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Benchmarking tomographic acquisition schemes for high-resolution structural biology.

Beata Turoňová¹, Wim J H Hagen¹, Martin Obr^2,3

¹European Molecular Biology Laboratory, Structure and Computational Biology Unit, 69124, Heidelberg, Germany.

Nature Communications

|February 15, 2020

Summary

Optimizing cryo-electron tomography (cryo-ET) for macromolecular complexes requires careful parameter selection. Dose-symmetric acquisition significantly improves resolution in subtomogram averaging, offering better structural insights.

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Area of Science:

Structural biology
Biophysics
Microscopy techniques

Background:

Cryo-electron tomography (cryo-ET) with subtomogram averaging (STA) enables high-resolution structural analysis of macromolecular complexes in situ.
Understanding the impact of experimental parameters on attainable resolution in cryo-ET-STA is crucial for optimizing data acquisition.

Purpose of the Study:

To systematically evaluate experimental parameters for optimizing resolution in subtomogram averaging.
To provide guidance for data acquisition in cryo-electron tomography studies.

Main Methods:

Utilized immature HIV-1 lattice as a benchmarking sample for cryo-ET-STA.
Systematically varied parameters including projection order, angular increments, and Volta phase plate usage.
Compared different acquisition schemes to assess their impact on resolution.

Main Results:

All commonly used acquisition schemes achieved subnanometer resolution.
Dose-symmetric acquisition demonstrated a considerably better outcome for subtomogram averaging resolution.
Identified key parameters influencing the attainable resolution in cryo-ET-STA.

Conclusions:

Dose-symmetric acquisition is recommended for superior resolution in cryo-ET-STA.
Findings provide practical guidance for optimizing cryo-ET data acquisition strategies.
Publicly available data can facilitate further development of processing routines.