Journal: Proc Natl Acad Sci U S A / Year: 2019 Title: Cryo-EM structure of pleconaril-resistant rhinovirus-B5 complexed to the antiviral OBR-5-340 reveals unexpected binding site. Authors: Jiri Wald / Marion Pasin / Martina Richter / Christin Walther / Neann Mathai / Johannes Kirchmair / Vadim A Makarov / Nikolaus Goessweiner-Mohr / Thomas C Marlovits / Irene Zanella / Antonio ...Authors: Jiri Wald / Marion Pasin / Martina Richter / Christin Walther / Neann Mathai / Johannes Kirchmair / Vadim A Makarov / Nikolaus Goessweiner-Mohr / Thomas C Marlovits / Irene Zanella / Antonio Real-Hohn / Nuria Verdaguer / Dieter Blaas / Michaela Schmidtke / Abstract: Viral inhibitors, such as pleconaril and vapendavir, target conserved regions in the capsids of rhinoviruses (RVs) and enteroviruses (EVs) by binding to a hydrophobic pocket in viral capsid protein 1 ...Viral inhibitors, such as pleconaril and vapendavir, target conserved regions in the capsids of rhinoviruses (RVs) and enteroviruses (EVs) by binding to a hydrophobic pocket in viral capsid protein 1 (VP1). In resistant RVs and EVs, bulky residues in this pocket prevent their binding. However, recently developed pyrazolopyrimidines inhibit pleconaril-resistant RVs and EVs, and computational modeling has suggested that they also bind to the hydrophobic pocket in VP1. We studied the mechanism of inhibition of pleconaril-resistant RVs using RV-B5 (1 of the 7 naturally pleconaril-resistant rhinoviruses) and OBR-5-340, a bioavailable pyrazolopyrimidine with proven in vivo activity, and determined the 3D-structure of the protein-ligand complex to 3.6 Å with cryoelectron microscopy. Our data indicate that, similar to other capsid binders, OBR-5-340 induces thermostability and inhibits viral adsorption and uncoating. However, we found that OBR-5-340 attaches closer to the entrance of the pocket than most other capsid binders, whose viral complexes have been studied so far, showing only marginal overlaps of the attachment sites. Comparing the experimentally determined 3D structure with the control, RV-B5 incubated with solvent only and determined to 3.2 Å, revealed no gross conformational changes upon OBR-5-340 binding. The pocket of the naturally OBR-5-340-resistant RV-A89 likewise incubated with OBR-5-340 and solved to 2.9 Å was empty. Pyrazolopyrimidines have a rigid molecular scaffold and may thus be less affected by a loss of entropy upon binding. They interact with less-conserved regions than known capsid binders. Overall, pyrazolopyrimidines could be more suitable for the development of new, broadly active inhibitors.
History
Deposition
Aug 14, 2019
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Header (metadata) release
Aug 28, 2019
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Map release
Sep 4, 2019
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Update
May 22, 2024
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Current status
May 22, 2024
Processing site: PDBe / Status: Released
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Structure visualization
Movie
Surface view with section colored by density value
Model: Homemade / Material: COPPER / Support film - Material: CARBON / Support film - topology: CONTINUOUS / Support film - Film thickness: 0.4 / Pretreatment - Type: GLOW DISCHARGE
Vitrification
Cryogen name: ETHANE / Chamber humidity: 100 % / Instrument: FEI VITROBOT MARK III
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Electron microscopy
Microscope
FEI POLARA 300
Image recording
Film or detector model: GATAN K2 SUMMIT (4k x 4k) / Detector mode: COUNTING / Digitization - Frames/image: 1-25 / Number real images: 5370 / Average exposure time: 5.0 sec. / Average electron dose: 40.0 e/Å2
Electron beam
Acceleration voltage: 300 kV / Electron source: FIELD EMISSION GUN
Electron optics
Illumination mode: FLOOD BEAM / Imaging mode: BRIGHT FIELD / Cs: 2.7 mm
Experimental equipment
Model: Tecnai Polara / Image courtesy: FEI Company
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Image processing
Startup model
Type of model: NONE
Final reconstruction
Applied symmetry - Point group: C60 (60 fold cyclic) / Resolution.type: BY AUTHOR / Resolution: 2.9 Å / Resolution method: FSC 0.143 CUT-OFF / Number images used: 107054
Chain - Source name: PDB / Chain - Initial model type: experimental model
Details
Rosetta: DiMaio, F. et al. Atomic-accuracy models from 4.5-A cryo-electron microscopy data with density-guided iterative local refinement. Nat. Meth (2015). doi:10.1038/nmeth.3286
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