Yeast-expressed recombinant protein of the receptor-binding domain in SARS-CoV spike protein with deglycosylated forms as a SARS vaccine candidate

TLDR

Vaccines against SARS‑CoV are urgently needed, and prior work has shown that the receptor‑binding domain (RBD) of the spike protein can elicit potent neutralizing antibodies and protection in animals. This study aimed to improve scalable production of the RBD vaccine by eliminating glycosylation sites. Two RBD variants (RBD193 and RBD219) and their deglycosylated forms were expressed in yeast, purified, and evaluated for antigenicity, functionality, and immunogenicity in alum‑adjuvanted mice. The deglycosylated RBD219‑N1 variant exhibited high yield, preserved antigenicity, and induced markedly stronger RBD‑specific and neutralizing antibody responses than the other constructs, indicating it as the optimal candidate for further development.

Abstract

Development of vaccines for preventing a future pandemic of severe acute respiratory syndrome (SARS) caused by SARS coronavirus (SARS-CoV) and for biodefense preparedness is urgently needed. Our previous studies have shown that a candidate SARS vaccine antigen consisting of the receptor-binding domain (RBD) of SARS-CoV spike protein can induce potent neutralizing antibody responses and protection against SARS-CoV challenge in vaccinated animals. To optimize expression conditions for scale-up production of the RBD vaccine candidate, we hypothesized that this could be potentially achieved by removing glycosylation sites in the RBD protein. In this study, we constructed two RBD protein variants: 1) RBD193-WT (193-aa, residues 318-510) and its deglycosylated forms (RBD193-N1, RBD193-N2, RBD193-N3); 2) RBD219-WT (219-aa, residues 318-536) and its deglycosylated forms (RBD219-N1, RBD219-N2, and RBD219-N3). All constructs were expressed as recombinant proteins in yeast. The purified recombinant proteins of these constructs were compared for their antigenicity, functionality and immunogenicity in mice using alum as the adjuvant. We found that RBD219-N1 exhibited high expression yield, and maintained its antigenicity and functionality. More importantly, RBD219-N1 induced significantly stronger RBD-specific antibody responses and a higher level of neutralizing antibodies in immunized mice than RBD193-WT, RBD193-N1, RBD193-N3, or RBD219-WT. These results suggest that RBD219-N1 could be selected as an optimal SARS vaccine candidate for further development.