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Journal of Structural Biology

Ahmad Ali-Ahmad, Marie-Line Garron, Véronique Zamboni, Nicolas Lenfant, Didier Nurizzo, Bernard Henrissat, Jean-Guy Berrin, Yves Bourne, Florence Vincent
Bacteria from the human gut are equipped with an arsenal of carbohydrate-active enzymes that degrade dietary and host-derived glycans. In this study, we present the 2.5Å resolution crystal structure of a member (GH39wh2) from the human gut bacteria Bacteroides cellulosilyticus WH2 representative of a new subgroup within family GH39. Together with 6 other GHs, GH39wh2 belongs to a polysaccharide utilization locus (PUL) that could be involved in detecting, binding and hydrolysing a specific carbohydrate species from the intestinal tract...
November 24, 2016: Journal of Structural Biology
Stefan A Arnold, Stefan Albiez, Andrej Bieri, Anastasia Syntychaki, Ricardo Adaixo, Robb A McLeod, Kenneth N Goldie, Henning Stahlberg, Thomas Braun
We present a sample preparation method for cryo-electron microscopy (cryo-EM) that requires only 3 to 20 nanoliters of sample to prepare a cryo-EM grid, depending on the protocol used. The sample is applied and spread on the grid by a microcapillary. The procedure does not involve any blotting steps, and real-time monitoring allows the water film thickness to be assessed and decreased to an optimum value prior to vitrification. We demonstrate that the method is suitable for high-resolution cryo-EM and will enable alternative electron microscopy approaches, such as single-cell visual proteomics...
November 15, 2016: Journal of Structural Biology
Rui Yan, Kunpeng Li, Wen Jiang
Minimization of the astigmatism of the objective lens is a critical daily instrument alignment task essential for high resolution TEM imaging. Fast and sensitive detection of astigmatism is needed to provide real-time feedback and adjust the stigmators to efficiently reduce astigmatism. Currently the method used by many microscopists isto visually examine the roundness of a diffractogram (Thon rings) and iteratively adjust the stigmators to make the Thon rings circular. This subjective method is limited by poor sensitivity and potentially biased by the astigmatism of human eyes...
November 9, 2016: Journal of Structural Biology
María L Cerutti, Lisandro H Otero, Clara Smal, Leonardo Pellizza, Fernando A Goldbaum, Sebastián Klinke, Martín Aran
The Pfam PF04536 TPM_phosphatase family is a broadly conserved family of domains found across prokaryotes, plants and invertebrates. Despite having a similar protein fold, members of this family have been implicated in diverse cellular processes and found in varied subcellular localizations. Very recently, the biochemical characterization of two evolutionary divergent TPM domains has shown that they are able to hydrolyze phosphate groups from different substrates. However, there are still incorrect functional annotations and uncertain relationships between the structure and function of this family of domains...
October 31, 2016: Journal of Structural Biology
Bogdan I Costescu, Sebastian Sturm, Frauke Gräter
Protein unfolding often does not obey a simple two-state behavior. Previous single molecule force spectroscopy studies demonstrated stretched exponential kinetics of protein unfolding under a constant pulling force, the molecular origin of which remains subject to debate. We here set out to extensively sample the mechanical unfolding of ubiquitin and NuG2 by Molecular Dynamics (MD) simulations. Both proteins show kinetics best fit by stretched exponentials, with stretching exponents similar to those found in experiments, even though static disorder is absent in our short MD simulations...
October 19, 2016: Journal of Structural Biology
Thai V Hoang, Caroline Kizilyaprak, Danièle Spehner, Bruno M Humbel, Patrick Schultz
Focused Ion Beam milling combined with Scanning Electron Microscopy is a powerful tool to determine the 3-D organization of whole cells and tissue at an isotropic resolution of 3-5nm. This opens the possibility to quantify several cellular parameters and to provide detailed phenotypic information in normal or disease states. Here we describe Biocomputing methods to extract in an automated way characteristic features of mouse rod photoreceptor nuclei such as the shape and the volume of the nucleus; the proportion of heterochromatin; the number, density and distribution of nuclear pore complexes (NPC)...
October 8, 2016: Journal of Structural Biology
M S Fernández, F Valenzuela, J I Arias, A Neira-Carrillo, J L Arias
Biominerals are inorganic-organic hybrid composites formed via self-assembled bottom up processes under mild conditions. Biominerals show interesting physical properties, controlled hierarchical structures and robust remodeling or repair mechanisms. Biological processes associated with biominerals remain to be developed into practical engineering processes. Therefore, the formation of biominerals is inspiring for the design of materials, especially those fabricated at ambient temperatures. The study described herein involves the influence of chicken outer eggshell membrane on the type of calcium carbonate (CaCO3) polymorph deposited on the shell of the land snail Helix aspersa during the repair process after an injury...
