD pathogenesis.
Collapse
Affiliation(s)
- Matthew R. Elkins
- Department of Chemistry, Massachusetts Institute of Technology, Cambridge MA 02139
| | - Tuo Wang
- Department of Chemistry, Massachusetts Institute of Technology, Cambridge MA 02139
| | - Mimi Nick
- Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158
| | - Hyunil Jo
- Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158
| | - Thomas Lemmin
- Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158
| | - Stanley B. Prusiner
- Institute for Neurodegenerative Diseases, Departments of Neurology, University of California, San Francisco, San Francisco, CA 94143
| | - William F. DeGrado
- Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, CA 94158
| | - Jan Stöhr
- Institute for Neurodegenerative Diseases, Departments of Neurology, University of California, San Francisco, San Francisco, CA 94143
| | - Mei Hong
- Department of Chemistry, Massachusetts Institute of Technology, Cambridge MA 02139
| |
Collapse
|
|
43
|
Atomic-resolution structure of a disease-relevant Aβ(1-42) amyloid fibril. Proc Natl Acad Sci U S A 2016; 113:E4976-84. [PMID: 27469165 DOI: 10.1073/pnas.1600749113] [Citation(s) in RCA: 668] [Impact Index Per Article: 74.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/17/2023] Open
Abstract
Amyloid-β (Aβ) is present in humans as a 39- to 42-amino acid residue metabolic product of the amyloid precursor protein. Although the two predominant forms, Aβ(1-40) and Aβ(1-42), differ in only two residues, they display different biophysical, biological, and clinical behavior. Aβ(1-42) is the more neurotoxic species, aggregates much faster, and dominates in senile plaque of Alzheimer's disease (AD) patients. Although small Aβ oligomers are believed to be the neurotoxic species, Aβ amyloid fibrils are, because of their presence in plaques, a pathological hallmark of AD and appear to play an important role in disease progression through cell-to-cell transmissibility. Here, we solved the 3D structure of a disease-relevant Aβ(1-42) fibril polymorph, combining data from solid-state NMR spectroscopy and mass-per-length measurements from EM. The 3D structure is composed of two molecules per fibril layer, with residues 15-42 forming a double-horseshoe-like cross-β-sheet entity with maximally buried hydrophobic side chains. Residues 1-14 are partially ordered and in a β-strand conformation, but do not display unambiguous distance restraints to the remainder of the core structure.
Collapse
|
|
44
|
NMR Meets Tau: Insights into Its Function and Pathology. Biomolecules 2016; 6:biom6020028. [PMID: 27338491 PMCID: PMC4919923 DOI: 10.3390/biom6020028] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2016] [Revised: 05/02/2016] [Accepted: 05/26/2016] [Indexed: 12/21/2022] Open
Abstract
In this review, we focus on what we have learned from Nuclear Magnetic Resonance (NMR) studies on the neuronal microtubule-associated protein Tau. We consider both the mechanistic details of Tau: the tubulin relationship and its aggregation process. Phosphorylation of Tau is intimately linked to both aspects. NMR spectroscopy has depicted accurate phosphorylation patterns by different kinases, and its non-destructive character has allowed functional assays with the same samples. Finally, we will discuss other post-translational modifications of Tau and its interaction with other cellular factors in relationship to its (dys)function.
Collapse
|
|
45
|
Wiegand T, Gardiennet C, Cadalbert R, Lacabanne D, Kunert B, Terradot L, Böckmann A, Meier BH. Variability and conservation of structural domains in divide-and-conquer approaches. JOURNAL OF BIOMOLECULAR NMR 2016; 65:79-86. [PMID: 27240588 DOI: 10.1007/s10858-016-0039-8] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/01/2016] [Accepted: 05/19/2016] [Indexed: 05/26/2023]
Abstract
The use of protein building blocks for the structure determination of multidomain proteins and protein-protein complexes, also known as the "divide and conquer" approach, is an important strategy for obtaining protein structures. Atomic-resolution X-ray or NMR data of the individual domains are combined with lower-resolution electron microscopy maps or X-ray data of the full-length protein or the protein complex. Doing so, it is often assumed that the individual domain structures remain invariant in the context of the superstructure. In this work, we show the potentials and limitations of NMR to validate this approach at the example of the dodecameric DnaB helicase from Helicobacter pylori. We investigate how sequentially assigned spectra, as well as unassigned spectral fingerprints can be used to indicate the conservation of individual domains, and also to highlight conformational differences.
