Electron crystallography of ultrathin 3D protein crystals: Atomic model with charges

Significance Electron crystallography has the potential to analyze crystals of membrane proteins and macromolecular complexes too small or too thin for X-ray crystallography, as electrons are scattered four to five orders of magnitude more strongly than X-rays. Electron crystallography yields Coulomb potential maps, rather than electron density maps as X-rays do, providing information on charged states of amino acids and metals. Here we present such Coulomb potential maps at 3.4-Å and 3.2-Å resolution, respectively, of Ca2+-ATPase and catalase obtained from crystals of just a few layers thick. These maps demonstrate that it is indeed possible to build atomic models from such crystals and charge information is included, often critical in understanding protein function. 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 Ca2+-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 Ca2+-binding sites of Ca2+-ATPase and that of the iron atom in the heme in catalase.