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Title: Crystal structures of the human 4-1BB receptor bound to its ligand 4-1BBL reveal covalent receptor dimerization as a potential signaling amplifier

Human (h)4-1BB (TNFRSF9 or CD137) is an inducible tumor necrosis factor receptor (TNFR) superfamily member that interacts with its cognate ligand h4-1BBL to promote T lymphocyte activation and proliferation. h4-1BB is currently being targeted with agonists in cancer immunotherapy. Here, we determined the crystal structures of unbound h4-1BBL and both WT h4-1BB and a dimerization-deficient h4-1BB mutant (C121S) in complex with h4-1BBL at resolutions between 2.7 and 3.2 Å. Here, we observed that the structural arrangement of 4-1BBL, both unbound and in the complex, represents the canonical bell shape as seen in other similar TNF proteins and differs from the previously reported three-bladed propeller structure of 4-1BBL. We also found that the binding site for the receptor is at the crevice formed between two protomers of h4-1BBL, but that h4-1BB interacts predominantly with only one ligand protomer. Moreover, h4-1BBL lacked the conserved tyrosine residue in the DE loop that forms canonical interactions between other TNFR family molecules and their ligands, suggesting h4-1BBL engages h4-1BB through a distinct mechanism. Of note, we discovered that h4-1BB forms a disulfide-linked dimer because of the presence of an additional cysteine residue found in its cysteine-rich domain 4 (CRD4). As a result, h4-1BB dimerization, inmore » addition to trimerization via h4-1BBL binding, could result in cross-linking of individual ligand–receptor complexes to form a 2D network that stimulates strong h4-1BB signaling. Finally, this work provides critical insights into the structural and functional properties of both h4-1BB and h4-1BBL and reveals that covalent receptor dimerization amplifies h4-1BB signaling.« less
 [1] ;  [2] ;  [3] ; ORCiD logo [4]
  1. La Jolla Inst. for Allergy and Immunology (LJI), La Jolla, CA (United States). Division of Immune Regulation
  2. SLAC National Accelerator Lab., Menlo Park, CA (United States). Stanford Synchrotron Radiation Lightsource (SSRL)
  3. La Jolla Inst. for Allergy and Immunology (LJI), La Jolla, CA (United States). Division of Immune Regulation; Univ. of California, San Diego, CA (United States). Dept. of Medicine
  4. La Jolla Inst. for Allergy and Immunology (LJI), La Jolla, CA (United States). Division of Immune Regulation; Ghent Univ., Ghent (Belgium). Dept. of Internal Medicine, Faculty of Medicine and Health Sciences
Publication Date:
Grant/Contract Number:
AC02-76SF00515; AI110929
Accepted Manuscript
Journal Name:
Journal of Biological Chemistry
Additional Journal Information:
Journal Volume: 293; Journal Issue: 26; Journal ID: ISSN 0021-9258
American Society for Biochemistry and Molecular Biology
Research Org:
SLAC National Accelerator Lab., Menlo Park, CA (United States)
Sponsoring Org:
USDOE; National Institutes of Health (NIH)
Country of Publication:
United States
59 BASIC BIOLOGICAL SCIENCES; tumor necrosis factor (TNF); protein-protein interaction; recombinant protein expression; cell surface receptor; protein structure X-ray crystallography
OSTI Identifier: