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oai:figshare.com:article/34047370 2026-10-01T17:22:55Z category_8 category_12 category_24 category_272 category_39 category_46 category_734 category_64 category_132 category_134 portal_5 item_type_1 month_year_10_2026

Cryo-EM structure of RhGB07 trimeric spike in a 3-RBD down state. Nazia Thakur (9904162) Joseph Newman (5638340) Dongchun Ni (15188356) Jeffrey Seow (9415776) Abigail L. Hay (25157576) Yeonjae Lee (12256119) Ayush Upadhyay (25157573) Babatunde E. Ekundayo (25157578) John A. Hammond (7431785) Thomas P. Peacock (6536120) Kelvin Lau (1847764) Katie J. Doores (346210) Dalan Bailey (193037) Microbiology Cell Biology Evolutionary Biology Environmental Sciences not elsewhere classified Ecology Immunology Biological Sciences not elsewhere classified Cancer Infectious Diseases Virology multiple bat coronaviruses g ., rc g ., civet clade 1 viruses restricted ace2 usage mammalian ace2 proteins g ., rhgb07 using monoclonal antibodies clade 1 sarbecoviruses div >< p determined host range converting enzyme 2 2 convalescent individuals ace2 ), via using binding assays antigenicity using sera related bat sarbecoviruses using sarbecoviruses host range clade 3 pangolin ), neutralising antibodies entry assays binding domain sarbecoviruses interact sarbecoviruses exhibited rodent ace2 like rhgb07 generalist ace2 related antigenicity zoonotic potential vitally important still evidence specialist </ shifting pattern robust evidence research indicates racoon dog novel structure largely maintained including human immunodominant target human primate generalist phenotypes generalist </ especially omicron entry phenotypes diverse panel continued evolution breakthrough infections (A) Left: CryoEM density of the trimer spike coloured by the protomer chains the density originates from. Glycans are highlighted in teal. Centre: the trimeric spike viewed from the top rendered in cartoon representation. Right: the isolated RhGB07 RBD represented as cartoon in grey with two glycans of interest shown. (B) The C-alpha trace of the RBD of RhGB07 rendered as balls-and-sticks superimposed with the RBD of SARS-CoV-1 (PDB 7DF3) and SARS-CoV-2 (PDB 5X5B). The insets show a zoomed-in view of two regions of the RBD that show the greatest changes in the position of the C-alpha of the RBD. The region containing the ins1-loop and delta4-loop coincides with putative ACE2-interacting residues based on mapping the equivalent residues of SARS-CoV-1 and SARS-CoV-2. The ACE2-interacting residues were determined by superimposing SARS-CoV-1 and RhGB07 RBD onto the SARS-CoV-2 RBD – ACE2 (PDB: 6M0J) structure and identifying interacting residues. Highlighted residues of RhGB07 show the putative interacting residues that differ in identity to SARS-CoV-2. (C) Schematic depicting mutants generated of RhGB07 and SARS-CoV-2 Spikes with either full RBD swaps, or insertions/deletions, respectively. Lentiviral pseudotypes bearing Spikes of RhGB07 or SARS-CoV-2 mutants (RhGB07 wildtype, RhGB07SARS-CoV-2 RBD, RhGB07437_438insGGNY, SARS-CoV-2 wildtype, SARS-CoV-2RhGB07 RBD, SARS-CoV-2 Δ446−449), were used to infect BHK-21 cells overexpressing human ACE2. A heatmap illustrating entry is shown, representing the mean log RLU of 3 separate experiments. (D) ELISA data depicting the binding curves and calculated EC50 values (µg/ml) of recombinant RhGB07, SARS-CoV-2, RhGB07 437_438insGGNY or SARS-CoV-2 Δ446−449 RBDs with recombinant human ACE2. (E) Alignment of the Δ4-loop region of RhGB07 with SARS-CoV-2, SARS-CoV-2, RaTG13, Rs4231, and Rc-o319 (RhGB07 numbering), with differences in amino acid residues shown. RhGB07 was mutated to introduce an insertion (437_438insGGNY, underlined) or additional mutations (mutated residues coloured in pink) in the Δ4-loop. A heatmap illustrating entry data is shown for pseudoviruses bearing these mutant Spikes, used to infect BHK-21 cells overexpressing Rh pusillus, Rh. affinis, Rh. ferrumequinum, Rh. cornutus, Rh. alcyone and M. lucifugus ACE2s. Data represents the mean log RLU of 4 separate experiments. A vector-only control (pDISPLAY) was included to demonstrate specificity and to set the background signal. The data underlying this Figure can be found in <a href="http://www.plosbiology.org/article/info:doi/10.1371/journal.pbio.3003944#pbio.3003944.s017" target="_blank">S1 Data</a>. 2026-10-01T17:40:07Z Image Figure 10.1371/journal.pbio.3003944.g002 https://figshare.com/articles/figure/_p_Cryo-EM_structure_of_RhGB07_trimeric_spike_in_a_3-RBD_down_state_p_/34047370 CC BY 4.0