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Critical Assessment of Structure Prediction (CASP) is an organization aimed at advancing the state of the art in computing protein structure from sequence. In the spring of 2020, CASP launched a community project to compute the structures of the most structurally challenging proteins coded for in the SARS‐CoV‐2 genome. Forty‐seven research groups submitted over 3000 three‐dimensional models and 700 sets of accuracy estimates on 10 proteins. The resulting models were released to the public. CASP community members also worked together to provide estimates of local and global accuracy and identify structure‐based domain boundaries for some proteins. Subsequently, two of these structures (ORF3a and ORF8) have been solved experimentally, allowing assessment of both model quality and the accuracy estimates. Models from the AlphaFold2 group were found to have good agreement with the experimental structures, with main chain GDT_TS accuracy scores ranging from 63 (a correct topology) to 87 (competitive with experiment).

Substantial progresses in protein structure prediction have been made by utilizing deep‐learning and residue‐residue distance prediction since CASP13. Inspired by the advances, we improve our CASP14 MULTICOM protein structure prediction system by incorporating three new components: (a) a new deep learning‐based protein inter‐residue distance predictor to improve template‐free (ab initio) tertiary structure prediction, (b) an enhanced template‐based tertiary structure prediction method, and (c) distance‐based model quality assessment methods empowered by deep learning. In the 2020 CASP14 experiment, MULTICOM predictor was ranked seventh out of 146 predictors in tertiary structure prediction and ranked third out of 136 predictors in inter‐domain structure prediction. The results demonstrate that the template‐free modeling based on deep learning and residue‐residue distance prediction can predict the correct topology for almost all template‐based modeling targets and a majority of hard targets (template‐free targets or targets whose templates cannot be recognized), which is a significant improvement over the CASP13 MULTICOM predictor. Moreover, the template‐free modeling performs better than the template‐based modeling on not only hard targets but also the targets that have homologous templates. The performance of the template‐free modeling largely depends on the accuracy of distance prediction closely related to the quality of multiple sequence alignments. The structural model quality assessment works well on targets for which enough good models can be predicted, but it may perform poorly when only a few good models are predicted for a hard target and the distribution of model quality scores is highly skewed. MULTICOM is available athttps://github.com/jianlin-cheng/MULTICOM_Human_CASP14/tree/CASP14_DeepRank3andhttps://github.com/multicom-toolbox/multicom/tree/multicom_v2.0.

Protein structure prediction is an important problem in bioinformatics and has been studied for decades. However, there are still few open-source comprehensive protein structure prediction packages publicly available in the field. In this paper, we present our latest open-source protein tertiary structure prediction system—MULTICOM2, an integration of template-based modeling (TBM) and template-free modeling (FM) methods. The template-based modeling uses sequence alignment tools with deep multiple sequence alignments to search for structural templates, which are much faster and more accurate than MULTICOM1. The template-free (ab initio or de novo) modeling uses the inter-residue distances predicted by DeepDist to reconstruct tertiary structure models without using any known structure as template. In the blind CASP14 experiment, the average TM-score of the models predicted by our server predictor based on the MULTICOM2 system is 0.720 for 58 TBM (regular) domains and 0.514 for 38 FM and FM/TBM (hard) domains, indicating that MULTICOM2 is capable of predicting good tertiary structures across the board. It can predict the correct fold for 76 CASP14 domains (95% regular domains and 55% hard domains) if only one prediction is made for a domain. The success rate is increased to 3% for both regular and hard domains if five predictions are made per domain. Moreover, the prediction accuracy of the pure template-free structure modeling method on both TBM and FM targets is very close to the combination of template-based and template-free modeling methods. This demonstrates that the distance-based template-free modeling method powered by deep learning can largely replace the traditional template-based modeling method even on TBM targets that TBM methods used to dominate and therefore provides a uniform structure modeling approach to any protein. Finally, on the 38 CASP14 FM and FM/TBM hard domains, MULTICOM2 server predictors (MULTICOM-HYBRID, MULTICOM-DEEP, MULTICOM-DIST) were ranked among the top 20 automated server predictors in the CASP14 experiment. After combining multiple predictors from the same research group as one entry, MULTICOM-HYBRID was ranked no. 5. The source code of MULTICOM2 is freely available athttps://github.com/multicom-toolbox/multicom/tree/multicom_v2.0.