Collapse
Affiliation(s)
- Thomas Wiegand
- Physical Chemistry, ETH Zurich, 8093, Zurich, Switzerland
| | - Carole Gardiennet
- Institut de Biologie et Chimie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France
- CRM2, UMR 7036, CNRS, Université de Lorraine, 54506, Vandoeuvre-lès-Nancy, France
| | | | - Denis Lacabanne
- Institut de Biologie et Chimie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France
| | - Britta Kunert
- Institut de Biologie et Chimie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France
| | - Laurent Terradot
- Institut de Biologie et Chimie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France.
| | - Anja Böckmann
- Institut de Biologie et Chimie des Protéines, Bases Moléculaires et Structurales des Systèmes Infectieux, Labex Ecofect, UMR 5086 CNRS, Université de Lyon, 7 passage du Vercors, 69367, Lyon, France.
| | - Beat H Meier
- Physical Chemistry, ETH Zurich, 8093, Zurich, Switzerland.
| |
Collapse
|
|
46
|
Brunori M, Gianni S. Molecular medicine - To be or not to be. Biophys Chem 2016; 214-215:33-46. [PMID: 27214761 DOI: 10.1016/j.bpc.2016.05.004] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2016] [Revised: 05/11/2016] [Accepted: 05/11/2016] [Indexed: 12/17/2022]
Abstract
Molecular medicine is founded on the synergy between Chemistry, Physics, Biology and Medicine, with the ambitious goal of tackling diseases from a molecular perspective. This Review aims at retracing a personal outlook of the birth and development of molecular medicine, as well as at highlighting some of the most urgent challenges linked to aging and represented by incurable neurodegenerative diseases caused by protein misfolding. Furthermore, we emphasize the emerging role of the retromer dysfunctions and improper protein sorting in Alzheimer's disease and other important neurological disordered.
Collapse
Affiliation(s)
- Maurizio Brunori
- Istituto Pasteur - Fondazione Cenci Bolognetti and Istituto di Biologia e Patologia Molecolari del CNR, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli", Sapienza Università di Roma, 00185 Rome, Italy.
| | - Stefano Gianni
- Istituto Pasteur - Fondazione Cenci Bolognetti and Istituto di Biologia e Patologia Molecolari del CNR, Dipartimento di Scienze Biochimiche "A. Rossi Fanelli", Sapienza Università di Roma, 00185 Rome, Italy.
| |
Collapse
|
|
47
|
Nagel-Steger L, Owen MC, Strodel B. An Account of Amyloid Oligomers: Facts and Figures Obtained from Experiments and Simulations. Chembiochem 2016; 17:657-76. [PMID: 26910367 DOI: 10.1002/cbic.201500623] [Citation(s) in RCA: 85] [Impact Index Per Article: 9.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2015] [Indexed: 12/27/2022]
Abstract
The deposition of amyloid in brain tissue in the context of neurodegenerative diseases involves the formation of intermediate species-termed oligomers-of lower molecular mass and with structures that deviate from those of mature amyloid fibrils. Because these oligomers are thought to be primarily responsible for the subsequent disease pathogenesis, the elucidation of their structure is of enormous interest. Nevertheless, because of the high aggregation propensity and the polydispersity of oligomeric species formed by the proteins or peptides in question, the preparation of appropriate samples for high-resolution structural methods has proven to be rather difficult. This is why theoretical approaches have been of particular importance in gaining insights into possible oligomeric structures for some time. Only recently has it been possible to achieve some progress with regard to the experimentally based structural characterization of defined oligomeric species. Here we discuss how theory and experiment are used to determine oligomer structures and what can be done to improve the integration of the two disciplines.
