Measurement of atom resolvability in cryo-EM maps with Q-scores

Pintilie, Grigore; Zhang, Kaiming; Su, Zhaoming; Li, Shanshan; Schmid, Michael F.; Chiu, Wah

doi:10.1038/s41592-020-0731-1

Cited by 281 publications

(293 citation statements)

References 34 publications

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“…The quality of these models was validated by MolProbity (15) (SI Appendix, Table S1). The Q-scores (16) developed for evaluating the density resolvability were used to assess the quality of these cryo-EM structures (SI Appendix, Figs. S5 and S6).…”

Section: Resultsmentioning

confidence: 99%

Inhibition mechanisms of AcrF9, AcrF8, and AcrF6 against type I-F CRISPR–Cas complex revealed by cryo-EM

Zhang

Wang

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Prokaryotes and viruses have fought a long battle against each other. Prokaryotes use CRISPR–Cas-mediated adaptive immunity, while conversely, viruses evolve multiple anti-CRISPR (Acr) proteins to defeat these CRISPR–Cas systems. The type I-F CRISPR–Cas system in Pseudomonas aeruginosa requires the crRNA-guided surveillance complex (Csy complex) to recognize the invading DNA. Although some Acr proteins against the Csy complex have been reported, other relevant Acr proteins still need studies to understand their mechanisms. Here, we obtain three structures of previously unresolved Acr proteins (AcrF9, AcrF8, and AcrF6) bound to the Csy complex using electron cryo-microscopy (cryo-EM), with resolution at 2.57 Å, 3.42 Å, and 3.15 Å, respectively. The 2.57-Å structure reveals fine details for each molecular component within the Csy complex as well as the direct and water-mediated interactions between proteins and CRISPR RNA (crRNA). Our structures also show unambiguously how these Acr proteins bind differently to the Csy complex. AcrF9 binds to key DNA-binding sites on the Csy spiral backbone. AcrF6 binds at the junction between Cas7.6f and Cas8f, which is critical for DNA duplex splitting. AcrF8 binds to a distinct position on the Csy spiral backbone and forms interactions with crRNA, which has not been seen in other Acr proteins against the Csy complex. Our structure-guided mutagenesis and biochemistry experiments further support the anti-CRISPR mechanisms of these Acr proteins. Our findings support the convergent consequence of inhibiting degradation of invading DNA by these Acr proteins, albeit with different modes of interactions with the type I-F CRISPR–Cas system.

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Section: Resultsmentioning

confidence: 99%

Inhibition mechanisms of AcrF9, AcrF8, and AcrF6 against type I-F CRISPR–Cas complex revealed by cryo-EM

Zhang

Wang

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…Overfitting is one of the main challenges during model refinement and new metrics to detect it are needed (Lawson et al, 2020). The Q-score has been indicated by the authors (Pintilie et al, 2020) that is particularly sensitive to overfitting. Furthermore, we have shown that our method has less dependence on sharpening than the map Q-score and that they complement each other to better estimate the resolvability of atoms.…”

Section: -Discussionmentioning

confidence: 99%

“…The Q-score method measures the correlation between the map values at points around the atom and a reference Gaussian-like function (Pintilie et al, 2020). In this way, the Q-score estimates the resolvability of each atom with respect to the map from which the model has been refined.…”

Section: Fsc-q As a Complement To Q-scorementioning

confidence: 99%

“…However, these measures are based on the model itself, regardless of the cryoEM map. Local map-to-model fit measurements have been introduced, such as EMRinger (Barad et al, 2015), which takes into account density values near carbon-ß atom, and the recent Q-score (Pintilie et al, 2020), which measures the correlation between the map values at points around the atom and a reference Gaussian-like function.…”

Section: -Introductionmentioning

confidence: 99%

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FSC-Q: A CryoEM map-to-atomic model quality validation based on the local Fourier Shell Correlation

Ramírez-Aportela

Maluenda

Fonseca

et al. 2020

Preprint

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In recent years, advances in cryoEM have dramatically increased the resolution of Coulomb potential maps and, with it, the number of solved atomic models. It is widely accepted that the quality of cryoEM maps varies locally; therefore, the evaluation of the maps-derived structural models must be done locally as well. In this article, a method for the local analysis of the map-tomodel fit is presented. The algorithm uses a comparison of two local resolution maps. The first is the local FSC (Fourier shell correlation) between the full map and the model, while the second is calculated between the half maps normally used in typical single particle analysis workflows. We call the new quality measure "FSC-Q", and it is a quantitative estimation of how much of the model is supported by the signal content of the map. Furthermore, we show that FSC-Q may be helpful to avoid overfitting. It can be used to complement other methods, such as the Q-score method that estimates the resolvability of atoms.

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“…We used high-resolution (1.8 Å, EMD-20026) and low-resolution (3.1 Å, EMD-20028) maps of human apoferritin available from the EMDB (Lawson et al, 2011, Pintilie et al, 2020. The crystal structure of human apoferritin (Masuda et al, 2010) was docked into and refined against the 1.8 Å cryo-EM map with the Phenix tools dock_in_map (Liebschner et al, 2019) and real_space_refine .…”

Section: Apoferritin Maps and Modelmentioning

confidence: 99%

Density modification of cryo-EM maps

Terwilliger

Sobolev

Afonine

et al. 2020

Preprint

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SynopsisThe prerequisites for density modification of maps from electron cryomicroscopy are examined and a procedure for incorporating model-based information is presented. Abstract Density modification uses expectations about features of a map such as a flat solvent and expected distributions of density in the region of the macromolecule to improve individual Fourier terms representing the map. This process transfers information from one part of a map to another and can improve the accuracy of a map. Here the assumptions behind density modification for maps from electron cryomicroscopy are examined and a procedure is presented that allows incorporation of modelbased information. Density modification works best in cases where unfiltered, unmasked maps with clear boundaries between macromolecule and solvent are visible and where there is substantial noise in the map, both in the region of the macromolecule and the solvent. It also is most effective if the characteristics of the map are relatively constant within regions of the macromolecule and the solvent.Model-based information can be used to improve density modification, but model bias can in principle occur. Here model bias is reduced by using ensemble models that allow estimation of model uncertainty.A test of model bias is presented suggesting that even if the expected density in a region of a map is specified incorrectly by using an incorrect model, the incorrect expectations do not strongly affect the final map.

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Measurement of atom resolvability in cryo-EM maps with Q-scores

Cited by 281 publications

References 34 publications

Inhibition mechanisms of AcrF9, AcrF8, and AcrF6 against type I-F CRISPR–Cas complex revealed by cryo-EM

Inhibition mechanisms of AcrF9, AcrF8, and AcrF6 against type I-F CRISPR–Cas complex revealed by cryo-EM

FSC-Q: A CryoEM map-to-atomic model quality validation based on the local Fourier Shell Correlation

Density modification of cryo-EM maps

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