Deep learning has emerged as a revolutionary technology for protein residue‐residue contact prediction since the 2012 CASP10 competition. Considerable advancements in the predictive power of the deep learning‐based contact predictions have been achieved since then. However, little effort has been put into interpreting the black‐box deep learning methods. Algorithms that can interpret the relationship between predicted contact maps and the internal mechanism of the deep learning architectures are needed to explore the essential components of contact inference and improve their explainability. In this study, we present an attention‐based convolutional neural network for protein contact prediction, which consists of two attention mechanism‐based modules: sequence attention and regional attention. Our benchmark results on the CASP13 free‐modeling targets demonstrate that the two attention modules added on top of existing typical deep learning models exhibit a complementary effect that contributes to prediction improvements. More importantly, the inclusion of the attention mechanism provides interpretable patterns that contain useful insights into the key fold‐determining residues in proteins. We expect the attention‐based model can provide a reliable and practically interpretable technique that helps break the current bottlenecks in explaining deep neural networks for contact prediction. The source code of our method is available athttps://github.com/jianlin-cheng/InterpretContactMap.

Accurate prediction of protein secondary structure (alpha‐helix, beta‐strand and coil) is a crucial step for protein inter‐residue contact prediction and ab initio tertiary structure prediction. In a previous study, we developed a deep belief network‐based protein secondary structure method (DNSS1) and successfully advanced the prediction accuracy beyond 80%. In this work, we developed multiple advanced deep learning architectures (DNSS2) to further improve secondary structure prediction. The major improvements over the DNSS1 method include (a) designing and integrating six advanced one‐dimensional deep convolutional/recurrent/residual/memory/fractal/inception networks to predict 3‐state and 8‐state secondary structure, and (b) using more sensitive profile features inferred from Hidden Markov model (HMM) and multiple sequence alignment (MSA). Most of the deep learning architectures are novel for protein secondary structure prediction. DNSS2 was systematically benchmarked on independent test data sets with eight state‐of‐art tools and consistently ranked as one of the best methods. Particularly, DNSS2 was tested on the protein targets of 2018 CASP13 experiment and achieved the Q3 score of 81.62%, SOV score of 72.19%, and Q8 score of 73.28%. DNSS2 is freely available at:https://github.com/multicom-toolbox/DNSS2.

This paper describes outcomes of the 2019 Cryo-EM Model Challenge. The goals were to (1) assess the quality of models that can be produced from cryogenic electron microscopy (cryo-EM) maps using current modeling software, (2) evaluate reproducibility of modeling results from different software developers and users and (3) compare performance of current metrics used for model evaluation, particularly Fit-to-Map metrics, with focus on near-atomic resolution. Our findings demonstrate the relatively high accuracy and reproducibility of cryo-EM models derived by 13 participating teams from four benchmark maps, including three forming a resolution series (1.8 to 3.1 Å). The results permit specific recommendations to be made about validating near-atomic cryo-EM structures both in the context of individual experiments and structure data archives such as the Protein Data Bank. We recommend the adoption of multiple scoring parameters to provide full and objective annotation and assessment of the model, reflective of the observed cryo-EM map density.

Many proteins are composed of several domains that pack together into a complex tertiary structure. Multidomain proteins can be challenging for protein structure modeling, particularly those for which templates can be found for individual domains but not for the entire sequence. In such cases, homology modeling can generate high quality models of the domains but not for the orientations between domains. Small‐angle X‐ray scattering (SAXS) reports the structural properties of entire proteins and has the potential for guiding homology modeling of multidomain proteins. In this article, we describe a novel multidomain protein assembly modeling method, SAXSDom that integrates experimental knowledge from SAXS with probabilistic Input‐Output Hidden Markov model to assemble the structures of individual domains together. Four SAXS‐based scoring functions were developed and tested, and the method was evaluated on multidomain proteins from two public datasets. Incorporation of SAXS information improved the accuracy of domain assembly for 40 out of 46 critical assessment of protein structure prediction multidomain protein targets and 45 out of 73 multidomain protein targets from the ab initio domain assembly dataset. The results demonstrate that SAXS data can provide useful information to improve the accuracy of domain‐domain assembly. The source code and tool packages are available athttps://github.com/jianlin-cheng/SAXSDom.

Methods to reliably estimate the accuracy of 3D models of proteins are both a fundamental part of most protein folding pipelines and important for reliable identification of the best models when multiple pipelines are used. Here, we describe the progress made from CASP12 to CASP13 in the field of estimation of model accuracy (EMA) as seen from the progress of the most successful methods in CASP13. We show small but clear progress, that is, several methods perform better than the best methods from CASP12 when tested on CASP13 EMA targets. Some progress is driven by applying deep learning and residue‐residue contacts to model accuracy prediction. We show that the best EMA methods select better models than the best servers in CASP13, but that there exists a great potential to improve this further. Also, according to the evaluation criteria based on local similarities, such as lDDT and CAD, it is now clear that single model accuracy methods perform relatively better than consensus‐based methods.