Work

Baited reconstruction with 2D template matching for high-resolution structure determination in vitro and in vivo without template bias

Year: 2023

Type: article

Abstract: Previously we showed that 2D template matching (2DTM) can be used to localize macromolecular complexes in images recorded by cryogenic electron microscopy (cryo-EM) with high precision, even in the presence of noise and cellular background (Lucas et al., 2021; Lucas et al., 2022). Here, we show that once localized, these particles may be averaged together to generate high-resolution 3D reconstructions. However, regions included in the template may suffer from template bias, leading to inflated resolution estimates and making the interpretation of high-resolution features unreliable. We evaluate conditions that minimize template bias while retaining the benefits of high-precision localization, and we show that molecular features not present in the template can be reconstructed at high resolution from targets found by 2DTM, extending prior work at low-resolution. Moreover, we present a quantitative metric for template bias to aid the interpretation of 3D reconstructions calculated with particles localized using high-resolution templates and fine angular sampling.

Source: eLife

DOI: 10.7554/elife.90486

Authors: Bronwyn A Lucas, Benjamin A Himes, Nikolaus Grigorieff

Institutions: University of Massachusetts Chan Medical School, University of California, Berkeley, Howard Hughes Medical Institute

Language: English

Topic: Advanced Electron Microscopy Techniques and Applications

Subfield: Structural Biology

Field: Biochemistry, Genetics and Molecular Biology

Domain: Life Sciences

Open Access status: gold

Estimated APC paid: $2,500

Funders: Howard Hughes Medical Institute, Chan Zuckerberg Initiative