The Experts below are selected from a list of 2934 Experts worldwide ranked by ideXlab platform
Ahmed H. Zewail - One of the best experts on this subject based on the ideXlab platform.
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structural dynamics of nanoscale gold by ultrafast Electron Crystallography
Chemical Physics Letters, 2011Co-Authors: Sascha Schafer, Wenxi Liang, Ahmed H. ZewailAbstract:By employing ultrafast Electron Crystallography in a transmission geometry for ultra-thin (2–3 nm) gold, here we show that structural dynamics of the transverse atomic motions and the atomic displacements around the equilibrium position can be separated from the measured change in Bragg diffraction, the positions and intensities of the peaks, respectively. The rate of intensity change provides the Electron-lattice equilibration time whereas the observed lattice expansion, which occurs on a slower time scale, maps the delayed response of transverse lattice strain. These textbook-type results provide the microscopic stress–strain profile that is critical for understanding dynamical deformations and the effect of morphological structures at surfaces.
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structural preablation dynamics of graphite observed by ultrafast Electron Crystallography
Physical Review Letters, 2008Co-Authors: Fabrizio Carbone, Peter Baum, Petra Rudolf, Ahmed H. ZewailAbstract:By means of time-resolved Electron Crystallography, we report direct observation of the structural dynamics of graphite, providing new insights into the processes involving coherent lattice motions and ultrafast graphene ablation. When graphite is excited by an ultrashort laser pulse, the excited carriers reach their equilibrium in less then one picosecond by transferring heat to a subset of strongly coupled optical phonons. The time-resolved diffraction data show that on such a time scale the crystal undergoes a contraction whose velocity depends on the excitation fluence. The contraction is followed by a large expansion which, at sufficiently high fluence, leads to the ablation of entire graphene layers, as recently predicted theoretically.
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nonequilibrium phase transitions in cuprates observed by ultrafast Electron Crystallography
Science, 2007Co-Authors: Nuh Gedik, Dingshyue Yang, G Logvenov, I Bozovic, Ahmed H. ZewailAbstract:Nonequilibrium phase transitions, which are defined by the formation of macroscopic transient domains, are optically dark and cannot be observed through conventional temperature- or pressure-change studies. We have directly determined the structural dynamics of such a nonequilibrium phase transition in a cuprate superconductor. Ultrafast Electron Crystallography with the use of a tilted optical geometry technique afforded the necessary atomic-scale spatial and temporal resolutions. The observed transient behavior displays a notable “structural isosbestic” point and a threshold effect for the dependence of c-axis expansion (Δc) on fluence (F), with Δc/F = 0.02 angstrom/(millijoule per square centimeter). This threshold for photon doping occurs at ∼0.12 photons per copper site, which is unexpectedly close to the density (per site) of chemically doped carriers needed to induce superconductivity.
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Ultrafast Electron Crystallography of Phospholipids
Angewandte Chemie, 2006Co-Authors: Songye Chen, Marco T. Seidel, Ahmed H. ZewailAbstract:The structure and dynamics of monolayer and bilayer (see picture) phospholipids have been studied with spatiotemporal resolutions by ultrafast Electron Crystallography. The expansion and restructuring of the chains were observed after a femtosecond temperature jump in the substrate, and a transient structural ordering was revealed. The atomic forces were identified to be coherent in the non-equilibrium state of the assembly.
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atomic scale dynamical structures of fatty acid bilayers observed by ultrafast Electron Crystallography
Proceedings of the National Academy of Sciences of the United States of America, 2005Co-Authors: Songye Chen, Marco T. Seidel, Ahmed H. ZewailAbstract:The structure and dynamics of a biological model bilayer are reported with atomic-scale resolution by using ultrafast Electron Crystallography. The bilayer was deposited as a Langmuir-Blodgett structure of arachidic (eicosanoic) fatty acids with the two chains containing 40 carbon atoms (≈50 A), on a hydrophobic substrate, the hydrogen terminated silicon(111) surface. We determined the structure of the 2D assembly, establishing the orientation of the chains and the subunit cell of the CH2 distances: a0 = 4.7 A, b0 = 8.0 A, and c0 = 2.54 A. For structural dynamics, the diffraction frames were taken every 1 picosecond after a femtosecond temperature jump. The observed motions, with sub-A resolution and monolayer sensitivity, clearly indicate the coherent anisotropic expansion of the bilayer solely along the aliphatic chains, followed by nonequilibrium contraction and restructuring at longer times. This motion is indicative of a nonlinear behavior among the anharmonically coupled bonds on the ultrashort time scale and energy redistribution and diffusion on the longer time scale. The ability to observe such atomic motions of complex structures and at interfaces is a significant leap forward for the determination of macromolecular dynamical structures by using ultrafast Electron Crystallography.
