Protein–protein interactions (PPIs) are fundamental to cellular functions, yet predicting and analyzing their 3D structures remains a critical and computationally demanding challenge. To address this, the HawkDock web server was developed as an integrated computational platform for predicting and analyzing protein–protein complexes. Over the past 6 years, HawkDock has successfully processed >234 000 computational tasks. In this study, an updated version of HawkDock was developed with the following advancements: (1) a deep learning-based flexible docking method, GeoDock, has been integrated to improve docking accuracy, particularly for apo-protein structures; (2) the VD-MM/GBSA method, which outperforms conventional MM/GBSA approaches in predicting binding affinities, has been implemented; (3) a new Mutation Analysis Module has been added to systematically evaluate the energetic impacts of amino acid mutations on protein–protein binding; (4) the server has been migrated to a high-performance cluster with Amber upgraded to version 24. Here, we describe the general protocol of HawkDock2, with a particular focus on its new features related to flexible docking, VD-MM/GBSA affinity prediction, and amino acid residue mutations. Comprehensive validation studies have demonstrated the reliability and effectiveness of these new features. HawkDock2 will remain freely accessible to all users at http://cadd.zju.edu.cn/hawkdock/.

Machine-learning (ML)-based scoring functions (MLSFs) have gradually emerged as a promising alternative for protein–ligand binding affinity prediction and structure-based virtual screening. However, clouds of doubts have still been raised against the benefits of this novel type of scoring functions (SFs). In this study, to benchmark the performance of target-specific MLSFs on a relatively unbiased dataset, the MLSFs trained from three representative protein–ligand interaction representations were assessed on the LIT-PCBA dataset, and the classical Glide SP SF and three types of ligand-based quantitative structure-activity relationship (QSAR) models were also utilized for comparison. Two major aspects in virtual screening campaigns, including prediction accuracy and hit novelty, were systematically explored. The calculation results illustrate that the tested target-specific MLSFs yielded generally superior performance over the classical Glide SP SF, but they could hardly outperform the 2D fingerprint-based QSAR models. Although substantial improvements could be achieved by integrating multiple types of protein–ligand interaction features, the MLSFs were still not sufficient to exceed MACCS-based QSAR models. In terms of the correlations between the hit ranks or the structures of the top-ranked hits, the MLSFs developed by different featurization strategies would have the ability to identify quite different hits. Nevertheless, it seems that target-specific MLSFs do not have the intrinsic attributes of a traditional SF and may not be a substitute for classical SFs. In contrast, MLSFs can be regarded as a new derivative of ligand-based QSAR models. It is expected that our study may provide valuable guidance for the assessment and further development of target-specific MLSFs.

Ribonucleic acid (RNA)–ligand interactions play a pivotal role in a wide spectrum of biological processes, ranging from protein biosynthesis to cellular reproduction. This recognition has prompted the broader acceptance of RNA as a viable candidate for drug targets. Delving into the atomic-scale understanding of RNA–ligand interactions holds paramount importance in unraveling intricate molecular mechanisms and further contributing to RNA-based drug discovery. Computational approaches, particularly molecular docking, offer an efficient way of predicting the interactions between RNA and small molecules. However, the accuracy and reliability of these predictions heavily depend on the performance of scoring functions (SFs). In contrast to the majority of SFs used in RNA–ligand docking, the end-point binding free energy calculation methods, such as molecular mechanics/generalized Born surface area (MM/GBSA) and molecular mechanics/Poisson Boltzmann surface area (MM/PBSA), stand as theoretically more rigorous approaches. Yet, the evaluation of their effectiveness in predicting both binding affinities and binding poses within RNA–ligand systems remains unexplored. This study first reported the performance of MM/PBSA and MM/GBSA with diverse solvation models, interior dielectric constants (ε_in) and force fields in the context of binding affinity prediction for 29 RNA–ligand complexes. MM/GBSA is based on short (5 ns) molecular dynamics (MD) simulations in an explicit solvent with the YIL force field; the GB^GBn2 model with higher interior dielectric constant (ε_in = 12, 16 or 20) yields the best correlation (R_p = −0.513), which outperforms the best correlation (R_p = −0.317, rDock) offered by various docking programs. Then, the efficacy of MM/GBSA in identifying the near-native binding poses from the decoys was assessed based on 56 RNA–ligand complexes. However, it is evident that MM/GBSA has limitations in accurately predicting binding poses for RNA–ligand systems, particularly compared with notably proficient docking programs like rDock and PLANTS. The best top-1 success rate achieved by MM/GBSA rescoring is 39.3%, which falls below the best results given by docking programs (50%, PLNATS). This study represents the first evaluation of MM/PBSA and MM/GBSA for RNA–ligand systems and is expected to provide valuable insights into their successful application to RNA targets.

Compound–protein interactions (CPI) play significant roles in drug development. To avoid side effects, it is also crucial to evaluate drug selectivity when binding to different targets. However, most selectivity prediction models are constructed for specific targets with limited data. In this study, we present a pretrained multi-functional model for compound–protein interaction prediction (PMF-CPI) and fine-tune it to assess drug selectivity. This model uses recurrent neural networks to process the protein embedding based on the pretrained language model TAPE, extracts molecular information from a graph encoder, and produces the output from dense layers. PMF-CPI obtained the best performance compared to outstanding approaches on both the binding affinity regression and CPI classification tasks. Meanwhile, we apply the model to analyzing drug selectivity after fine-tuning it on three datasets related to specific targets, including human cytochrome P450s. The study shows that PMF-CPI can accurately predict different drug affinities or opposite interactions toward similar targets, recognizing selective drugs for precise therapeutics.Kindly confirm if corresponding authors affiliations are identified correctly and amend if any.Yes, it is correct.

Six folding states and three folding pathways are identified for HP35 with U and F being unfolded and folded states and I1, I2, I3 and I4 being intermediate states.