Decision letter: Two new polymorphic structures of human full-length alpha-synuclein fibrils solved by cryo-electron microscopy

Parkinson’s disease is a progressive neuropathological disorder that belongs to the class of synucleinopathies, in which the protein alpha-synuclein is found at abnormally high concentrations in affected neurons. Its hallmark are intracellular inclusions called Lewy bodies and Lewy neurites. We here report the structure of cytotoxic alpha-synuclein fibrils (residues 1–121), determined by cryo-electron microscopy at a resolution of 3.4 Å. Two protofilaments form a polar fibril composed of staggered β-strands. The backbone of residues 38 to 95, including the fibril core and the non-amyloid component region, are well resolved in the EM map. Residues 50–57, containing three of the mutation sites associated with familial synucleinopathies, form the interface between the two protofilaments and contribute to fibril stability. A hydrophobic cleft at one end of the fibril may have implications for fibril elongation, and invites for the design of molecules for diagnosis and treatment of synucleinopathies.

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Structural insight into the function of full-length human Fanconi Anemia FANCD2/FANCI complex by cryo-electron microscopy

European Microscopy Congress 2016: Proceedings ◽

10.1002/9783527808465.emc2016.6465 ◽

2016 ◽

pp. 47-48

Author(s):

Zhuolun Li ◽

Chih-Chao Liang ◽

William Nicholson ◽

Martin Cohn ◽

Catherine Venien-Bryan

Keyword(s):

Electron Microscopy ◽

Fanconi Anemia ◽

Full Length ◽

Cryo Electron Microscopy ◽

Structural Insight ◽

Insight Into

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Domain topology of human Rasal

Biological Chemistry ◽

10.1515/hsz-2017-0159 ◽

2017 ◽

Vol 399 (1) ◽

pp. 63-72 ◽

Cited By ~ 2

Author(s):

Jorge Cuellar ◽

José María Valpuesta ◽

Alfred Wittinghofer ◽

Begoña Sot

Keyword(s):

Electron Microscopy ◽

Small Gtpase ◽

Full Length ◽

Gtpase Activating Protein ◽

Ras Gtpase ◽

Cryo Electron Microscopy ◽

Domain Protein ◽

Negative Staining Electron Microscopy ◽

Resolution Structure

AbstractRasal is a modular multi-domain protein of the GTPase-activating protein 1 (GAP1) family; its four known members, GAP1m, Rasal, GAP1IP4BPand Capri, have a Ras GTPase-activating domain (RasGAP). This domain supports the intrinsically slow GTPase activity of Ras by actively participating in the catalytic reaction. In the case of Rasal, GAP1IP4BPand Capri, their remaining domains are responsible for converting the RasGAP domains into dual Ras- and Rap-GAPs, via an incompletely understood mechanism. Although Rap proteins are small GTPase homologues of Ras, their catalytic residues are distinct, which reinforces the importance of determining the structure of full-length GAP1 family proteins. To date, these proteins have not been crystallized, and their size is not adequate for nuclear magnetic resonance (NMR) or for high-resolution cryo-electron microscopy (cryoEM). Here we present the low resolution structure of full-length Rasal, obtained by negative staining electron microscopy, which allows us to propose a model of its domain topology. These results help to understand the role of the different domains in controlling the dual GAP activity of GAP1 family proteins.

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Cryo-Electron Microscopy of Arabidopsis thaliana Phytochrome A in Its Pr State Reveals Head-to-Head Homodimeric Architecture

Frontiers in Plant Science ◽

10.3389/fpls.2021.663751 ◽

2021 ◽

Vol 12 ◽

Author(s):

Weixiao Yuan Wahlgren ◽

David Golonka ◽

Sebastian Westenhoff ◽

Andreas Möglich

Keyword(s):

Electron Microscopy ◽

Arabidopsis Thaliana ◽

Electron Density ◽

Molecular Mechanisms ◽

Signal Propagation ◽

Full Length ◽

Particle Analysis ◽

Phytochrome A ◽

Cryo Electron Microscopy ◽

Bacterial Phytochromes

Phytochrome photoreceptors regulate vital adaptations of plant development, growth, and physiology depending on the ratio of red and far-red light. The light-triggered Z/E isomerization of a covalently bound bilin chromophore underlies phytochrome photoconversion between the red-absorbing Pr and far-red-absorbing Pfr states. Compared to bacterial phytochromes, the molecular mechanisms of signal propagation to the C-terminal module and its regulation are little understood in plant phytochromes, not least owing to a dearth of structural information. To address this deficit, we studied the Arabidopsis thaliana phytochrome A (AtphyA) at full length by cryo-electron microscopy (cryo-EM). Following heterologous expression in Escherichia coli, we optimized the solvent conditions to overcome protein aggregation and thus obtained photochemically active, near-homogenous AtphyA. We prepared grids for cryo-EM analysis of AtphyA in its Pr state and conducted single-particle analysis. The resulting two-dimensional class averages and the three-dimensional electron density map at 17 Å showed a homodimeric head-to-head assembly of AtphyA. Docking of domain structures into the electron density revealed a separation of the AtphyA homodimer at the junction of its photosensor and effector modules, as reflected in a large void in the middle of map. The overall architecture of AtphyA resembled that of bacterial phytochromes, thus hinting at commonalities in signal transduction and mechanism between these receptors. Our work paves the way toward future studies of the structure, light response, and interactions of full-length phytochromes by cryo-EM.