Tamir Gonen - One of the best experts on this subject based on the ideXlab platform.
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Overview of Electron Crystallography of membrane proteins: crystallization and screening strategies using negative stain Electron microscopy.
Current protocols in protein science, 2020Co-Authors: Brent L Nannenga, Matthew G Iadanza, Breanna S Vollmar, Tamir GonenAbstract:Electron cryomicroscopy, or cryoEM, is an emerging technique for studying the three-dimensional structures of proteins and large macromolecular machines. Electron Crystallography is a branch of cryoEM in which structures of proteins can be studied at resolutions that rival those achieved by X-ray Crystallography. Electron Crystallography employs two-dimensional crystals of a membrane protein embedded within a lipid bilayer. The key to a successful Electron crystallographic experiment is the crystallization, or reconstitution, of the protein of interest. This unit describes ways in which protein can be expressed, purified, and reconstituted into well-ordered two-dimensional crystals. A protocol is also provided for negative stain Electron microscopy as a tool for screening crystallization trials. When large and well-ordered crystals are obtained, the structures of both protein and its surrounding membrane can be determined to atomic resolution.
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from Electron Crystallography of 2d crystals to microed of 3d crystals
Current Opinion in Colloid and Interface Science, 2018Co-Authors: Michael W Martynowycz, Tamir GonenAbstract:Electron Crystallography is widespread in material science applications, but for biological samples its use has been restricted to a handful of examples where two-dimensional (2D) crystals or helical samples were studied either by Electron diffraction and/or imaging. Electron Crystallography in cryoEM, was developed in the mid-1970s and used to solve the structure of several membrane proteins and some soluble proteins. In 2013, a new method for cryoEM was unveiled and named Micro-crystal Electron Diffraction, or MicroED, which is essentially three-dimensional (3D) Electron Crystallography of microscopic crystals. This method uses truly 3D crystals, that are about a billion times smaller than those typically used for X-ray Crystallography, for Electron diffraction studies. There are several important differences and some similarities between Electron Crystallography of 2D crystals and MicroED. In this review, we describe the development of these techniques, their similarities and differences, and offer our opinion of future directions in both fields.
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three dimensional Electron Crystallography of protein microcrystals
eLife, 2013Co-Authors: Brent L Nannenga, Matthew G Iadanza, Tamir GonenAbstract:X-ray Crystallography has been used to work out the atomic structure of a large number of proteins. In a typical X-ray Crystallography experiment, a beam of X-rays is directed at a protein crystal, which scatters some of the X-ray photons to produce a diffraction pattern. The crystal is then rotated through a small angle and another diffraction pattern is recorded. Finally, after this process has been repeated enough times, it is possible to work backwards from the diffraction patterns to figure out the structure of the protein. The crystals used for X-ray Crystallography must be large to withstand the damage caused by repeated exposure to the X-ray beam. However, some proteins do not form crystals at all, and others only form small crystals. It is possible to overcome this problem by using extremely short pulses of X-rays, but this requires a very large number of small crystals and ultrashort X-ray pulses are only available at a handful of research centers around the world. There is, therefore, a need for other approaches that can determine the structure of proteins that only form small crystals. Electron Crystallography is similar to X-ray Crystallography in that a protein crystal scatters a beam to produce a diffraction pattern. However, the interactions between the Electrons in the beam and the crystal are much stronger than those between the X-ray photons and the crystal. This means that meaningful amounts of data can be collected from much smaller crystals. However, it is normally only possible to collect one diffraction pattern from each crystal because of beam induced damage. Researchers have developed methods to merge the diffraction patterns produced by hundreds of small crystals, but to date these techniques have only worked with very thin two-dimensional crystals that contain only one layer of the protein of interest. Now Shi et al. report a new approach to Electron Crystallography that works with very small three-dimensional crystals. Called MicroED, this technique involves placing the crystal in a transmission Electron cryo-microscope, which is a fairly standard piece of equipment in many laboratories. The normal ‘low-dose’ Electron beam in one of these microscopes would normally damage the crystal after a single diffraction pattern had been collected. However, Shi et al. realized that it was possible to obtain diffraction patterns without severely damaging the crystal if they dramatically reduced the normal low-dose Electron beam. By reducing the Electron dose by a factor of 200, it was possible to collect up to 90 diffraction patterns from the same, very small, three-dimensional crystal, and then—similar to what happens in X-ray Crystallography—work backwards to figure out the structure of the protein. Shi et al. demonstrated the feasibility of the MicroED approach by using it to determine the structure of lysozyme, which is widely used as a test protein in Crystallography, with a resolution of 2.9 A. This proof-of principle study paves the way for crystallographers to study protein that cannot be studied with existing techniques.
