Fig. 1: Self-assembling SARS-CoV-2 S6P-HBsAg nanoparticles.

A Schematic representation of full-length SARS-CoV-2 WA1 spike, S2P(1-1273), S2P(1-1206), S6P(1-1206) and S6P-HBsAg fusion protein. The individual ___domain of the spike or its ectodomains was colored and indicated. S2P(1-1273) is full-length spike harboring K986P and V987P mutations. S2P(1-1206) and S6P(1-1206) represent the ectodomains of S2P and S6P spanning from amino acids 1 to 1206, respectively. S6P(1-1206) contains six-proline substitutions: F817P, A892P, A899P, A942P, K986P and V987P. Both S2P(1-1206) and S6P(1-1206) contain RRAR-to-GSAS substitution at the furin cleavage site. S6P-HBsAg fusion was constructed by joining S6P(1-1206) and HBsAg from amino acid 1 to 226 with a linker of GS repeats. B Constructs tested for nanoparticle formation on a HBsAg core. The spike-HBsAg constructs for SARS-CoV-2 WA1 spike, S2P and S6P with a 12-GS linker were named S-12-HBsAg, S2P-12-HBsAg and S6P-12-HBsAg, respectively. Varying linkers of 8- to 24-GS were tested and named accordingly. C Negative-stain EM images of S6P-8-HBsAg (left), S6P-12-HBsAg (middle) and S6P-16-HBsAg (right) from the 50% sucrose fractions. The insets show 2D class averages of corresponding nanoparticle constructs. Scale bars represent 100 nm in micrographs and 20 nm in 2D class average images. D Human ACE2 binding and antigenicity of S6P-HBsAgs. Plates were coated with 1 µg/ml of SARS-CoV-2 S2P, S6P or S6P-HBsAg nanoparticles. Human ACE2 was assessed at a concentration ranging from 6.1 ng/ml to 100 µg/ml. The mAbs specific to the NTD, SD1, S2 or RBD ___domain of SARS-CoV-2 spike were tested from 1 pg/ml to 4 µg/ml. All plots were color-coded according to the legends on the right.