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Benchmarking tomographic acquisition schemes for high-resolution structural biology

By Beata Turoňová, Wim J. H. Hagen, Martin Obr, Hans-Georg Kraeusslich, Martin Beck

Posted 21 Aug 2019
bioRxiv DOI: 10.1101/742254 (published DOI: 10.1038/s41467-020-14535-2)

Cryo electron tomography with subsequent subtomogram averaging is a powerful technique to structurally analyze macromolecular complexes in their native context. Although close to atomic resolution, in principle, can be obtained, it is not clear how individual experimental parameters contribute to the attainable resolution. Here, we have used immature HIV-1 lattice as a benchmarking sample to optimize the attainable resolution for subtomogram averaging. We systematically tested various experimental parameters such as the order of projections, different angular increments and the use of the Volta phase plate. We find that although any of the prominently used acquisition schemes is sufficient to obtain subnanometer resolution, dose-symmetric acquisition provides considerably better outcome. We discuss our findings in order to provide guidance for data acquisition. Our data is publicly available at EMPIAR-10277 as well as EMD-10207 and might be used to further develop processing routines.

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