Electron microscopy projections

In this work the Rb. sphaeroides LHl was modeled as a circular aggregate of 16 ap-heterodimers, i.e., a hexadecamer, as shown in the side and top views, respectively, in Color plates 5 (A) and (B). The 16 aP-heterodimers form two concentric cylinders, with the a-polypeptides on the inside and the P-polypep-tides on the outside. The diameters ofthe inner and outer cylinders are 78 and 108 A, respectively. Color plate 5 (C) shows the calculated projection map of Rb. sphaeroides LHl superimposed onto the 8.5-A electron-microscopy projection map of Rs. rubrum LHl measured previously by Karrasch et al The contours of the two projection maps appear to be a good match, indicating that the polypeptides in the calculated model are probably positioned correctly. 

Thus, the interaction of the primary beam with the sample provides a wealth of information on morphology, crystallography and chemical composition. Using transmission electron microscopy to make a projection of the sample density is a routine way to study particle sizes in catalysts. 

The dimer chains of Ca -ATPase can also be observed by freeze-fracture electron microscopy [119,165,166,172-174], forming regular arrays of oblique parallel ridges on the concave P fracture faces of the membrane, with complementary grooves or furrows on the convex E fracture faces. Resolution of the surface projections of individual Ca -ATPase molecules within the crystalline arrays has also been achieved on freeze-dried rotary shadowed preparations of vanadate treated rabbit sarcoplasmic reticulum [163,166,173,175]. The unit cell dimensions derived from these preparations are a = 6.5 nm b = 10.7 nm and 7 = 85.5° [175], in reasonable agreement with earlier estimates on negatively stained preparations [88]. 

Two distinct patterns of repeats were observed by electron microscopy of sectioned, negatively stained, frozen-hyd rated, or freeze-fractured specimens of Ca -ATPase crystals that represent different projections of the same structure... 

An unusually extensive battery of experimental techniques was brought to bear on these comparisons of enantiomers with their racemic mixtures and of diastereomers with each other. A very sensitive Langmuir trough was constructed for the project, with temperature control from 15 to 40°C. In addition to the familiar force/area isotherms, which were used to compare all systems, measurements of surface potentials, surface shear viscosities, and dynamic suface tensions (for hysteresis only) were made on several systems with specially designed apparatus. Several microscopic techniques, epi-fluorescence optical microscopy, scanning tunneling microscopy, and electron microscopy, were applied to films of stearoylserine methyl ester, the most extensively investigated surfactant. 

Lee, J.Y., Urbatsch, I.L., Senior, A.E. and Wilkens, S. (2002) Projection structure of P-glycoprotein by electron microscopy. Evidence for a closed conformation of the nucleotide binding domains. Journal of Biological Chemistry, 277, 40125-40131. 

Dark-field electron microscopy (in which the image is formed from the scattered beam), when combined with improved techniques of sample handling and preparation and minimal radiation exposure, can lead to images of sufficiently undamaged DNA at a resolution of 10 A (116). Figure 45 shows such an image in which the two-dimensional projection of the helix is clearly visible on the undamaged part of the molecule. 

We have reconstructed the 3D structure of a complex quasicrystal approximant v-AlCrFe (P6 m, a = 40.687 and c = 12.546 A) (Zou et al, 2004). Due to the huge unit cell, it was necessary to combine crystallographic data from 13 projections to resolve the atoms. Electron microscopy images containing both amplitude and phase information were combined with amplitudes from electron diffraction patterns. 124 of the 129 unique atoms (1176 in the unit cell) were found in the remarkably clean calculated potential maps. This investigation demonstrates that inorganic crystals of any complexity can be solved by electron crystallography. 

In chemistry, we are often interested in the bulk of the material, and for this purpose we must view the structures with a probe that penetrates through the object. Transmission electron microscopy is ideal for this. The 3D structure of a transparent object is much more complex than the surface, which can be considered as a 2D object, although it often is not at all flat. In the case of a crystal, the object may be hundreds of atoms thick, resulting in a massive overlap of atoms in any direction we chose to look at the crystal from. It is easily realized that even three orthogonal views are not sufficient for resolving all overlapping reflections, unless the structure is very simple. The larger the unit cell is, the more projections are needed in order to obtain a structure with all atoms resolved. 

D reconstruction can be performed by restoring the 3D Fourier space of the object from a series of 2D Fourier transforms of the projections. Then the 3D object can be reconstructed by inverse Fourier transformation of the 3D Fourier space. For crystalline objects, the Fourier transforms are discrete spots, i.e. reflections. In electron microscopy, the Fourier transform of the projection of the 3D electrostatic potential distribution inside a crystal, or crystal structure factors, can be obtained from HREM images of thin crystals. So one can obtain the 3D electrostatic potential distribution (p(r) inside a crystal from a series of projections by... 

Voigt-Martin et al. [13] have used MICE to solve the stmcture of 4-(4 -(N,N-dimethyl)aminobenzylidene)-pyrazolidine-3,5-dione at 1.4A in projection using 42 reflections. The potential maps do not resolve atoms with these data and models have to be fitted to the map density in a way reminiscent of macromolecular crystallography. This can pose problems in structure validation which were overcome in this case by simulation calculations. There is an excellent agreement between the solution and independent model building and high resolution electron microscopy studies. 

Transmission electron microscopy (TEM) can provide valuable information on particle size, shape, and structure, as well as on the presence of different types of colloidal structures within the dispersion. As a complication, however, all electron microscopic techniques applicable for solid lipid nanoparticles require more or less sophisticated specimen preparation procedures that may lead to artifacts. Considerable experience is often necessary to distinguish these artifacts from real structures and to decide whether the structures observed are representative of the sample. Moreover, most TEM techniques can give only a two-dimensional projection of the three-dimensional objects under investigation. Because it may be difficult to conclude the shape of the original object from electron micrographs, additional information derived from complementary characterization methods is often very helpful for the interpretation of electron microscopic data. 

Electron microscopy of sectioned chloroplasts shows ATP synthase complexes as knoblike projections on the outside (stromal or N) surface of thylalcoid membranes these complexes correspond to the ATP synthase complexes seen to project on the inside (matrix or N) surface of the inner mitochondrial membrane. Thus the relationship between the orientation of the ATP synthase and the direction of proton pumping is the same in chloroplasts and mitochondria. In both cases, the Fl portion of ATP synthase is located on the more alkaline (N) side of the membrane through which protons flow down their concentration gradient the direction of proton flow relative to Fi is the same in both cases P to N (Fig. 19-58). 

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