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Cryo electron microscopy

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https://www.readbyqxmd.com/read/28819229/cryo-em-analysis-of-homodimeric-full-length-lrrk2-and-lrrk1-protein-complexes
#1
Kushal Sejwal, Mohamed Chami, Hervé Rémigy, Renée Vancraenenbroeck, William Sibran, Rosmarie Sütterlin, Paul Baumgartner, Robert McLeod, Marie-Christine Chartier-Harlin, Veerle Baekelandt, Henning Stahlberg, Jean-Marc Taymans
Leucine-rich repeat kinase 2 (LRRK2) is a large multidomain protein implicated in the pathogenesis of both familial and sporadic Parkinson's disease (PD), and currently one of the most promising therapeutic targets for drug design in Parkinson's disease. In contrast, LRRK1, the closest homologue to LRRK2, does not play any role in PD. Here, we use cryo-electron microscopy (cryo-EM) and single particle analysis to gain structural insight into the full-length dimeric structures of LRRK2 and LRRK1. Differential scanning fluorimetry-based screening of purification buffers showed that elution of the purified LRRK2 protein in a high pH buffer is beneficial in obtaining high quality cryo-EM images...
August 17, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28817608/data-driven-coarse-graining-of-large-biomolecular-structures
#2
Yi-Ling Chen, Michael Habeck
Advances in experimental and computational techniques allow us to study the structure and dynamics of large biomolecular assemblies at increasingly higher resolution. However, with increasing structural detail it can be challenging to unravel the mechanism underlying the function of molecular machines. One reason is that atomistic simulations become computationally prohibitive. Moreover it is difficult to rationalize the functional mechanism of systems composed of tens of thousands to millions of atoms by following each atom's movements...
2017: PloS One
https://www.readbyqxmd.com/read/28815272/gold-nanoparticle-based-localized-surface-plasmon-immunosensor-for-staphylococcal-enterotoxin-a-sea-detection
#3
Maroua Ben Haddada, David Hu, Michèle Salmain, Lu Zhang, Chen Peng, Yi Wang, Bo Liedberg, Souhir Boujday
We describe the engineering of stable gold nanoparticle (AuNP) bioconjugates for the detection of staphylococcal enterotoxin A (SEA) using localized surface plasmon resonance (LSPR). Two types of AuNP bioconjugates were prepared by covalently attaching anti-SEA antibody (Ab) or SEA to AuNPs. This was achieved by reacting Traut's reagent with lysine residues of both proteins to generate thiol groups that bind to gold atoms on the AuNP surface. These bioconjugates were characterized in-depth by absorption spectroscopy, cryo-transmission electron microscopy, dynamic light scattering, and zeta potential measurements...
August 16, 2017: Analytical and Bioanalytical Chemistry
https://www.readbyqxmd.com/read/28811473/plant-made-polio-type-3-stabilized-vlps-a-candidate-synthetic-polio-vaccine
#4
Johanna Marsian, Helen Fox, Mohammad W Bahar, Abhay Kotecha, Elizabeth E Fry, David I Stuart, Andrew J Macadam, David J Rowlands, George P Lomonossoff
Poliovirus (PV) is the causative agent of poliomyelitis, a crippling human disease known since antiquity. PV occurs in two distinct antigenic forms, D and C, of which only the D form elicits a robust neutralizing response. Developing a synthetically produced stabilized virus-like particle (sVLP)-based vaccine with D antigenicity, without the drawbacks of current vaccines, will be a major step towards the final eradication of poliovirus. Such a sVLP would retain the native antigenic conformation and the repetitive structure of the original virus particle, but lack infectious genomic material...
