SARS‐CoV‐2 is responsible for a disruptive worldwide viral pandemic, and renders a severe respiratory disease known as COVID‐19. Spike protein of SARS‐CoV‐2 mediates viral entry into host cells by binding ACE2 through the receptor‐binding domain (RBD). RBD is an important target for development of virus inhibitors, neutralizing antibodies, and vaccines. RBD expressed in mammalian cells suffers from low expression yield and high cost. E. coli is a popular host for protein expression, which has the advantage of easy scalability with low cost. However, RBD expressed by E. coli (RBD‐1) lacks the glycosylation, and its antigenic epitopes may not be sufficiently exposed. In the present study, RBD‐1 was expressed by E. coli and purified by a Ni Sepharose Fast Flow column. RBD‐1 was structurally characterized and compared with RBD expressed by the HEK293 cells (RBD‐2). The secondary structure and tertiary structure of RBD‐1 were largely maintained without glycosylation. In particular, the major β‐sheet content of RBD‐1 was almost unaltered. RBD‐1 could strongly bind ACE2 with a dissociation constant (KD) of 2.98 × 10–8 M. Thus, RBD‐1 was expected to apply in the vaccine development, screening drugs and virus test kit.

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The diseases caused by Zika virus (ZIKV) have received widespread concerns. As a key viral element of ZIKV, E protein was an ideal antigen for vaccine development. However, the poor immunogenicity of E protein necessitated the formulation with adjuvants. Formulation of E protein by conjugation with β-glucan was a strategy to improve the immunogenicity of E protein, where β-glucan was a polysaccharide adjuvant that could activate macrophages and trigger intracellular processes. However, the antigenic epitopes of E protein and the immunomodulatory sites of β-glucan were shielded in the conjugate. Moreover, the conjugate might elicit the undesired immune response to β-glucan. Thus, the acidic-labile hydrazone and the thiol-sensitive disulfide bonds were used as the linkers between E protein and β-glucan. Hydrazone hydrolysis and disulfide reduction could sufficiently detach the two components in the immune cells to overcome the two disadvantages. As compared with the conjugate without the two linkers, the conjugate with the two linkers (E-PS-4) elicited high E protein-specific IgG titers and low β-glucan-specific IgG titers. E-PS-4 elicited high levels of IFN-γ, TNF-α, IL-2, and IL-10. Moreover, E-PS-4 greatly facilitated the activation of dendritic cells without significant toxicity to the organs. A pharmacokinetic study revealed that the serum duration of E-PS-4 was longer than that of E protein. Accordingly, conjugation of E protein with β-glucan by the hydrazone and disulfide linkers could promote a potent cellular and humoral immune response to E protein. Thus, our study could facilitate the development of an effective vaccine against ZIKV.