Collapse
Affiliation(s)
- Luitgard Nagel-Steger
- Institute of Complex Systems: Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany.,Institut für Physikalische Biologie, Heinrich-Heine-Universität Düsseldorf, Universitätstrasse 1, 40225, Düsseldorf, Germany
| | - Michael C Owen
- Institute of Complex Systems: Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany
| | - Birgit Strodel
- Institute of Complex Systems: Structural Biochemistry (ICS-6), Forschungszentrum Jülich, 52425, Jülich, Germany. .,Institute of Theoretical and Computational Chemistry, Heinrich Heine University Düsseldorf, Universitätstrasse 1, 40225, Düsseldorf, Germany.
| |
Collapse
|
|
48
|
|
|
49
|
Schubeis T, Yuan P, Ahmed M, Nagaraj M, van Rossum B, Ritter C. Untangling a Repetitive Amyloid Sequence: Correlating Biofilm‐Derived and Segmentally Labeled Curli Fimbriae by Solid‐State NMR Spectroscopy. Angew Chem Int Ed Engl 2015; 54:14669-72. [DOI: 10.1002/anie.201506772] [Citation(s) in RCA: 28] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2015] [Indexed: 11/09/2022]
Affiliation(s)
- Tobias Schubeis
- Laboratory of Macromolecular Interactions, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstrasse 7, 38124 Braunschweig (Germany)
| | - Puwei Yuan
- Laboratory of Macromolecular Interactions, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstrasse 7, 38124 Braunschweig (Germany)
| | - Mumdooh Ahmed
- Laboratory of Macromolecular Interactions, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstrasse 7, 38124 Braunschweig (Germany)
- Department of Physics, Faculty of Science, Suez University, Suez, 43533 (Egypt)
| | - Madhu Nagaraj
- Laboratory of Macromolecular Interactions, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstrasse 7, 38124 Braunschweig (Germany)
- NMR‐Supported Structural Biology, Leibniz‐Institut für Molekulare Pharmakologie (FMP), Robert‐Rössle‐Strasse 10, 13125 Berlin (Germany)
| | - Barth‐Jan van Rossum
- NMR‐Supported Structural Biology, Leibniz‐Institut für Molekulare Pharmakologie (FMP), Robert‐Rössle‐Strasse 10, 13125 Berlin (Germany)
| | - Christiane Ritter
- Laboratory of Macromolecular Interactions, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstrasse 7, 38124 Braunschweig (Germany)
| |
Collapse
|
|
50
|
Schubeis T, Yuan P, Ahmed M, Nagaraj M, van Rossum B, Ritter C. Entschlüsselung einer repetitiven Amyloidsequenz: Korrelation von Biofilm‐extrahierten und segmental markierten Curli‐Fimbrien mittels Festkörper‐NMR‐Spektroskopie. Angew Chem Int Ed Engl 2015. [DOI: 10.1002/ange.201506772] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Affiliation(s)
- Tobias Schubeis
- Laboratorium für makromolekulare Interaktionen, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstraße 7, 38124 Braunschweig (Deutschland)
| | - Puwei Yuan
- Laboratorium für makromolekulare Interaktionen, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstraße 7, 38124 Braunschweig (Deutschland)
| | - Mumdooh Ahmed
- Laboratorium für makromolekulare Interaktionen, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstraße 7, 38124 Braunschweig (Deutschland)
- Department of Physics, Faculty of Science, Suez University, Suez, 43533 (Ägypten)
| | - Madhu Nagaraj
- Laboratorium für makromolekulare Interaktionen, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstraße 7, 38124 Braunschweig (Deutschland)
- NMR‐Supported Structural Biology, Leibniz‐Institut für Molekulare Pharmakologie (FMP), Robert‐Rössle‐Straße 10, 13125 Berlin (Deutschland)
| | - Barth‐Jan van Rossum
- NMR‐Supported Structural Biology, Leibniz‐Institut für Molekulare Pharmakologie (FMP), Robert‐Rössle‐Straße 10, 13125 Berlin (Deutschland)
| | - Christiane Ritter
- Laboratorium für makromolekulare Interaktionen, Helmholtz‐Zentrum für Infektionsforschung, Inhoffenstraße 7, 38124 Braunschweig (Deutschland)
| |
Collapse
|