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An update for AlphaFold3 versus experimental structures: assessing the precision of small molecule binding in GPCRs

Acta Pharmacologica Sinica (2025)Cite this article

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Abstract

G protein-coupled receptors (GPCRs) are key drug discovery targets with many of them modulated by small molecules via diverse binding mechanisms. AlphaFold3, a leading structure prediction tool, models GPCR-small molecule complexes, but its accuracy remains insufficiently evaluated. In this study we compared 74 AlphaFold3-predicted structures to experimental counterparts. We showed that while AlphaFold3 accurately captured global receptor architecture and orthosteric binding pockets, which was consistent with our previous research, its ligand positioning was highly variable and often inaccurate, rendering predictions unreliable, particularly for allosteric modulators. The significant divergence from experimental structures, particularly for complex ligand interactions, highlighted AlphaFold3’s limitations and underscored that experimental structures remained essential for validating ligand-binding accuracy in GPCR complexes. These findings suggest that while AlphaFold3 offers potential for structure-based drug design, its current inaccuracies necessitate substantial refinement and integration with experimental data. This study highlights the limitation of AlphaFold3 in predicting small molecule binding and reinforces the critical role of high-resolution experimental validation for reliable GPCR-ligand interactions.

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Fig. 1: Evaluation of AlphaFold3 prediction accuracy for small-molecule-GPCR Complexes.
Fig. 2: Evaluation of AlphaFold3 prediction accuracy for class A and B GPCR complexes bound with small-molecule ligands.
Fig. 3: Evaluation of AlphaFold3 prediction accuracy for class C GPCR complexes bound with small-molecule ligands.
Fig. 4: Evaluation of AlphaFold3 prediction accuracy for GPCR complexes bound with small-molecule PAMs or NAMs.
Fig. 5: Evaluation of AlphaFold3 prediction of Global Cα RMSD of TM6 of inactive GPCRs.
Fig. 6: Comparison of molecular docking with AlphaFold3.

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  • 23 July 2025

    In this article the acknowledgment section has been omitted. This has been corrected.

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Acknowledgments

This work was supported by National Natural Science Foundation of China (32130022 and 82121005 to HEX); the National Key R&D Program of China (2022YFC2703105 to HEX, 2019YFA0904200); CAS Strategic Priority Research Program (XDB37030103 to HEX); Shanghai Municipal Science and Technology Major Project (2019SHZDZX02 to HEX); the Lingang Laboratory (LG-GG-202204-01 to HEX); State Key Laboratory of Drug Research (SKLDR-2023-TT-04 to HEX).

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Authors and Affiliations

  1. State Key Laboratory of Drug Research and CAS Key Laboratory of Receptor Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, 201203, China

    Shi-yi Shen, Jun-rui Li, H. Eric Xu & Xin-heng He

  2. University of Chinese Academy of Sciences, Beijing, 100049, China

    Shi-yi Shen, H. Eric Xu & Xin-heng He

  3. Institute of Artificial Intelligence and Robotics, Xi’an Jiaotong University, Xi’an, 710049, China

    Yu-song Wang

  4. Department of Computer Science and Engineering, The Chinese University of Hong Kong, Hong Kong, China

    Shao-ning Li

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  1. Shi-yi Shen
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  2. Jun-rui Li
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  5. H. Eric Xu
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Contributions

XHH, JRL, YSW and SNL deployed AlphaFold3 locally; SYS evaluated and analyzed the structures and data, and performed molecular docking; SYS, XHH and HEX wrote the paper.

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Correspondence to H. Eric Xu or Xin-heng He.

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Shen, Sy., Li, Jr., Wang, Ys. et al. An update for AlphaFold3 versus experimental structures: assessing the precision of small molecule binding in GPCRs. Acta Pharmacol Sin (2025). https://doi.org/10.1038/s41401-025-01617-4

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