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High-throughput methods for Electron Crystallography.
Methods of Molecular Biology, 2012Co-Authors: David L. Stokes, Iban Ubarretxena-belandia, Tamir Gonen, Andreas EngelAbstract:Membrane proteins play a tremendously important role in cell physiology and serve as a target for an increasing number of drugs. Structural information is key to understanding their function and for developing new strategies for combating disease. However, the complex physical chemistry associated with membrane proteins has made them more difficult to study than their soluble cousins. Electron Crystallography has historically been a successful method for solving membrane protein structures and has the advantage of providing the natural environment of a lipid membrane. Specifically, when membrane proteins form two-dimensional arrays within a lipid bilayer, images and diffraction can be recorded by Electron microscopy. The corresponding data can be combined to produce a three-dimensional reconstruction which, under favorable conditions, can extend to atomic resolution. Like X-ray Crystallography, the quality of the structures are very much dependent on the order and size of the crystals. However, unlike X-ray Crystallography, high-throughput methods for screening crystallization trials for Electron Crystallography are not in general use. In this chapter, we describe two alternative and potentially complementary methods for high-throughput screening of membrane protein crystallization within the lipid bilayer. The first method relies on the conventional use of dialysis for removing detergent and thus reconstituting the bilayer; an array of dialysis wells in the standard 96-well format allows the use of a liquid-handling robot and greatly increases throughput. The second method relies on detergent complexation by cyclodextrin; a specialized pipetting robot has been designed not only to titrate cyclodextrin, but to use light scattering to monitor the reconstitution process. In addition, the use of liquid-handling robots for making negatively stained grids and methods for automatically imaging samples in the Electron microscope are described.
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Advances in Structural and Functional Analysis of Membrane Proteins by Electron Crystallography
Structure, 2011Co-Authors: Goragot Wisedchaisri, Steve L. Reichow, Tamir GonenAbstract:Electron Crystallography is a powerful technique for the study of membrane protein structure and function in the lipid environment. When well-ordered two-dimensional crystals are obtained the structure of both protein and lipid can be determined and lipid-protein interactions analyzed. Protons and ionic charges can be visualized by Electron Crystallography and the protein of interest can be captured for structural analysis in a variety of physiologically distinct states. This review highlights the strengths of Electron Crystallography and the momentum that is building up in automation and the development of high throughput tools and methods for structural and functional analysis of membrane proteins by Electron Crystallography.
Werner Kuhlbrandt - One of the best experts on this subject based on the ideXlab platform.
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introduction to Electron Crystallography
Methods of Molecular Biology, 2013Co-Authors: Werner KuhlbrandtAbstract:: From the earliest work on regular arrays in negative stain, Electron Crystallography has contributed greatly to our understanding of the structure and function of biological macromolecules. The development of Electron cryo-microscopy (cryo-EM) then lead to the first groundbreaking atomic models of the membrane proteins bacteriorhodopsin and light harvesting complex II within lipid bilayers. Key contributions towards cryo-EM and Electron Crystallography methods included specimen preparation and vitrification, liquid-helium cooling, data collection, and image processing. These methods are now applied almost routinely to both membrane and soluble proteins. Here we outline the advances and the breakthroughs that paved the way towards high-resolution structures by Electron Crystallography, both in terms of methods development and biological milestones.
