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Cryo electron microscopy

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https://www.readbyqxmd.com/read/28925789/motile-cilia-defects-in-diseases-other-than-primary-ciliary-dyskinesia-the-contemporary-diagnostic-and-research-role-for-transmission-electron-microscopy
#1
Hannah M Mitchison, Amelia Shoemark
Ultrastructural studies have underpinned the cell biological and clinical investigations of the varied roles of motile cilia in health and disease, with a long history since the 1950s. Recent developments from transmission electron microscopy (TEM; cryo-electron microscopy, electron tomography) have yielded higher resolution and fresh insights into the structure and function of these complex organelles. Microscopy in ciliated organisms, disease models, and in patients with ciliopathy diseases has dramatically expanded our understanding of the ubiquity, multisystem involvement, and importance of cilia in normal human development...
September 19, 2017: Ultrastructural Pathology
https://www.readbyqxmd.com/read/28925040/thermal-melt-circular-dichroism-spectroscopic-studies-for-identifying-stabilising-amphipathic-molecules-for-the-voltage-gated-sodium-channel-navms
#2
Sam M Ireland, Altin Sula, B A Wallace
Purified integral membrane proteins require amphipathic molecules to maintain their solubility in aqueous solutions. These complexes, in turn, are used in studies to characterise the protein structures by a variety of biophysical and structural techniques, including spectroscopy, crystallography, and cryo-electron microscopy. Typically the amphilphiles used have been detergent molecules, but more recently they have included amphipols, which are polymers of different sizes and compositions designed to create smaller, more well-defined solubilised forms of the membrane proteins...
September 19, 2017: Biopolymers
https://www.readbyqxmd.com/read/28922082/combinatorial-peptide-based-epitope-mapping-from-ebola-virus-dna-vaccines-and-infections-reveals-residue-level-determinants-of-antibody-binding
#3
Daniel R Ripoll, Daniel A J Mitchell, Lesley C Dupuy, Anders Wallqvist, Connie Schmaljohn, Sidhartha Chaudhury
Ebola virus (EBOV) infection is highly lethal and results in severe febrile bleeding disorders that affect humans and non-human primates. One of the therapeutic approaches for treating EBOV infection focus largely on cocktails of monoclonal antibodies (mAbs) that bind to specific regions of the EBOV glycoprotein (GP) and neutralize the virus. Recent structural studies using cryo-electron microscopy have identified key epitopes for several EBOV mAbs. While such information has yielded deep insights into antibody binding, limitations on resolution of these structures often preclude a residue-level analysis of EBOV epitopes...
September 18, 2017: Human Vaccines & Immunotherapeutics
https://www.readbyqxmd.com/read/28916391/zika-virus-structural-biology-and-progress-in-vaccine-development
#4
REVIEW
Hsiao-Han Lin, Bak-Sau Yip, Li-Min Huang, Suh-Chin Wu
The growing number of zika virus (ZIKV) infections plus a 20-fold increase in neonatal microcephaly in newborns in Brazil have raised alarms in many countries regarding the threat to pregnant women. Instances of microcephaly and central nervous system malformations continue to increase in ZIKV outbreak regions. ZIKV is a small enveloped positive-strand RNA virus belonging to the Flavivirus genus of the Flaviviridae family. High-resolution ZIKV structures recently identified by cryo-electron microscopy indicate that the overall ZIKV structure is similar to those of other flaviviruses...
September 12, 2017: Biotechnology Advances
https://www.readbyqxmd.com/read/28915442/electron-cryo-tomography-captures-macromolecular-complexes-in-native-environments
#5
REVIEW
Lindsay A Baker, Michael Grange, Kay Grünewald
Transmission electron microscopy has a long history in cellular biology. Fixed and stained samples have been used for cellular imaging for over 50 years, but suffer from sample preparation induced artifacts. Electron cryo-tomography (cryoET) instead uses frozen-hydrated samples, without chemical modification, to determine the structure of macromolecular complexes in their native environment. Recent developments in electron microscopes and associated technologies have greatly expanded our ability to visualize cellular features and determine the structures of macromolecular complexes in situ...
