The Experts below are selected from a list of 113232 Experts worldwide ranked by ideXlab platform
Tamir Gonen - One of the best experts on this subject based on the ideXlab platform.
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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.
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, Richard K Hite, Yoshinori Fujiyoshi, 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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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.
Robert Bucker - One of the best experts on this subject based on the ideXlab platform.
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serial electron diffraction data processing with diffractem and crystfel
Frontiers in Molecular Biosciences, 2021Co-Authors: Robert Bucker, Pascal Hoganlamarre, R Dwayne J MillerAbstract:Serial electron diffraction (SerialED) is an emerging technique, which applies the snapshot data-collection mode of serial X-ray Crystallography to three-dimensional electron diffraction (3D Electron Diffraction), forgoing the conventional rotation method. Similarly to serial X-ray Crystallography, this approach leads to almost complete absence of radiation damage effects even for the most sensitive samples, and allows for a high level of automation. However, SerialED also necessitates new techniques of data processing, which combine existing pipelines for rotation electron diffraction and serial X-ray Crystallography with some more particular solutions for challenges arising in SerialED specifically. Here, we introduce our analysis pipeline for SerialED data, and its implementation using the CrystFEL and diffractem program packages. Detailed examples are provided in extensive supplementary code.
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serial electron diffraction data processing with diffractem and crystfel
arXiv: Data Analysis Statistics and Probability, 2020Co-Authors: Robert Bucker, Pascal Hoganlamarre, R Dwayne J MillerAbstract:Serial electron diffraction (SerialED) is an emerging technique, which applies the snapshot data-collection mode of serial X-ray Crystallography to three-dimensional electron diffraction (3D ED), forgoing the conventional rotation method. Similarly to serial X-ray Crystallography, this approach leads to almost complete absence of radiation damage effects even for the most sensitive samples, and allows for a high level of automation. However, SerialED also necessitates new techniques of data processing, which combine existing pipelines for rotation electron diffraction and serial X-ray Crystallography with some more particular solutions for challenges arising in SerialED specifically. Here, we introduce our analysis pipeline for SerialED data, and its implementation using the CrystFEL and diffractem program packages. Detailed examples are provided in extensive supplementary code.
Samuel H. Gellman - One of the best experts on this subject based on the ideXlab platform.
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Retention of Native Quaternary Structure in Racemic Melittin Crystals.
Journal of the American Chemical Society, 2019Co-Authors: Kathleen W. Kurgan, Adam F. Kleman, Craig A. Bingman, Dale F. Kreitler, Bernard Weisblum, Katrina T. Forest, Samuel H. GellmanAbstract:Racemic Crystallography has been used to elucidate the secondary and tertiary structures of peptides and small proteins that are recalcitrant to conventional crystallization. It is unclear, however, whether racemic Crystallography can capture native quaternary structure, which could be disrupted by heterochiral associations. We are exploring the use of racemic Crystallography to characterize the self-assembly behavior of membrane-associated peptides, very few of which have been crystallized. We report a racemic crystal structure of the membrane-active peptide melittin; the new structure allows comparison with a previously reported crystal structure of L-melittin. The tetrameric assembly observed in crystalline L-melittin has been proposed to represent the tetrameric state detected in solution for this peptide. This tetrameric assembly is precisely reproduced in the racemic crystal, which strengthens the conclusion that the tetramer is biologically relevant. More broadly, these findings suggest that racemic Crystallography can provide insight on native quaternary structure.
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Retention of Native Quaternary Structure in Racemic Melittin Crystals
2019Co-Authors: Kathleen W. Kurgan, Adam F. Kleman, Craig A. Bingman, Dale F. Kreitler, Bernard Weisblum, Katrina T. Forest, Samuel H. GellmanAbstract:Racemic Crystallography has been used to elucidate the secondary and tertiary structures of peptides and small proteins that are recalcitrant to conventional crystallization. It is unclear, however, whether racemic Crystallography can capture native quaternary structure, which could be disrupted by heterochiral associations. We are exploring the use of racemic Crystallography to characterize the self-assembly behavior of membrane-associated peptides, very few of which have been crystallized. We report a racemic crystal structure of the membrane-active peptide melittin; the new structure allows comparison with a previously reported crystal structure of L-melittin. The tetrameric assembly observed in crystalline L-melittin has been proposed to represent the tetrameric state detected in solution for this peptide. This tetrameric assembly is precisely reproduced in the racemic crystal, which strengthens the conclusion that the tetramer is biologically relevant. More broadly, these findings suggest that racemic Crystallography can provide insight on native quaternary structure
R Dwayne J Miller - One of the best experts on this subject based on the ideXlab platform.
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serial electron diffraction data processing with diffractem and crystfel
Frontiers in Molecular Biosciences, 2021Co-Authors: Robert Bucker, Pascal Hoganlamarre, R Dwayne J MillerAbstract:Serial electron diffraction (SerialED) is an emerging technique, which applies the snapshot data-collection mode of serial X-ray Crystallography to three-dimensional electron diffraction (3D Electron Diffraction), forgoing the conventional rotation method. Similarly to serial X-ray Crystallography, this approach leads to almost complete absence of radiation damage effects even for the most sensitive samples, and allows for a high level of automation. However, SerialED also necessitates new techniques of data processing, which combine existing pipelines for rotation electron diffraction and serial X-ray Crystallography with some more particular solutions for challenges arising in SerialED specifically. Here, we introduce our analysis pipeline for SerialED data, and its implementation using the CrystFEL and diffractem program packages. Detailed examples are provided in extensive supplementary code.
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serial electron diffraction data processing with diffractem and crystfel
arXiv: Data Analysis Statistics and Probability, 2020Co-Authors: Robert Bucker, Pascal Hoganlamarre, R Dwayne J MillerAbstract:Serial electron diffraction (SerialED) is an emerging technique, which applies the snapshot data-collection mode of serial X-ray Crystallography to three-dimensional electron diffraction (3D ED), forgoing the conventional rotation method. Similarly to serial X-ray Crystallography, this approach leads to almost complete absence of radiation damage effects even for the most sensitive samples, and allows for a high level of automation. However, SerialED also necessitates new techniques of data processing, which combine existing pipelines for rotation electron diffraction and serial X-ray Crystallography with some more particular solutions for challenges arising in SerialED specifically. Here, we introduce our analysis pipeline for SerialED data, and its implementation using the CrystFEL and diffractem program packages. Detailed examples are provided in extensive supplementary code.