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High-Resolution Electron Crystallography of Membrane Proteins
Membrane Protein Structure, 1994Co-Authors: Werner KuhlbrandtAbstract:For the foreseeable future, progress in determining high-resolution structures of membrane proteins will depend on crystallographic techniques. Until recently, x-ray Crystallography seemed to be the only promising method. Progress with this technique, however, has not been as rapid as originally hoped because it is difficult to grow large and sufficiently well-ordered three-dimensional crystals. Electron Crystallography of two-dimensional crystals is now a viable alternative that is particularly suitable for structural studies of membrane proteins because of their natural propensity to form two-dimensional arrays.
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atomic model of plant light harvesting complex by Electron Crystallography
Nature, 1994Co-Authors: Werner Kuhlbrandt, Daneng Wang, Yoshinori FujiyoshiAbstract:The structure of the light-harvesting chlorophyll a/b–protein complex, an integral membrane protein, has been determined at 3.4 A resolution by Electron Crystallography of two-dimensional crystals. Two of the three membrane-spanning α-helices are held together by ion pairs formed by charged residues that also serve as chlorophyll ligands. In the centre of the complex, chlorophyll a is in close contact with chlorophyll b for rapid energy transfer, and with two carotenoids that prevent the formation of toxic singlet oxygen.
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three dimensional structure of plant light harvesting complex determined by Electron Crystallography
Nature, 1991Co-Authors: Werner Kuhlbrandt, Daneng WangAbstract:The structure of the light-harvesting chlorophyll a/b-protein complex, a membrane protein serving as the major antenna of solar energy in plant photosynthesis, has been determined at 6 A resolution by Electron Crystallography. Within the complex, three membrane-spanning α helices and 15 chlorophyll molecules are resolved. There is an intramolecular diad relating two of the α helices and some of the chlorophylls. The spacing of the chlorophylls suggests energy transfer by delocalized exciton coupling and Forster mechanisms.
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Structure of the Light-Harvesting Chlorophyll a/b Protein Complex by High-Resolution Electron Crystallography
Current Research in Photosynthesis, 1990Co-Authors: Werner KuhlbrandtAbstract:High-resolution Electron Crystallography (1,2) is emerging as an alternative to X-ray Crystallography for determining the structure of biological macromolecules at near-atomic resolution. The technique combines Electron diffraction, high-resolution Electron microscopy and image processing. Crystals for Electron Crystallography need to be very thin. Crystalline monolayers, often referred to as two-dimensional crystals, are ideal. With membrane proteins, order in two dimensions may be easier to achieve than in three dimensions. Indeed, many membrane proteins tend to form two-dimensional crystals. A well-known example is bacteriorhodopsin which occurs naturally in highly ordered two-dimensional arrays. Two-dimensional crystals can also be grown from purified detergent-solubilized membrane proteins, as is the case with the light-harvesting chlorophyll a/b-protein complex (LHC-II) (3) and bacterial porins (4,5).
Yoshinori Fujiyoshi - One of the best experts on this subject based on the ideXlab platform.
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Water channel structures analysed by Electron Crystallography.
Biochimica et Biophysica Acta, 2013Co-Authors: Kazutoshi Tani, Yoshinori FujiyoshiAbstract:Abstract Background The mechanisms underlying water transport through aquaporin (AQP) have been debated for two decades. The water permeation phenomenon of AQP seems inexplicable because the Grotthuss mechanism does not allow for simultaneous fast water permeability and inhibition of proton transfer through the hydrogen bonds of water molecules. Scope of review The AQP1 structure determined by Electron Crystallography provided the first insights into the proton exclusion mechanism despite fast water permeation. Although several studies have provided clues about the mechanism based on the AQP structure, each proposed mechanism remains incomplete. The present review is focused on AQP function and structure solved by Electron Crystallography in an attempt to fill the gaps between the findings in the absence and presence of lipids. Major conclusions Many AQP structures can be superimposed regardless of the determination method. The AQP fold is preserved even under conditions lacking lipids, but the water arrangement in the channel pore differs. The differences might be explained by dipole moments formed by the two short helices in the lipid bilayer. In addition, structure analyses of double-layered two-dimensional crystals of AQP suggest an array formation and cell adhesive function. General significance Electron Crystallography findings not only have contributed to resolve some of the water permeation mechanisms, but have also elucidated the multiple functions of AQPs in the membrane. The roles of AQPs in the brain remain obscure, but their multiple activities might be important in the regulation of brain and other biological functions. This article is part of a Special Issue entitled Aquaporins.
