Coronavirus disease 2019 , caused by the novel human coronavirus SARS-CoV-2, is currently a major threat to public health worldwide. The viral spike protein binds the host receptor angiotensin-converting enzyme 2 (ACE2) via the receptor-binding domain (RBD), and thus is believed to be a major target to block viral entry. Both SARS-CoV-2 and SARS-CoV share this mechanism.Here we functionally analyzed the key amino acid residues located within receptor binding motif of RBD that may interact with human ACE2 and available neutralizing antibodies. The in vivo experiments showed that immunization with either the SARS-CoV RBD or SARS-CoV-2 RBD was able to induce strong clade-specific neutralizing antibodies in mice; however, the cross-neutralizing activity was much weaker, indicating that there are distinct antigenic features in the RBDs of the two viruses. This finding was confirmed with the available neutralizing monoclonal antibodies against SARS-CoV or SARS-CoV-2. It is worth noting that a newly developed SARS-CoV-2 human antibody, HA001, was able to neutralize SARS-CoV-2, but failed to recognize SARS-CoV. Moreover, the potential epitope residues of HA001 were identified as A475 and F486 in the SARS-CoV-2 RBD, representing new binding sites for neutralizing antibodies. Overall, our study has revealed the presence of different key epitopes between SARS-CoV and SARS-CoV-2, which indicates the necessity to develop new prophylactic vaccine and antibody drugs for specific control of the COVID-19 pandemic although the available agents obtained from the SARS-CoV study are unneglectable.

Fourteen ORFs have been identified in the severe acute respiratory syndrome-associated coronavirus (SARS-CoV) genome. ORF 3a of SARS-CoV codes for a recently identified transmembrane protein, but its function remains unknown. In this study we confirmed the 3a protein expression and investigated its localization at the surface of SARS-CoV-infected or 3a-cDNA-transfected cells. Our experiments showed that recombinant 3a protein can form a homotetramer complex through interprotein disulfide bridges in 3a-cDNA-transfected cells, providing a clue to ion channel function. The putative ion channel activity of this protein was assessed in 3a-complement RNA-injected Xenopus oocytes by two-electrode voltage clamp. The results suggest that 3a protein forms a potassium sensitive channel, which can be efficiently inhibited by barium. After FRhK-4 cells were transfected with an siRNA, which is known to suppress 3a expression, followed by infection with SARS-CoV, the released virus was significantly decreased, whereas the replication of the virus in the infected cells was not changed. Our observation suggests that SARS-CoV ORF 3a functions as an ion channel that may promote virus release. This finding will help to explain the highly pathogenic nature of SARS-CoV and to develop new strategies for treatment of SARS infection. caused alarm all over the world. The newly discovered human coronavirus named SARS-associated coronavirus (SARS-CoV) was identified as the causative agent for this disease (1, 2). SARSCoV has a large single-positive-strand RNA genome that contains 14 ORFs. Some of these ORFs encode viral structural proteins, such as spike protein, membrane protein, small envelope protein, and nucleocapsid protein, as well as viral replicase and protease (3). Those proteins play important roles in viral infection and replication. However, functions for other ORFs are not clear. Therefore, identification and characterization of new functional proteins from the ORFs will be helpful for understanding the pathogenesis of SARS-CoV. Up to now there are still no effective drugs or vaccines against SARS-CoV. The identification of new viral proteins and the elucidation of their functions will provide potential targets for design of drugs or vaccines against SARS.Our previous work has revealed that ORF 3a of SARS-CoV is such a viral protein (4). Since then, other publications have concurred in this observation and have shown that it is a structural protein (5-8). ORF 3a is located between the S and E protein loci and encodes a protein of 274 aa. The only available information based on proteomics and immunoblotting suggests that 3a protein is structural in nature, but its localization, topology, and biological function have not been identified.A computed biology analysis of the amino acid sequence of the 3a protein revealed that it has low similarity with any other known protein. Its C-terminal region shares Ϸ50% similarity to Plasmodium calcium pump protein and to the Shewanella outer-membrane porin. Interestingly, comparison of ORFs ...