In situ structure determination at nanometer resolution using TYGRESS

Autor: Kangkang Song, Daniela Nicastro, Xiaochu Lou, Zhiguo Shang, Xiaofeng Fu, Nikolaus Grigorieff
Rok vydání: 2019
Předmět:
Zdroj: Nature methods
ISSN: 1548-7105
1548-7091
DOI: 10.1038/s41592-019-0651-0
Popis: The resolution of subtomogram averages calculated from cryo-electron tomograms (cryo-ET) of crowded cellular environments is often limited owing to signal loss in, and misalignment of, the subtomograms. By contrast, single-particle cryo-electron microscopy (SP-cryo-EM) routinely reaches near-atomic resolution of isolated complexes. We report a method called ‘tomography-guided 3D reconstruction of subcellular structures’ (TYGRESS) that is a hybrid of cryo-ET and SP-cryo-EM, and is able to achieve close-to-nanometer resolution of complexes inside crowded cellular environments. TYGRESS combines the advantages of SP-cryo-EM (images with good signal-to-noise ratio and contrast, as well as minimal radiation damage) and subtomogram averaging (three-dimensional alignment of macromolecules in a complex sample). Using TYGRESS, we determined the structure of the intact ciliary axoneme with up to resolution of 12 A. These results reveal many structural details that were not visible by cryo-ET alone. TYGRESS is generally applicable to cellular complexes that are amenable to subtomogram averaging. The unique advantages of single-particle cryo-electron microscopy and cryo-electron tomography are combined in a method called TYGRESS, here applied to determine the structure of the intact ciliary axoneme at a resolution of 12 A.
Databáze: OpenAIRE
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