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Future Directions of Electron Crystallography
Methods of Molecular Biology, 2012Co-Authors: Yoshinori FujiyoshiAbstract:: In biological science, there are still many interesting and fundamental yet difficult questions, such as those in neuroscience, remaining to be answered. Structural and functional studies of membrane proteins, which are key molecules of signal transduction in neural and other cells, are essential for understanding the molecular mechanisms of many fundamental biological processes. Technological and instrumental advancements of Electron microscopy have facilitated comprehension of structural studies of biological components, such as membrane proteins. While X-ray Crystallography has been the main method of structure analysis of proteins including membrane proteins, Electron Crystallography is now an established technique to analyze structures of membrane proteins in the lipid bilayer, which is close to their natural biological environment. By utilizing cryo-Electron microscopes with helium-cooled specimen stages, structures of membrane proteins were analyzed at a resolution better than 3 A. Such high-resolution structural analysis of membrane proteins by Electron Crystallography opens up the new research field of structural physiology. Considering the fact that the structures of integral membrane proteins in their native membrane environment without artifacts from crystal contacts are critical in understanding their physiological functions, Electron Crystallography will continue to be an important technology for structural analysis. In this chapter, I will present several examples to highlight important advantages and to suggest future directions of this technique.
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Electron Crystallography for structural and functional studies of membrane proteins.
Journal of Electron Microscopy, 2011Co-Authors: Yoshinori FujiyoshiAbstract:: Membrane proteins are important research targets for basic biological sciences and drug design, but studies of their structure and function are considered difficult to perform. Studies of membrane structures have been greatly facilitated by technological and instrumental advancements in Electron microscopy together with methodological advancements in biology. Electron Crystallography is especially useful in studying the structure and function of membrane proteins. Electron Crystallography is now an established method of analyzing the structures of membrane proteins in lipid bilayers, which resembles their natural biological environment. To better understand the neural system function from a structural point of view, we developed the cryo-Electron microscope with a helium-cooled specimen stage, which allows for analysis of the structures of membrane proteins at a resolution higher than 3 A. This review introduces recent instrumental advances in cryo-Electron microscopy and presents some examples of structure analyses of membrane proteins, such as bacteriorhodopsin, water channels and gap junction channels. This review has two objectives: first, to provide a personal historical background to describe how we came to develop the cryo-Electron microscope and second, to discuss some of the technology required for the structural analysis of membrane proteins based on cryo-Electron microscopy.
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Structural physiology based on Electron Crystallography.
Protein Science, 2011Co-Authors: Yoshinori FujiyoshiAbstract:There are many questions in brain science, which are extremely interesting but very difficult to answer. For example, how do education and other experiences during human development influence the ability and personality of the adult? The molecular mechanisms underlying such phenomena are still totally unclear. However, technological and instrumental advancements of Electron microscopy have facilitated comprehension of the structures of biological components, cells, and organelles. Electron Crystallography is especially good for studying the structure and function of membrane proteins, which are key molecules of signal transduction in neural and other cells. Electron Crystallography is now an established technique to analyze the structures of membrane proteins in lipid bilayers, which are close to their natural biological environment. By utilizing cryo-Electron microscopes with helium cooled specimen stages, which were developed through a personal motivation to understand functions of neural systems from a structural point of view, structures of membrane proteins were analyzed at a resolution higher than 3 A. This review has four objectives. First, it is intended to introduce the new research field of structural physiology. Second, it introduces some of the personal struggles, which were involved in developing the cryo-Electron microscope. Third, it discusses some of the technology for the structural analysis of membrane proteins based on cryo-Electron microscopy. Finally, it reviews structural and functional analyses of membrane proteins.
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Electron Crystallography and aquaporins
Methods in Enzymology, 2010Co-Authors: Andreas D Schenk, Yoshinori Fujiyoshi, Richard K Hite, Andreas Engel, Thomas WalzAbstract:Abstract Electron Crystallography of two-dimensional (2D) crystals can provide information on the structure of membrane proteins at near-atomic resolution. Originally developed and used to determine the structure of bacteriorhodopsin (bR), Electron Crystallography has recently been applied to elucidate the structure of aquaporins (AQPs), a family of membrane proteins that form pores mostly for water but also other solutes. While Electron Crystallography has made major contributions to our understanding of the structure and function of AQPs, structural studies on AQPs, in turn, have fostered a number of technical developments in Electron Crystallography. In this contribution, we summarize the insights Electron Crystallography has provided into the biology of AQPs, and describe technical advancements in Electron Crystallography that were driven by structural studies on AQP 2D crystals. In addition, we discuss some of the lessons that were learned from Electron crystallographic work on AQPs.