October 4, 2016: Journal of Structural Biology
Thomas H Sharp, Frank G A Faas, Abraham J Koster, Piet Gros
Phase plates in cryo-electron tomography (cryoET) improve contrast, increasing the ability to discern separate molecules and molecular complexes in dense biomolecular environments. Here, we applied this new technology to the activation of the human complement system. Binding of C1 to antigen-antibody complexes initiates a cascade of proteolytic events that deposits molecules onto adjacent surfaces and terminates with the formation of membrane-attack-complex (MAC) pores in the targeted membranes. We imaged steps in this process using a Volta phase plate mounted on a Titan Krios equipped with a Falcon-II direct electron detector...
September 20, 2016: Journal of Structural Biology
Matthew A Hood, Hanna Leemreize, André Scheffel, Damien Faivre
During biomineralization, organisms control the formation and morphology of a mineral using biomacromolecules. The biomacromolecules that most strongly interact with the growing crystals frequently get occluded within. Such an observation has been recently obtained for the calcium carbonate producing coccolithophore species Pleurochrysis carterae. Coccolithophores are unicellular algae that produce calcified scales built from complex-shaped calcite crystals, termed coccoliths. It is unclear how widespread the phenomenon of biomacromolecular occlusion within calcite crystals is in calcifying haptophytes such as coccolithophores...
September 16, 2016: Journal of Structural Biology
Silvia Pabisch, Chika Akabane, Wolfgang Wagermaier, Andreas Roschger, Taku Ogura, Ryo Hyodo, Shinsuke Kataoka, Norio Tobori, Tomomichi Okano, Shinya Murakami, Peter Fratzl, Richard Weinkamer
Alveolar bone - the bony ridge containing the tooth sockets - stands out by its remodeling activity where bone is being formed and resorbed at a much higher rate than in any other bony tissue. Teeth that are anchored in the jaw through the periodontal ligament exert very large localized loads during mastication that could lead to a unique adaptation of the collagen/mineral structure in the bone. Our aim was to characterize the nanostructure of alveolar bone and to determine the influence of diabetes on structural characteristics of the mineralized matrix...
September 13, 2016: Journal of Structural Biology
Carlos Rodriguez-Navarro, Encarnacion Ruiz-Agudo, Joe Harris, Stephan E Wolf
Recent research has shown that biominerals and their biomimetics (i) typically form via an amorphous precursor phase, and (ii) commonly display a nanogranular texture. Apparently, these two key features are closely related, underlining the fact that the formation of biominerals and their biomimetics does not necessarily follow classical crystallization routes, and leaves a characteristic nanotextural imprint which may help to disclose their origins and formation mechanisms. Here we present a general overview of the current theories and models of nonclassical crystallization and their applicability for the advance of our current understanding of biomineralization and biomimetic mineralization...
September 9, 2016: Journal of Structural Biology
Lionel Sacconnay, Pierre-Alain Carrupt, Alessandra Nurisso
In recent years, sirtuins (SIRTs), members of histone deacetylases (HDACs) class III, have been found to modulate cellular processes related to the development of human aging-related pathologies (i.e. cancer, neurodegeneration, metabolic disorders). Several crystallographic structures and computational studies have shed light into their catalytic mechanism of action, identifying also the structural elements for the design of selective drug candidates. In this review, we first aim at summarizing the structural features characterizing human SIRTs...
December 2016: Journal of Structural Biology
J L Vilas, J Navas, J Gómez-Blanco, J M de la Rosa-Trevín, R Melero, I Peschiera, I Ferlenghi, J Cuenca, R Marabini, J M Carazo, J Vargas, C O S Sorzano
Random conical tilt (RCT) and orthogonal tilt reconstruction (OTR) are two remarkable methods for reconstructing the three-dimensional structure of macromolecules at low resolution. These techniques use two images at two different sample tilts. One of the most demanding steps in these methods at the image processing level is to identify corresponding particles on both micrographs, and manual or semiautomatic matching methods are usually used. Here we present an approach to solve this bottleneck with a fully automatic method for assigning particle tilt pairs...