September 12, 2017: Current Opinion in Structural Biology
https://www.readbyqxmd.com/read/28905417/investigating-the-structural-dynamics-of-the-piezo1-channel-activation-and-inactivation-by-coarse-grained-modeling
#6
Wenjun Zheng, Frederick Sachs
The PIEZO channels, a family of mechanosensitive channels in vertebrates, feature a fast activation by mechanical stimuli (e.g., membrane tension) followed by a slower inactivation. Although a medium-resolution structure of the trimeric form of PIEZO1 was solved by cryo-electron microscopy (cryo-EM), key structural changes responsible for the channel activation and inactivation are still unknown. Toward decrypting the structural mechanism of the PIEZO1 activation and inactivation, we performed systematic coarse-grained modeling using an elastic network model and related modeling/analysis tools (i...
September 14, 2017: Proteins
https://www.readbyqxmd.com/read/28902838/the-cryo-electron-microscopy-structure-of-human-transcription-factor-iih
#7
Basil J Greber, Thi Hoang Duong Nguyen, Jie Fang, Pavel V Afonine, Paul D Adams, Eva Nogales
Human transcription factor IIH (TFIIH) is part of the general transcriptional machinery required by RNA polymerase II for the initiation of eukaryotic gene transcription. Composed of ten subunits that add up to a molecular mass of about 500 kDa, TFIIH is also essential for nucleotide excision repair. The seven-subunit TFIIH core complex formed by XPB, XPD, p62, p52, p44, p34, and p8 is competent for DNA repair, while the CDK-activating kinase subcomplex, which includes the kinase activity of CDK7 as well as the cyclin H and MAT1 subunits, is additionally required for transcription initiation...
September 13, 2017: Nature
https://www.readbyqxmd.com/read/28901669/automating-tasks-in-protein-structure-determination-with-the-clipper-python-module
#8
Stuart McNicholas, Tristan Croll, Tom Burnley, Colin M Palmer, Soon Wen Hoh, Huw T Jenkins, Eleanor Dodson, Kevin Cowtan, Jon Agirre
Scripting programming languages provide the fastest means of prototyping complex functionality. Those with a syntax and grammar resembling human language also greatly enhance the maintainability of the produced source code. Furthermore, the combination of a powerful, machine-independent scripting language with binary libraries tailored for each computer architecture allows programs to break free from the tight boundaries of efficiency traditionally associated with scripts. In the present work, we describe how an efficient C++ crystallographic library such as Clipper can be wrapped, adapted and generalised for use in both crystallographic and electron cryo-microscopy applications, scripted with the Python language...
September 13, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28898733/manipulation-of-pickering-emulsion-rheology-using-hydrophilically-modified-silica-nanoparticles-in-brine
#9
Christopher Griffith, Hugh Daigle
HYPOTHESIS: Previous work on Pickering emulsions has shown that bromohexadecane-in-water emulsions (50% oil) stabilized with fumed and spherical particles modified with hexadecyl groups develop a noticeable zero shear elastic storage modulus (G'0) of 200Pa and 9Pa, respectively, while in just 50mM NaCl. This high G'0 can be problematic for subsurface applications where brine salinities are higher and on the order of 600mM NaCl. High reservoir salinity coupled with low formation pressure drops could prevent an emulsion with a high G'0 from propagating deep into formation...
September 1, 2017: Journal of Colloid and Interface Science
https://www.readbyqxmd.com/read/28891250/guidelines-for-using-bsoft-for-high-resolution-reconstruction-and-validation-of-biomolecular-structures-from-electron-micrographs
#10
J Bernard Heymann
Cryo-electron microscopy (cryoEM) is becoming popular as a tool to solve biomolecular structures with the recent availability of direct electron detectors allowing automated acquisition of high resolution data. The Bsoft software package, developed over 20 years for analyzing electron micrographs, offers a full workflow for validated single particle analysis with extensive functionality, enabling customization for specific cases. With the increasing use of cryoEM and its automation, proper validation of the results is a bigger concern...
