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Rapid structural analysis of bacterial ribosomes in situ.

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Název: Rapid structural analysis of bacterial ribosomes in situ.
Autoři: Powell, Barrett M., Brant, Tyler S., Davis, Joseph H., Mosalaganti, Shyamal
Zdroj: Communications Biology; 1/28/2025, Vol. 8 Issue 1, p1-9, 9p
Témata: ESCHERICHIA coli, CELL analysis, PROTEIN analysis, INTEGRATED software, ELECTRONIC data processing
Abstrakt: Rapid structural analysis of purified proteins and their complexes has become increasingly common thanks to key methodological advances in cryo-electron microscopy (cryo-EM) and associated data processing software packages. In contrast, analogous structural analysis in cells via cryo-electron tomography (cryo-ET) remains challenging due to critical technical bottlenecks, including low-throughput sample preparation and imaging, and laborious data processing methods. Here, we describe a rapid in situ cryo-ET sample preparation and data analysis workflow that results in the routine determination of sub-nm resolution ribosomal structures. We apply this workflow to E. coli, producing a 5.8 Å structure of the 70S ribosome from cells in less than 10 days and facilitating the discovery of a minor population of 100S-like disomes. We envision our approach to be widely applicable to related bacterial samples. A workflow for rapid cryo-ET structural characterization of bacterial ribosomes in situ and the identification of a 100S-like disome structure. [ABSTRACT FROM AUTHOR]
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Abstrakt:Rapid structural analysis of purified proteins and their complexes has become increasingly common thanks to key methodological advances in cryo-electron microscopy (cryo-EM) and associated data processing software packages. In contrast, analogous structural analysis in cells via cryo-electron tomography (cryo-ET) remains challenging due to critical technical bottlenecks, including low-throughput sample preparation and imaging, and laborious data processing methods. Here, we describe a rapid in situ cryo-ET sample preparation and data analysis workflow that results in the routine determination of sub-nm resolution ribosomal structures. We apply this workflow to E. coli, producing a 5.8 Å structure of the 70S ribosome from cells in less than 10 days and facilitating the discovery of a minor population of 100S-like disomes. We envision our approach to be widely applicable to related bacterial samples. A workflow for rapid cryo-ET structural characterization of bacterial ribosomes in situ and the identification of a 100S-like disome structure. [ABSTRACT FROM AUTHOR]
ISSN:23993642
DOI:10.1038/s42003-025-07586-y