December 2016: Journal of Structural Biology
Vladan Lučić, Rubén Fernández-Busnadiego, Ulrike Laugks, Wolfgang Baumeister
Molecular complexes, arguably the basic units carrying cellular function, can be visualized directly in their native environment by cryo-electron tomography. Here we describe a procedure for the detection of small, pleomorphic membrane-bound molecular complexes in cryo-tomograms by a hierarchical connectivity segmentation. Validation on phantom and real data showed above 90% true positive rates. This segmentation procedure is implemented in the Pyto software package, together with methods for quantitative characterization and classification of complexes detected by our segmentation procedure and for statistical analysis between experimental conditions...
December 2016: Journal of Structural Biology
J Vargas, E Franken, C O S Sorzano, J Gomez-Blanco, R Schoenmakers, A J Koster, J M Carazo
Automatic or semiautomatic data collection approaches on a transmission electron microscope (TEM) for Single Particle Analysis, capable of acquiring large datasets composed of only high quality images, are of great importance to obtain 3D density maps with the highest resolution possible. Typically, this task is performed by an experienced microscopist, who manually decides to keep or discard images according to subjective criteria. Therefore, this methodology is slow, intensive in human work and subjective...
December 2016: Journal of Structural Biology
Ake Vastermark, Adelle Driker, Jingwei Weng, Xiaochun Li, Jiawei Wang, Milton H Saier
We propose that the alternative crystal forms of outward open UlaA (which are experimental, not simulated, and contain the substrate in the cavity) can be used to interpret/validate the MD results from MalT (the substrate capture step, which involves the mobile second TMSs of the V-motifs, TMSs 2 and 7). Since the crystal contacts are the same between the two alternative crystal forms of outward open UlaA, the striking biological differences noted, including rearranged hydrogen bonds and salt bridge coordination, are not attributable to crystal packing differences...
December 2016: Journal of Structural Biology
Netta Vidavsky, Anat Akiva, Ifat Kaplan-Ashiri, Katya Rechav, Lia Addadi, Steve Weiner, Andreas Schertel
Many important biological questions can be addressed by studying in 3D large volumes of intact, cryo fixed hydrated tissues (⩾10,000μm(3)) at high resolution (5-20nm). This can be achieved using serial FIB milling and block face surface imaging under cryo conditions. Here we demonstrate the unique potential of the cryo-FIB-SEM approach using two extensively studied model systems; sea urchin embryos and the tail fin of zebrafish larvae. We focus in particular on the environment of mineral deposition sites...
December 2016: Journal of Structural Biology
Takatsugu Miyazaki, Atsushi Nishikawa, Takashi Tonozuka
Glycoside hydrolases are divided into two groups, known as inverting and retaining enzymes, based on their hydrolytic mechanisms. Glycoside hydrolase family 63 (GH63) is composed of inverting α-glycosidases, which act mainly on α-glucosides. We previously found that Escherichia coli GH63 enzyme, YgjK, can hydrolyze 2-O-α-d-glucosyl-d-galactose. Two constructed glycosynthase mutants, D324N and E727A, which catalyze the transfer of a β-glucosyl fluoride donor to galactose, lactose, and melibiose. Here, we determined the crystal structures of D324N and E727A soaked with a mixture of glucose and lactose at 1...
December 2016: Journal of Structural Biology
Peter Rez, Thomas Larsen, Michael Elbaum
Scanning transmission electron microscope (STEM) imaging has recently been applied to the cryo-tomography of thick biological specimens. As previously shown for plastic sections, STEM has a number of advantages for cryo-imaging compared to conventional wide-field TEM imaging. STEM is insensitive to phase coherence and is therefore suitable for much thicker specimens than TEM. Imaging in focus, with a long depth of field, also circumvents the complications of an oscillatory contrast transfer function and missing information at low spatial frequencies...
December 2016: Journal of Structural Biology
Xiaojing Wang, Baobin Li, Yu Guo, Shu Shen, Liang Zhao, Peisheng Zhang, Yuna Sun, Sen-Fang Sui, Fei Deng, Zhiyong Lou
Negative-sense single-strand RNA (-ssRNA) viruses comprise a large family of pathogens that cause severe human infectious diseases. All -ssRNA viruses encode a nucleocapsid protein (NP) to encapsidate the viral genome, which, together with polymerase, forms a ribonucleoprotein complex (RNP) that is packaged into virions and acts as the template for viral replication and transcription. In our previous work, we solved the monomeric structure of NP encoded by Crimean-Congo hemorrhagic fever virus (CCHFV), which belongs to the Nairovirus genus within the Bunyaviridae family, and revealed its unusual endonuclease activity...
December 2016: Journal of Structural Biology
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