A COVID-19 antibody curbs SARS-CoV-2 nucleocapsid protein-induced complement hyperactivation (preprint)

Although human antibodies elicited by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nucleocapsid (N) protein are profoundly boosted upon infection, little is known about the function of N-reactive antibodies. Herein, we isolated and profiled a panel of 32 N protein-specific monoclonal antibodies (mAbs) from a quick recovery coronavirus disease-19 (COVID-19) convalescent patient who had dominant antibody responses to the SARS-CoV-2 N protein rather than to the SARS-CoV-2 spike (S) protein. The complex structure of the N protein RNA binding domain with the mAb with the highest binding affinity (nCoV396) revealed changes in the epitopes and antigen’s allosteric regulation. Functionally, a virus-free complement hyper-activation analysis demonstrated that nCoV396 specifically compromises the N protein-induced complement hyper-activation, which is a risk factor for the morbidity and mortality of COVID-19 patients, thus laying the foundation for the identification of functional anti-N protein mAbs.

Structural Insight Reveals SARS-CoV-2 Orf7a as an Immunomodulating Factor for Human CD14+ Monocytes (preprint)

Dysregulated immune cell responses have been linked to the severity of Coronavirus Disease 2019 (COVID-19). However, the specific viral factor of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) contributing to the immune-dysregulation is currently unclear. Herein, we identified the ectodomain of Ig-like fold viral proteinSARS-CoV-2 Orf7a interacted with CD14+ monocytes at the highest efficiency in human peripheral blood mononuclear cells, but not for the relative highly pathogenic protein SARS-CoV Orf7a. The 2.2 Å resolutioncrystal structure of SARS-CoV-2 Orf7a reveals three remarkable changes in the amphipathic side of the four-strand β-sheet, implying the potential functional interface of the viral protein. Structure-based superimposition of SARS-CoV-2 Orf7a with SARS-CoV Orf7a - LFA1 working model suggests that SARS-CoV-2 Orf7a utilizes different binding patterns to recognize the specific immune cells. Importantly, SARS-CoV-2 Orf7a co-incubation with CD14+ monocytes ex vivo triggers a decrease of HLA-DR/DP/DQ and significant pro-inflammatory cytokines expressions, including IL-6, IL-1β, IL-8, and TNF-α. Our work demonstrates that SARS-CoV-2 Orf7a is an immunomodulating factor for immune cells binding and trigging aberrant inflammatory responses, providing promising therapeutic drug targets in the pandemic disease COVID-19.Funding: This work is supported by the National Key R&D Program of China (2019YFA0110300) granted to J.C.; the Special Fund for Scientific and Technological Innovation Strategy of Guangdong Province of China (2018B030306029), COVID-19 Emerging Prevention Products, Research Special Fund of Zhuhai City (ZH22036302200016PWC) granted to S.C.; the Fundamental Research Funds for the Central Universities (19ykzd36) granted to J.C; and the Science and Technology Program of Guangzhou (202002030069) granted to J.C. Conflict of Interest: The authors declare no conflict of interest.Ethical Approval: The human peripheral blood samples for the experiments were collected through The Health Management Center, The Fifth Affiliated Hospital, Sun Yat-sen University. This study was approved by The Medical Ethics Committee of The Fifth Affiliated Hospital, Sun Yat-sen University (2020-K195-1).

A COVID-19 antibody curbs SARS-CoV-2 nucleocapsid protein-induced complement hyper-activation (preprint)

Although human antibodies elicited by severe acute respiratory distress syndrome coronavirus-2 (SARS-CoV-2) nucleocapsid (N) protein are profoundly boosted upon infection, little is known about the function of N-directed antibodies. Herein, we isolated and profiled a panel of 32 N protein-specific monoclonal antibodies (mAb) from a quick recovery coronavirus disease-19 (COVID-19) convalescent, who had dominant antibody responses to SARS-CoV-2 N protein rather than to Spike protein. The complex structure of N protein RNA binding domain with the highest binding affinity mAb nCoV396 reveals the epitopes and antigens allosteric changes. Functionally, a virus-free complement hyper-activation analysis demonstrates that nCoV396 specifically compromises N protein-induced complement hyper-activation, a risk factor for morbidity and mortality in COVID-19, thus paving the way for functional anti-N mAbs identification. One Sentence SummaryB cell profiling, structural determination, and protease activity assays identify a functional antibody to N protein.

Crystal structure of SARS-CoV-2 nucleocapsid protein RNA binding domain reveals potential unique drug targeting sites (preprint)

The outbreak of coronavirus disease (COVID-19) in China caused by SARS-CoV-2 virus continually lead to worldwide human infections and deaths. It is currently no specific viral protein targeted therapeutics yet. Viral nucleocapsid protein is a potential antiviral drug target, serving multiple critical functions during the viral life cycle. However, the structural information of SARS-CoV-2 nucleocapsid protein is yet to be clear. Herein, we have determined the 2.7 [A] crystal structure of the N-terminal RNA binding domain of SARS-CoV-2 nucleocapsid protein. Although overall structure is similar with other reported coronavirus nucleocapsid protein N-terminal domain, the surface electrostatic potential characteristics between them are distinct. Further comparison with mild virus type HCoV-OC43 equivalent domain demonstrates a unique potential RNA binding pocket alongside the {beta}-sheet core. Complemented by in vitro binding studies, our data provide several atomic resolution features of SARS-CoV-2 nucleocapsid protein N-terminal domain, guiding the design of novel antiviral agents specific targeting to SARS-CoV-2.