Membrane proteins and macromolecular complexes often yield crystals too small or too thin for even the modern synchrotron X-ray beam. Electron crystallography could provide a powerful means for structure determination with such undersized crystals, as protein atoms diffract electrons four to five orders of magnitude more strongly than they do X-rays. Furthermore, as electron crystallography yields Coulomb potential maps rather than electron density maps, it could provide a unique method to visualize the charged states of amino acid residues and metals. Here we describe an attempt to develop a methodology for electron crystallography of ultrathin (only a few layers thick) 3D protein crystals and present the Coulomb potential maps at 3.4-Å and 3.2-Å resolution, respectively, obtained from Ca 2+ -ATPase and catalase crystals. These maps demonstrate that it is indeed possible to build atomic models from such crystals and even to determine the charged states of amino acid residues in the Ca 2+ -binding sites of Ca 2+ -ATPase and that of the iron atom in the heme in catalase.electron crystallography | protein crystal | Coulomb potential | Ca 2+ -ATPase | catalase P rotein atoms scatter electrons four to five orders of magnitude more strongly than they do X-rays, thus allowing individual protein molecules to be imaged by electron microscopy (1). Although not fully exploited so far, electron protein crystallography has great potential and indeed has yielded superb high-resolution (∼2.0-Å resolution) atomic structures from 2D crystals (2). However, electron crystallography of 3D crystals is problematic, as stacking of even a few layers makes diffraction patterns discrete in all directions, and methods developed for conventional electron crystallography of 2D crystals (3) are not useful (SI Appendix, Fig. S1A, Left). This problem can be overcome, however, as Gonen and coworkers demonstrated (4, 5), by rotating the crystal to spatially integrate the intensities of diffraction spots as in X-ray crystallography (SI Appendix, Fig. S1A, Right) or, in certain cases, even combining simple tilt series.Another important feature of electron scattering is that the diffraction pattern formed by elastically scattered electrons is directly related to the distribution of Coulomb potential. This is in marked contrast to X-rays, which, because they are scattered by electrons, yield an electron density map. Coulomb potential maps may be more difficult to interpret, compared with electron density maps by X-ray crystallography, as the appearance of the same residues may differ depending on their charged state, resolution, and surrounding environment ( Fig. 1 and SI Appendix, Fig. S2), but they provide unique information, not attainable by X-rays (6). Theoretical potential maps (F calc maps; Fig. 1 B-E) calculated from an atomic model of Ca 2+ -ATPase (7) using scattering factors for 300-keV electrons highlight these features. For instance, densities of acidic residues are absent or weak when lower-resolution data are included in the map calc...

The possibility of photo-transmutation of long-lived nuclide Cs135 by ultrashort ultraintense laser was analytically evaluated. The yield of Cs135(γ,n) Cs134 was strongly dependent on the laser intensity at around 1020W∕cm2. If Cs135 were irradiated by such a laser with the intensity of 1021W∕cm2 and 10 Hz for 30 min, characteristic γ-ray counting rate was estimated to be 3 Bq.

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