Zernike phase contrast cryo-electron microscopy and tomography for structure determination at nanometer and subnanometer resolutions.
Zernike phase contrast cryo-electron microscopy (ZPC-cryoEM) is an emerging technique that is capable of producing higher image contrast than conventional cryoEM. By combining this technique with advanced image processing methods, we achieved subnanometer resolution for two biological specimens: 2D bacteriorhodopsin crystal and epsilon15 bacteriophage. For an asymmetric reconstruction of epsilon15 bacteriophage, ZPC-cryoEM can reduce the required amount of data by a factor of approximately 3, compared with conventional cryoEM. The reconstruction was carried out to 13 A resolution without the need to correct the contrast transfer function. New structural features at the portal vertex of the epsilon15 bacteriophage are revealed in this reconstruction. Using ZPC cryo-electron tomography (ZPC-cryoET), a similar level of data reduction and higher resolution structures of epsilon15 bacteriophage can be obtained relative to conventional cryoET. These results show quantitatively the benefits of ZPC-cryoEM and ZPC-cryoET for structural determinations of macromolecular machines at nanometer and subnanometer resolutions.
Murata
Kazuyoshi
K
National Center for Macromolecular Imaging, Verna and Marrs Mclean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, TX 77030, USA.
Liu
Xiangan
X
Danev
Radostin
R
Jakana
Joanita
J
Schmid
Michael F
MF
King
Jonathan
J
Nagayama
Kuniaki
K
Chiu
Wah
W
D001435:Bacteriophages / Q000648:ultrastructure; D001436:Bacteriorhodopsins / Q000648:ultrastructure; D020285:Cryoelectron Microscopy / Q000379:methods*; D003461:Crystallography / Q000379:methods*; D055032:Electron Microscope Tomography / Q000379:methods*; D020565:Epsilonproteobacteria / Q000821:virology; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D008858:Microscopy, Phase-Contrast / Q000379:methods*; D008958:Models, Molecular
D001436:Bacteriorhodopsins