September 10, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28888105/cryoem-structures-of-spliceosomal-complexes-reveal-the-molecular-mechanism-of-pre-mrna-splicing
#11
REVIEW
Sjors Hw Scheres, Kiyoshi Nagai
The spliceosome is an intricate molecular machine which catalyses the removal of introns from eukaryotic mRNA precursors by two trans-esterification reactions (branching and exon ligation) to produce mature mRNA with uninterrupted protein coding sequences. The structures of the spliceosome in several key states determined by electron cryo-microscopy have greatly advanced our understanding of its molecular mechanism. The catalytic RNA core is formed during the activation of the fully assembled B to Bact complex and remains largely unchanged throughout the splicing cycle...
September 6, 2017: Current Opinion in Structural Biology
https://www.readbyqxmd.com/read/28881141/elongated-plant-virus-based-nanoparticles-for-enhanced-delivery-of-thrombolytic-therapies
#12
Andrzej Stanislaw Pitek, Yunmei Wang, Sahil Gulati, Huiyun Gao, Phoebe L Stewart, Daniel I Simon, Nicole F Steinmetz
Thrombotic cardiovascular disease, including acute myocardial infarction, ischemic stroke, and venous thromboembolic disease, is the leading cause of morbidity and mortality worldwide. While reperfusion therapy with thrombolytic agents reduces mortality from acute myocardial infarction and disability from stroke, thrombolysis is generally less effective than mechanical reperfusion and is associated with fatal intra-cerebral hemorrhage in up to 2-4% of patients. To address these limitations, we propose the tobacco mosaic virus (TMV)-based platform technology for targeted delivery of thrombolytic therapies...
September 7, 2017: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/28880323/wormlike-micelles-versus-water-soluble-polymers-as-rheology-modifiers-similarities-and-differences
#13
Ji Wang, Yujun Feng, Niti R Agrawal, Srinivasa R Raghavan
Wormlike micelles (WLMs) formed from surfactants have attracted much attention for their ability to thicken water in a manner similar to water-soluble polymers. It is known that WLMs are cylindrical filaments that can attain very long contour lengths (∼few μm), akin to chains of polymers with ultra-high molecular weights (UHMWs). In this study, we aim to make a direct comparison between the thickening capabilities of WLMs and UHMW polymers. The chosen surfactant is erucyl dimethyl amidopropyl betaine (EDAB), a C22-tailed zwitterionic surfactant known to form particularly long WLMs independent of salt...
September 7, 2017: Physical Chemistry Chemical Physics: PCCP
https://www.readbyqxmd.com/read/28878270/membrane-association-of-the-bacterial-riboregulator-hfq-and-functional-perspectives
#14
Antoine Malabirade, Javier Morgado-Brajones, Sylvain Trépout, Frank Wien, Ileana Marquez, Jérôme Seguin, Sergio Marco, Marisela Velez, Arluison Véronique
Hfq is a bacterial RNA binding protein that carries out several roles in genetic expression regulation, mainly at the post-transcriptional level. Previous studies have shown its importance in growth and virulence of bacteria. Here, we provide the direct observation of its ability to interact with membranes. This was established by co-sedimentation assay, cryo-transmission electron (cryo-TEM) and atomic force (AFM) microscopies. Furthermore, our results suggest a role for its C-terminus amyloidogenic domain in membrane disruption...
September 6, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28871556/lessons-from-a-tarantula-new-insights-into-muscle-thick-filament-and-myosin-interacting-heads-motif-structure-and-function
#15
REVIEW
Lorenzo Alamo, Natalia Koubassova, Antonio Pinto, Richard Gillilan, Andrey Tsaturyan, Raúl Padrón
The tarantula skeletal muscle X-ray diffraction pattern suggested that the myosin heads were helically arranged on the thick filaments. Electron microscopy (EM) of negatively stained relaxed tarantula thick filaments revealed four helices of heads allowing a helical 3D reconstruction. Due to its low resolution (5.0 nm), the unambiguous interpretation of densities of both heads was not possible. A resolution increase up to 2.5 nm, achieved by cryo-EM of frozen-hydrated relaxed thick filaments and an iterative helical real space reconstruction, allowed the resolving of both heads...