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Cryo-EM structure of alpha-synuclein fibrils

10.1101/276436 ◽

2018 ◽

Cited By ~ 1

Author(s):

Ricardo Guerrero-Ferreira ◽

Nicholas M. I. Taylor ◽

Daniel Mona ◽

Philippe Ringler ◽

Matthias E. Lauer ◽

...

Keyword(s):

Electron Microscopy ◽

Rational Design ◽

Alpha Synuclein ◽

Diagnosis And Treatment ◽

Cryo Electron Microscopy ◽

Cell Propagation ◽

Hydrophobic Cleft

AbstractIntracellular inclusions of alpha-synuclein are the neuropathological hallmark of progressive disorders called synucleinopathies. Alpha-synuclein fibrils are associated with transmissive cell-to-cell propagation of pathology. We report the structure of an alpha-synuclein fibril (residues 1-121) determined by cryo-electron microscopy at 3.4Å resolution. Two protofilaments form a polar fibril composed of staggered β-strands. The backbone of residues 38 to 95, including the fibril core and the non-amyloid component region, are well resolved in the EM map. Residues 50-57, containing three mutation sites associated with familial synucleinopathies, form the interface between the two protofilaments and contribute to fibril stability. A hydrophobic cleft may have implications for fibril elongation, and inform the rational design of molecules for diagnosis and treatment of synucleinopathies.

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Cryo-Electron Microscopy Uncovers Key Residues within the Core of Alpha-Synuclein Fibrils

ACS Chemical Neuroscience ◽

10.1021/acschemneuro.9b00090 ◽

2019 ◽

Vol 10 (3) ◽

pp. 1135-1136 ◽

Cited By ~ 3

Author(s):

Ritobrita Chakraborty ◽

Krishnananda Chattopadhyay

Keyword(s):

Electron Microscopy ◽

Alpha Synuclein ◽

The Core ◽

Cryo Electron Microscopy ◽

Key Residues

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New insights on the structure of alpha-synuclein fibrils using cryo-electron microscopy

Current Opinion in Neurobiology ◽

10.1016/j.conb.2020.01.014 ◽

2020 ◽

Vol 61 ◽

pp. 89-95 ◽

Cited By ~ 7

Author(s):

Ricardo Guerrero-Ferreira ◽

Lubomir Kovacik ◽

Dongchun Ni ◽

Henning Stahlberg

Keyword(s):

Electron Microscopy ◽

Alpha Synuclein ◽

Cryo Electron Microscopy

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Cryo-EM structures of the Caspase activated protein XKR9 involved in apoptotic lipid scrambling

10.1101/2021.05.03.442400 ◽

2021 ◽

Author(s):

Raimund Dutzler ◽

Monique S. Straub ◽

Carolina Alvadia ◽

Marta Sawicka

Keyword(s):

Electron Microscopy ◽

Cell Surface ◽

Protein Function ◽

Caspase 3 ◽

Full Length ◽

Cryo Electron Microscopy ◽

C Terminus ◽

Hydrophobic Site ◽

Membrane Spanning ◽

Insight Into

The exposure of the negatively charged lipid phosphatidylserine on the cell-surface, catalyzed by lipid scramblases, is an important signal for the clearance of apoptotic cells by macrophages. The protein XKR9 is a member of a conserved family that has been associated with apoptotic lipid scrambling. Here, we describe structures of full-length and caspase-treated XKR9 in complex with a synthetic nanobody determined by cryo-electron microscopy. The 43 kDa monomeric membrane protein contains eight membrane-spanning helices, two segments that are partly inserted into the lipid bilayer and is organized as two structurally related repeats. In the full-length protein, the C-terminus interacts with a hydrophobic site located at the intracellular side acting as an inhibitor of protein function. Cleavage by caspase-3 at a specific site releases 16 residues of the C-terminus thus making the binding site accessible to the cytoplasm. Collectively, the work has revealed the unknown architecture of the XKR family and has provided initial insight into its activation by caspases.

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Two new polymorphic structures of human full-length alpha-synuclein fibrils solved by cryo-electron microscopy

eLife ◽

10.7554/elife.48907 ◽

2019 ◽

Vol 8 ◽

Cited By ~ 43

Author(s):

Ricardo Guerrero-Ferreira ◽

Nicholas MI Taylor ◽

Ana-Andreea Arteni ◽

Pratibha Kumari ◽

Daniel Mona ◽

...

Keyword(s):

Electron Microscopy ◽

Amino Acids ◽

Alpha Synuclein ◽

Fibril Formation ◽

Salt Bridges ◽

Atomic Structures ◽

Cryo Electron Microscopy ◽

N Terminus ◽

Hydrophobic Cleft ◽

Fibril Diameter

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.

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