August 15, 2017: Nature Communications
https://www.readbyqxmd.com/read/28809539/polymersomes-with-endosomal-ph-induced-vesicle-to-micelle-morphology-transition-and-a-potential-application-for-controlled-doxorubicin-delivery
#5
Turgay Yildirim, Anja Traeger, Pelin Sungur, Stephanie Hoeppener, Carolin Kellner, Ilknur Yildirim, David Pretzel, Stephanie Schubert, Ulrich S Schubert
A series of pH responsive block copolymers were synthesized via the reversible addition-fragmentation chain transfer (RAFT) polymerization of 3,4-dihydro-2H-pyran (DHP) protected 2-hydroxyethyl methacrylate (HEMA) (2-((tetrahydro-2H-pyran-2-yl)oxy)ethyl methacrylate (THP-HEMA)) and 2-(dimethylamino) ethyl methacrylate (DMAEMA) using p(THP-HEMA) as a macro chain transfer agent (mCTA). The degree of polymerization (DP) of the p(THP-HEMA) block was fixed to 35, whereas the DP of the p(DMAEMA) block was systematically varied from 21 to 50...
August 15, 2017: Biomacromolecules
https://www.readbyqxmd.com/read/28809395/cryo-em-structures-of-the-80s-ribosomes-from-human-parasites-trichomonas-vaginalis-and-toxoplasma-gondii
#6
Zhifei Li, Qiang Guo, Lvqin Zheng, Yongsheng Ji, Yi-Ting Xie, De-Hua Lai, Zhao-Rong Lun, Xun Suo, Ning Gao
As an indispensable molecular machine universal in all living organisms, the ribosome has been selected by evolution to be the natural target of many antibiotics and small-molecule inhibitors. High-resolution structures of pathogen ribosomes are crucial for understanding the general and unique aspects of translation control in disease-causing microbes. With cryo-electron microscopy technique, we have determined structures of the cytosolic ribosomes from two human parasites, Trichomonas vaginalis and Toxoplasma gondii, at resolution of 3...
August 15, 2017: Cell Research
https://www.readbyqxmd.com/read/28808031/changes-in-aggregation-states-of-light-harvesting-complexes-as-a-mechanism-for-modulating-energy-transfer-in-desert-crust-cyanobacteria
#7
Leeat Bar Eyal, Reza Ranjbar Choubeh, Eyal Cohen, Ido Eisenberg, Carmen Tamburu, Márta Dorogi, Renata Ünnep, Marie-Sousai Appavou, Reinat Nevo, Uri Raviv, Ziv Reich, Győző Garab, Herbert van Amerongen, Yossi Paltiel, Nir Keren
In this paper we propose an energy dissipation mechanism that is completely reliant on changes in the aggregation state of the phycobilisome light-harvesting antenna components. All photosynthetic organisms regulate the efficiency of excitation energy transfer (EET) to fit light energy supply to biochemical demands. Not many do this to the extent required of desert crust cyanobacteria. Following predawn dew deposition, they harvest light energy with maximum efficiency until desiccating in the early morning hours...
August 14, 2017: Proceedings of the National Academy of Sciences of the United States of America
https://www.readbyqxmd.com/read/28803776/mechanism-of-enzyme-repair-by-the-aaa-chaperone-rubisco-activase
#8
Javaid Y Bhat, Goran Miličić, Gabriel Thieulin-Pardo, Andreas Bracher, Andrew Maxwell, Susanne Ciniawsky, Oliver Mueller-Cajar, John R Engen, F Ulrich Hartl, Petra Wendler, Manajit Hayer-Hartl
How AAA+ chaperones conformationally remodel specific target proteins in an ATP-dependent manner is not well understood. Here, we investigated the mechanism of the AAA+ protein Rubisco activase (Rca) in metabolic repair of the photosynthetic enzyme Rubisco, a complex of eight large (RbcL) and eight small (RbcS) subunits containing eight catalytic sites. Rubisco is prone to inhibition by tight-binding sugar phosphates, whose removal is catalyzed by Rca. We engineered a stable Rca hexamer ring and analyzed its functional interaction with Rubisco...
July 20, 2017: Molecular Cell
https://www.readbyqxmd.com/read/28801059/editorial-overview-cryo-electron-microscopy-exciting-advances-in-cryoem-herald-a-new-era-in-structural-biology
#9
EDITORIAL
Wah Chiu, Kenneth H Downing
No abstract text is available yet for this article.