September 4, 2017: Biophysical Reviews
https://www.readbyqxmd.com/read/28870772/the-dissolvable-bead-a-novel-in-vitro-biofilm-model-for-evaluating-antimicrobial-resistance
#16
G F Dall, S T J Tsang, P J Gwynne, A J Wilkinson, A H R W Simpson, S J B Breusch, M P Gallagher
In vitro biofilm assays are a vital first step in the assessment of therapeutic effectiveness. Current biofilm models have been found to be limited by throughput, reproducibility, and cost. We present a novel in vitro biofilm model, utilising a sodium alginate substratum for surface biofilm colony formation, which can be readily dissolved for accurate evaluation of viable organisms. The dissolving bead biofilm assay was evaluated using a range of clinically relevant strains. The reproducibility and responsiveness of the assay to an antimicrobial challenge was assessed using standardised methods...
September 1, 2017: Journal of Microbiological Methods
https://www.readbyqxmd.com/read/28869968/structural-basis-of-msba-mediated-lipopolysaccharide-transport
#17
Wei Mi, Yanyan Li, Sung Hwan Yoon, Robert K Ernst, Thomas Walz, Maofu Liao
Lipopolysaccharide (LPS) in the outer membrane of Gram-negative bacteria is critical for the assembly of their cell envelopes. LPS synthesized in the cytoplasmic leaflet of the inner membrane is flipped to the periplasmic leaflet by MsbA, an ATP-binding cassette transporter. Despite substantial efforts, the structural mechanisms underlying MsbA-driven LPS flipping remain elusive. Here we use single-particle cryo-electron microscopy to elucidate the structures of lipid-nanodisc-embedded MsbA in three functional states...
September 14, 2017: Nature
https://www.readbyqxmd.com/read/28867613/high-resolution-cryo-em-maps-and-models-a%C3%A2-crystallographer-s-perspective
#18
Alexander Wlodawer, Mi Li, Zbigniew Dauter
The appearance of ten high-resolution cryoelectron microscopy (cryo-EM) maps of proteins, ribosomes, and viruses was compared with the experimentally phased crystallographic electron density maps of four proteins. We found that maps calculated at a similar resolution by the two techniques are quite comparable in their appearance, although cryo-EM maps, even when sharpened, seem to be a little less detailed. An analysis of models fitted to the cryo-EM maps indicated the presence of significant problems in almost all of them, including incorrect geometry, clashes between atoms, and discrepancies between the map density and the fitted models...
August 10, 2017: Structure
https://www.readbyqxmd.com/read/28859647/blob-ology-and-biology-of-cryo-em-an-interview-with-helen-saibil
#19
Helen R Saibil
Helen Saibil is Bernal Professor of Structural Biology at Birkbeck, University of London. After undergraduate work at McGill University, Canada, Helen moved to London for her PhD at Kings College. After stints at CEA Grenoble and the University of Oxford, she moved to Birkbeck where her lab studies the operation of macromolecular machinery-including molecular chaperones, protein folding/misfolding, and host cell interactions with pathogens. Helen is a Fellow of the Royal Society, Fellow of the Academy of Medical Sciences, and an Honorary Member of both the British Biophysical Society and the Royal Microscopical Society...
August 31, 2017: BMC Biology
https://www.readbyqxmd.com/read/28855679/pea-psii-lhcii-supercomplexes-form-pairs-by-making-connections-across-the-stromal-gap
#20
Pascal Albanese, Roberto Melero, Benjamin D Engel, Alessandro Grinzato, Paola Berto, Marcello Manfredi, Angelica Chiodoni, Javier Vargas, Carlos Óscar Sánchez Sorzano, Emilio Marengo, Guido Saracco, Giuseppe Zanotti, Jose-Maria Carazo, Cristina Pagliano
In higher plant thylakoids, the heterogeneous distribution of photosynthetic protein complexes is a determinant for the formation of grana, stacks of membrane discs that are densely populated with Photosystem II (PSII) and its light harvesting complex (LHCII). PSII associates with LHCII to form the PSII-LHCII supercomplex, a crucial component for solar energy conversion. Here, we report a biochemical, structural and functional characterization of pairs of PSII-LHCII supercomplexes, which were isolated under physiologically-relevant cation concentrations...
August 30, 2017: Scientific Reports
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