2. Academic Validation
  3. Design of minibinder proteins as universal antagonists against canine and human TNFα

Design of minibinder proteins as universal antagonists against canine and human TNFα

  • Commun Biol. 2025 Nov 24;8(1):1639. doi: 10.1038/s42003-025-09030-7.
Jun Weng # 1 2 3,  Zhiyong Wu # 1 2,  Banbin Xing # 1 2,  Yang Hu # 1 2,  Xiaoyu Hu # 1 2,  Meng Mei 1 2,  Jiaxin Xu 1 2,  Mengqing Lu 1 2,  Yibin Chen 1 2,  Lin Wei 1 2,  Ke Ming 1 2,  Zhizheng Wang 1 2,  Zhuang Li 1 2,  Zigong Wei 4 5 6
Affiliations

Affiliations

  • 1 State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan, Hubei, China.
  • 2 National & Local Joint Engineering Research Center of High-throughput Drug Screening Technology, School of Life Sciences, Hubei University, Wuhan, Hubei, China.
  • 3 Key Laboratory of Molecular Biophysics of Ministry of Education, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei, China.
  • 4 State Key Laboratory of Biocatalysis and Enzyme Engineering, School of Life Sciences, Hubei University, Wuhan, Hubei, China. [email protected].
  • 5 National & Local Joint Engineering Research Center of High-throughput Drug Screening Technology, School of Life Sciences, Hubei University, Wuhan, Hubei, China. [email protected].
  • 6 Hubei Jiangxia Laboratory, Wuhan, Hubei, China. [email protected].
  • # Contributed equally.
PMID: 41286500 DOI: 10.1038/s42003-025-09030-7
Abstract

In this study, we resolve the complex structure of canine TNFα (cTNFα) trimer bound to nanobody molecules at a resolution of 3.1 Å using cryo-electron microscopy. Structural comparison between cTNFα and human TNFα (hTNFα) reveals that the non-conserved residue Phe83 in cTNFα induces a conformational change in a nearby loop, which significantly reduces the binding affinity of cTNFα to nanobody TNF30. By analyzing the simulated complex structures of cTNFα with its receptors and resolved structures of hTNFα in complex with its receptors, we identify a conserved hydrophobic region involved in ligand-receptor interaction on both TNFαs. Five hydrophobic residues within this region are determined as target hotspots to allow de novo computational design of universal minibinders against both canine and human TNFα. Purified top-ranked designs exhibit sub-nanomolar to nanomolar affinity toward cTNFα as well as hTNFα, and show anti-TNFα activities to both TNFαs in cell-based assays that are comparable or superior to the anti-hTNFα activity of TNF30. This work provides a practical approach to generate universal TNFα antagonists as promising lead molecules for the development of potent cross-species biologics applicable in both humans and canines.

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