August 8, 2017: Current Opinion in Structural Biology
https://www.readbyqxmd.com/read/28799758/ph-sensitive-interactions-between-cellulose-nanocrystals-and-dopc-liposomes
#10
Yotam Navon, Harisoa Radavidson, Jean-Luc Putaux, Bruno Jean, Laurent Heux
The interaction of DOPC vesicles with cellulose nanocrystals (CNCs) have been investigated using several complementary techniques. Dynamic light scattering, zeta-potential, cryo-transmission electron microscopy and isothermal titration calorimetry (ITC) analyses confirmed the formation of pH-dependent CNC-liposome complexes. ITC was used to characterize the thermodynamic properties of this interaction. Positive values of enthalpy were found at pH lower than 5 where the charge sign of the constituents was opposite...
August 11, 2017: Biomacromolecules
https://www.readbyqxmd.com/read/28798962/structural-pathway-of-regulated-substrate-transfer-and-threading-through-an-hsp100-disaggregase
#11
Célia Deville, Marta Carroni, Kamila B Franke, Maya Topf, Bernd Bukau, Axel Mogk, Helen R Saibil
Refolding aggregated proteins is essential in combating cellular proteotoxic stress. Together with Hsp70, Hsp100 chaperones, including Escherichia coli ClpB, form a powerful disaggregation machine that threads aggregated polypeptides through the central pore of tandem adenosine triphosphatase (ATPase) rings. To visualize protein disaggregation, we determined cryo-electron microscopy structures of inactive and substrate-bound ClpB in the presence of adenosine 5'-O-(3-thiotriphosphate), revealing closed AAA+ rings with a pronounced seam...
August 2017: Science Advances
https://www.readbyqxmd.com/read/28795863/a-new-era-of-studying-p53-mediated-transcription-activation
#12
Wei-Li Liu, Robert A Coleman, Sameer K Singh
To prevent tumorigenesis, p53 stimulates transcription by facilitating the recruitment of the transcription machinery on target gene promoters. Cryo-Electron Microscopy studies on p53-bound RNA Polymerase II (Pol II) reveal that p53 structurally regulates Pol II to affect its DNA binding and elongation, providing new insights into p53-mediated transcriptional regulation.
August 10, 2017: Transcription
https://www.readbyqxmd.com/read/28795512/single-particle-cryo-em-improved-ab-initio-3d-reconstruction-with-simple-prime
#13
Cyril F Reboul, Michael Eager, Dominika Elmlund, Hans Elmlund
Cryogenic electron microscopy (cryo-EM) and single-particle analysis now enables the determination of high-resolution structures of macromolecular assemblies that have resisted X-ray crystallography and other approaches. We developed the SIMPLE open-source image-processing suite for analysing cryo-EM images of single-particles. A core component of SIMPLE is the probabilistic PRIME algorithm for identifying clusters of images in 2D and determine relative orientations of single-particle projections in 3D. Here we extend our previous work on PRIME and introduce new stochastic optimisation algorithms that improve the robustness of the approach...
August 10, 2017: Protein Science: a Publication of the Protein Society
https://www.readbyqxmd.com/read/28794948/glucosomes-glycosylated-vesicle-in-vesicle-aggregates-in-water-from-ph-responsive-microbial-glycolipid
#14
Niki Baccile, Patrick Le Griel, Sylvain Prévost, Bernd Everaert, Inge N A Van Bogaert, Sophie Roelants, Wim Soetaert
Vesicle-in-vesicle self-assembled containers, or vesosomes, are promising alternatives to liposomes because of their possible hierarchical encapsulation and high stability. We report herein the first example of sugar-based vesicles-in-vesicles, which we baptize glucosomes. These were prepared by using a natural microbial glycolipid (branched C22 sophorolipid) extracted from the culture medium of the yeast Pseudohyphozyma bogoriensis. Glucosomes spontaneously formed in water between pH 6 and pH 4 at room temperature, without the requirement of any additive...
August 2017: ChemistryOpen
https://www.readbyqxmd.com/read/28782000/particle-segmentation-algorithm-for-flexible-single-particle-reconstruction
#15
Qiang Zhou, Niyun Zhou, Hong-Wei Wang
As single particle cryo-electron microscopy has evolved to a new era of atomic resolution, sample heterogeneity still imposes a major limit to the resolution of many macromolecular complexes, especially those with continuous conformational flexibility. Here, we describe a particle segmentation algorithm towards solving structures of molecules composed of several parts that are relatively flexible with each other. In this algorithm, the different parts of a target molecule are segmented from raw images according to their alignment information obtained from a preliminary 3D reconstruction and are subjected to single particle processing in an iterative manner...
