2dx—User-friendly image processing for 2D crystals

Gipson, Bryant; Zeng, Xiangyan; Zhang, Zi Yan; Stahlberg, Henning

doi:10.1016/j.jsb.2006.07.020

Cited by 184 publications

(166 citation statements)

References 40 publications

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“…Image processing (28) led to a 2D electron density map that showed clear density consistent with paired α-helices in the coiled-coil fibrils (Fig. 3 B and C).…”

Section: Resultsmentioning

confidence: 98%

“…The model hexagonal array of coiled coils was used to simulate an electron density map filtered to 10 Å. The electron density of the model was projected over angular steps of 5°(Movie S1) and the resulting projection images compared and aligned to the processed images output from 2dx (28). Matching projection images and processed micrographs (shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Micrographs of fibers displaying striations were treated as 2D crystals and analyzed using 2dx (28). Layer and row lines were identified by eye and followed by automatic spot selection and unbending.…”

Section: Methodsmentioning

confidence: 99%

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Cryo-transmission electron microscopy structure of a gigadalton peptide fiber of de novo design

Sharp

Bruning

Mantell

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Nature presents various protein fibers that bridge the nanometer to micrometer regimes. These structures provide inspiration for the de novo design of biomimetic assemblies, both to address difficulties in studying and understanding natural systems, and to provide routes to new biomaterials with potential applications in nanotechnology and medicine. We have designed a self-assembling fiber system, the SAFs, in which two small α-helical peptides are programmed to form a dimeric coiled coil and assemble in a controlled manner. The resulting fibers are tens of nm wide and tens of μm long, and, therefore, comprise millions of peptides to give gigadalton supramolecular structures. Here, we describe the structure of the SAFs determined to approximately 8 Å resolution using cryotransmission electron microscopy. Individual micrographs show clear ultrastructure that allowed direct interpretation of the packing of individual α-helices within the fibers, and the construction of a 3D electron density map. Furthermore, a model was derived using the cryotransmission electron microscopy data and side chains taken from a 2.3 Å X-ray crystal structure of a peptide building block incapable of forming fibers. This was validated using singleparticle analysis techniques, and was stable in prolonged molecular-dynamics simulation, confirming its structural viability. The level of self-assembly and self-organization in the SAFs is unprecedented for a designed peptide-based material, particularly for a system of considerably reduced complexity compared with natural proteins. This structural insight is a unique high-resolution description of how α-helical fibrils pack into larger protein fibers, and provides a basis for the design and engineering of future biomaterials.fibrous proteins | protein design | helical reconstruction | X-ray crystallography

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“…Image processing (28) led to a 2D electron density map that showed clear density consistent with paired α-helices in the coiled-coil fibrils (Fig. 3 B and C).…”

Section: Resultsmentioning

confidence: 98%

Section: Resultsmentioning

confidence: 99%

See 1 more Smart Citation

Cryo-transmission electron microscopy structure of a gigadalton peptide fiber of de novo design

Sharp

Bruning

Mantell

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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“…For structure determination of membrane proteins by cryo-TEM, images from non-tilted and tilted 2D crystals are recorded, and the structural information is then extracted from electron micrographs by applying computer image processing. Typical software are the 2dx (Gipson et al, 2007) and the Image Processing Library & Toolbox (IPLT) (Philippsen et al, 2003(Philippsen et al, , 2007 systems, and the Medical Research Council (MRC) programs (Crowther et al, 1996). In contrast to XRC, 3D structure determination at high resolution by cryo-TEM/EC is still a tedious and time-consuming task, although the methodology has made major advances over the past years.…”

Section: Cryo-transmission Electron Microscopymentioning

confidence: 99%

Structure determination of channel and transport proteins by high-resolution microscopy techniques

Meury

Harder²,

Ucurum³

et al. 2011

Biological Chemistry

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High-resolution microscopy techniques provide a plethora of information on biological structures from the cellular level down to the molecular level. In this review, we present the unique capabilities of transmission electron and atomic force microscopy to assess the structure, oligomeric state, function and dynamics of channel and transport proteins in their native environment, the lipid bilayer. Most importantly, membrane proteins can be visualized in the frozen-hydrated state and in buffer solution by cryo-transmission electron and atomic force microscopy, respectively. We also illustrate the potential of the scintillation proximity assay to study substrate binding of detergent-solubilized transporters prior to crystallization and structural characterization.

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“…We also produce a software system 2dx for the user-friendly image processing of electron crystallography data [67]. 2dx provides a graphical user-interface (GUI) that guides the user through the required processing steps, displays processing parameters and processing results in a clearly structured way, and offers extensive help information and functions in all phases of the image processing, see Figure 3.…”

Section: Information Exchange -2dxorgmentioning

confidence: 99%

Milestones in electron crystallography

Renault

Chou

Chiu

et al. 2006

J Comput Aided Mol Des

Self Cite

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SummaryElectron crystallography determines the structure of membrane embedded proteins in the twodimensionally crystallized state by cryo-transmission electron microscopy imaging and computer structure reconstruction. Milestones on the path to the structure are high-level expression, purification of functional protein, reconstitution into two-dimensional lipid membrane crystals, high-resolution imaging, and structure determination by computer image processing. Here we review the current state of these methods. We also created an Internet information exchange platform for electron crystallography, where guidelines for the imaging and data processing method are maintained. The server (http://2dx.org) provides the electron crystallography community with a central information exchange platform, which is structured in blog and Wiki form, allowing visitors to add comments or discussions. It currently offers a detailed step-by-step introduction to image processing with the MRC software program. The server is also a repository for the 2dx software package, a user-friendly image processing system for 2D membrane protein crystals.

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2dx—User-friendly image processing for 2D crystals

Cited by 184 publications

References 40 publications

Cryo-transmission electron microscopy structure of a gigadalton peptide fiber of de novo design

Cryo-transmission electron microscopy structure of a gigadalton peptide fiber of de novo design

Structure determination of channel and transport proteins by high-resolution microscopy techniques

Milestones in electron crystallography

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