Thomas Walz - One of the best experts on this subject based on the ideXlab platform.
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Electron Crystallography and aquaporins
Methods in Enzymology, 2010Co-Authors: Andreas D Schenk, Yoshinori Fujiyoshi, Richard K Hite, Andreas Engel, Thomas WalzAbstract:Abstract Electron Crystallography of two-dimensional (2D) crystals can provide information on the structure of membrane proteins at near-atomic resolution. Originally developed and used to determine the structure of bacteriorhodopsin (bR), Electron Crystallography has recently been applied to elucidate the structure of aquaporins (AQPs), a family of membrane proteins that form pores mostly for water but also other solutes. While Electron Crystallography has made major contributions to our understanding of the structure and function of AQPs, structural studies on AQPs, in turn, have fostered a number of technical developments in Electron Crystallography. In this contribution, we summarize the insights Electron Crystallography has provided into the biology of AQPs, and describe technical advancements in Electron Crystallography that were driven by structural studies on AQP 2D crystals. In addition, we discuss some of the lessons that were learned from Electron crystallographic work on AQPs.
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Electron Crystallography as a technique to study the structure on membrane proteins in a lipidic environment
Annual Review of Biophysics, 2009Co-Authors: Stefan Raunser, Thomas WalzAbstract:The native environment of integral membrane proteins is a lipid bilayer. The structure of a membrane protein is thus ideally studied in a lipidic environment. In the first part of this review we describe some membrane protein structures that revealed the surrounding lipids and provide a brief overview of the techniques that can be used to study membrane proteins in a lipidic environment. In the second part of this review we focus on Electron Crystallography of two-dimensional crystals as potentially the most suitable technique for such studies. We describe the individual steps involved in the Electron crystallographic determination of a membrane protein structure and discuss current challenges that need to be overcome to transform Electron Crystallography into a technique that can be routinely used to analyze the structure of membrane proteins embedded in a lipid bilayer.
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revival of Electron Crystallography
Current Opinion in Structural Biology, 2007Co-Authors: Richard K Hite, Stefan Raunser, Thomas WalzAbstract:Since the structure determination of bacteriorhodopsin in 1990, much progress has been made in the further development and use of Electron Crystallography. In this review, we provide a concise overview of the new developments in Electron Crystallography concerning 2D crystallization, data collection and data processing. Based on Electron crystallographic work on bacteriorhodopsin, the acetylcholine receptor and aquaporins, we highlight the unique advantages and future perspectives of Electron Crystallography for the structural study of membrane proteins. These advantages include the visualization of membrane proteins in their native environment without detergent-induced artifacts, the trapping of different states in a reaction pathway by time-resolved experiments, the study of non-specific protein-lipid interactions and the characterization of the charge state of individual residues in membrane proteins.
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Electron Crystallography of Two-Dimensional Crystals of Membrane Proteins☆
Journal of Structural Biology, 1998Co-Authors: Thomas Walz, Nikolaus GrigorieffAbstract:Abstract Electron microscopy has become a powerful technique, along with X-ray Crystallography and nuclear magnetic resonance spectroscopy, to study the three-dimensional structure of biological molecules. It has evolved into a number of methods dealing with a wide range of biological samples, with Electron Crystallography of two-dimensional crystals being so far the only method allowing data collection at near-atomic resolution. In this paper, we review the methodology of Electron Crystallography and its application to membrane proteins, starting with the pioneering work on bacteriorhodopsin, which led to the first visualization of the secondary structure of a membrane protein in 1975. Since then, improvements in instrumentation, sample preparation, and data analysis have led to atomic models for bacteriorhodopsin and light-harvesting complex II from higher plants. The structures of many more membrane proteins have been studied by Electron Crystallography and in this review examples are included where a resolution of better than 10 A has been achieved. Indeed, in some of the given examples an atomic model can be expected in the near future. Finally, a brief outlook is given on current and future developments of Electron crystallographic methods.