2017: Biophysics Reports
https://www.readbyqxmd.com/read/28781997/using-integrated-correlative-cryo-light-and-electron-microscopy-to-directly-observe-syntaphilin-immobilized-neuronal-mitochondria-in-situ
#16
Shengliu Wang, Shuoguo Li, Gang Ji, Xiaojun Huang, Fei Sun
Correlative cryo-fluorescence and cryo-electron microscopy (cryo-CLEM) system has been fast becoming a powerful technique with the advantage to allow the fluorescent labeling and direct visualization of the close-to-physiologic ultrastructure in cells at the same time, offering unique insights into the ultrastructure with specific cellular function. There have been various engineered ways to achieve cryo-CLEM including the commercial FEI iCorr system that integrates fluorescence microscope into the column of transmission electron microscope...
2017: Biophysics Reports
https://www.readbyqxmd.com/read/28781996/opinion-hazards-faced-by-macromolecules-when-confined-to-thin-aqueous-films
#17
REVIEW
Robert M Glaeser, Bong-Gyoon Han
Samples prepared for single-particle electron cryo-microscopy (cryo-EM) necessarily have a very high surface-to-volume ratio during the short period of time between thinning and vitrification. During this time, there is an obvious risk that macromolecules of interest may adsorb to the air-water interface with a preferred orientation, or that they may even become partially or fully unfolded at the interface. In addition, adsorption of macromolecules to an air-water interface may occur even before thinning. This paper addresses the question whether currently used methods of sample preparation might be improved if one could avoid such interfacial interactions...
2017: Biophysics Reports
https://www.readbyqxmd.com/read/28780349/implications-of-coordinated-cell-body-rotations-for-leptospira-motility
#18
Kyosuke Takabe, Akihiro Kawamoto, Hajime Tahara, Seishi Kudo, Shuichi Nakamura
The spirochete Leptospira has a coiled cell body and two periplasmic flagella (PFs) that reside beneath the outer sheath. PFs extend from each end of the cell body and are attached to the right-handed spiral protoplasmic cylinder (PC) via a connection with the flagellar motor embedded in the inner membrane. PFs bend each end of the cell body into left-handed spiral (S) or planar hook (H) shapes, allowing leptospiral cells to swim using combined anterior S-end and posterior H-end gyrations with PC rotations...
August 2, 2017: Biochemical and Biophysical Research Communications
https://www.readbyqxmd.com/read/28779844/saposin-lipoprotein-scaffolds-for-structure-determination-of-membrane-transporters
#19
Joseph A Lyons, Andreas Bøggild, Poul Nissen, Jens Frauenfeld
Membrane proteins depend on their natural lipid environment for function, which makes them more difficult to study in isolation. A number of approaches that mimic the lipid bilayer of biological membranes have been described (nanodiscs, SMALPs), enabling novel ways to assay activity and elucidate structures of this important class of proteins. More recently, the use of saposin A, a protein that is involved in lipid transport, to form Salipro (saposin-lipid-protein) complexes was demonstrated for a range of membrane protein targets (Frauenfeld et al...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28768871/isolation-and-characterization-of-metallosphaera-turreted-icosahedral-virus-mtiv-a-founding-member-of-a-new-family-of-archaeal-viruses
#20
Cassia Wagner, Vijay Reddy, Francisco Asturias, Maryam Khoshouei, John E Johnson, Pilar Manrique, Jacob Munson-McGee, Wolfgang Baumeister, C Martin Lawrence, Mark J Young
Our understanding of archaeal virus diversity and structure is just beginning to emerge. Here we describe a new archaeal virus, tentatively named Metallosphaera turreted icosahedral virus (MTIV), that was isolated from an acidic hot spring in Yellowstone National Park, USA. Two strains of the virus were identified and found to replicate in an archaeal host species closely related to Metallosphaera yellowstonensis Each strain encodes for a 9.8-9.9 kb, linear dsDNA genome with large inverted terminal repeats...
August 2, 2017: Journal of Virology
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