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

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 112; no. 11; pp. 3368 - 3373
• Main Authors: Yonekura, Koji, 米倉功治, Kato, Kazuyuki, 加藤一幸, Ogasawara, Mitsuo, 小笠原光雄, Tomita, Masahiro, 富田正弘, Toyoshima, Chikashi, 豊島近
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences 17.03.2015; National Acad Sciences
• Subjects: Animals; Binding sites; Biological Sciences; Calcium-Transporting ATPases - chemistry; Catalase - chemistry; Cattle; Crystallography; Crystallography, X-Ray - methods; Crystals; Electrons; Models, Molecular; Proteins; Static Electricity; Coulomb potential; protein crystal; Ca2+-ATPase; catalase; electron crystallography
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1500724112

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-ÅÅ resolution, respectively, of Ca ²⁺-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-ÅÅÅ and 3.2-ÅÅÅÅ 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.