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Rapid structural analysis of bacterial ribosomes in situ.


ABSTRACT: 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 the development of 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 we expect this workflow will be widely applicable to related bacterial samples.

SUBMITTER: Powell BM 

PROVIDER: S-EPMC10996489 | biostudies-literature | 2024 Mar

REPOSITORIES: biostudies-literature

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Rapid structural analysis of bacterial ribosomes <i>in situ</i>.

Powell Barrett M BM   Brant Tyler S TS   Davis Joseph H JH   Mosalaganti Shyamal S  

bioRxiv : the preprint server for biology 20240326


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 the developm  ...[more]

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