Human Anti-SARS-CoV-2 NP (N-term) Monoclonal Antibody, clone CO19

SARS-CoV-2 belongs to the Coronavirus family, genus Beta-coronavirus, and is an enveloped single-stranded positive-strand RNA virus with a genome of approximately 30kb and four structural proteins, i.e., spike (S) protein, envelope (E) protein, membrane (M) protein, and nucleocapsid (N) protein. Among them, the S protein is a baseball-shaped glycoprotein extending out of the envelope. It plays a key role in the binding of the virus to the host cell surface receptor and mediating membrane fusion to enter the cell, and determines the host nature of the virus. It is also the main antigenic protein of the coronavirus. The E protein is mainly distributed on the viral envelope. The M protein plays an important role in the formation and budding of the viral envelope. The NP protein binds to the viral RNA, supports the formation of the nucleocapsid, and assists in viral budding, RNA replication, and mRNA replication. The upstream of the genome mainly encodes non-structural proteins, while the downstream mainly encodes structural proteins. Due to the complexity of the genome, the new classification standard is based on the amino acid sequences of several proteins, including 3C protease (3CLpro), related nucleosyltransferase (NiRAN), RNA-dependent RNA polymerase (RdRp), zinc-binding domain covalently linked HEL1 (ZBDHEL1), and superfamily 1 helicase (HEL1). The first two-thirds of the genome encodes ORF1a and ORF1b polyproteins, which are cleaved into multiple proteins by a protease of the virus itself, which has unique protease activity. The cleaved proteins are non-structural proteins 1-16 (nsp1-16), including RdRp, Hel, 3CLpro and other important domains that play multiple functions during genome replication. Other scattered accessory genes such as ORF3, ORF4, ORF5, etc. are redundant genes. There is evidence that these genes have specific functions, such as regulating host immune responses and potentially promoting viral replication. Similar to SARS-CoV and MERS-CoV, SARS-CoV-2 mainly infects the respiratory tract, with symptoms and diseases ranging from mild respiratory infections to severe acute respiratory syndrome, the latter of which causes organ failure and ultimately death in some patients. SARS-CoV and SARS-CoV-2 are mainly transmitted and entered through the respiratory tract, with acute severe pneumonia as the main symptom. However, compared with SARS-CoV, SARS-CoV-2 is reported to have a wider range of tissue infections, including the intestines, kidneys, and nervous system. In addition, SARS-CoV-2 has a higher transmissibility and a longer incubation period of infection, with a median incubation period of 5.4 days, 95% of symptomatic cases up to 13.7 days, and some asymptomatic cases even longer. SARS-CoV-2 is mainly transmitted by direct contact or respiratory droplets in a close and time-dependent manner, usually requiring close contact at close range for 15 minutes or more. However, the possibility of airborne transmission has also been confirmed in some cases, including prolonged exposure in closed or poorly ventilated spaces. Although SARS-CoV-2 has a lower lethality rate than SARS-CoV, its rapid and easy transmission, long incubation period, and absence of moderate symptoms make it very difficult to identify, track, and eliminate diseases caused by SARS-CoV-2 infection.

Figure 1. Sequence and structural summary of N protein. (Sources: Cubuk J, et al. 2021)

SARS-CoV NP protein contains two RNA binding domains, the N-terminal domain (NTD) and the C-terminal domain (CTD), between which there is a poorly structured connection (LKR). Due to the presence of positive amino acids, SARS-CoV N-NTD and N-CTD can bind to the viral RNA genome, and LKR can improve the ability to oligomerize. Nucleocapsid protein (NP) is highly expressed during viral infection and has a high content in mature viral particles. It also has good immunogenicity. During viral infection, NP protein enters the host cell together with viral RNA, promotes its replication, and participates in the assembly and release of viral particles. Serological diagnosis has found that specific antibodies against NP protein in the serum of SARS patients have higher sensitivity and longer persistence than other structural proteins of SARS-CoV. The N gene is more conservative and stable. For example, the amino acid homology of SARS-CoV-2 and SARS-CoV NP proteins is 90%, and there are fewer mutations during the long-term epidemic. Therefore, NP protein is often used in serological tests and is also an important target for vaccine design. The use of NP protein to prevent and treat infection has been demonstrated in chimpanzees with hepatitis B NP antigen to protect against hepatitis B challenge; human melanoma NP protein vaccine with CpG-ODN as adjuvant has good antitumor effect in tumor-bearing mice. NP protein is the most abundant protein in coronavirus and has a high level of protection. Although specific antibodies against NP protein have been detected in patient sera, its use in vaccination remains controversial. Some studies have shown that strong NP-specific humoral and cellular immune responses can be induced, while others have shown that NP protein contributes little to the production of neutralizing antibodies. Antibody responses to NP were generated in SARS-CoV patients, and its injection also induced T cell responses in BALB/C mice. However, immunization against NP protein did not provide protection against SARS-CoV, indicating that NP protein may be a peripheral target for improving T cell immunity. NP protein is also considered a promising vaccine material because it is highly immunogenic and expressed in large quantities in SARS-CoV-2. T cell responses to S, M, and NP subunit proteins have been shown to be the most significant and persistent in SARS-CoV-2, and their subunit vaccines have been extensively studied in clinical trials. The researchers analyzed the average antigenic tendency of the NP protein and selected the SARS-CoV-2 NP protein to design a multi-epitope vaccine; based on the advantages of NP protein as a SARS-CoV-2 vaccine antigen: sequence conservation, stability, strong immunogenicity, and low mutation during infection, NP protein can be used as a high-quality candidate vaccine for SARS-CoV-2. At present, there are not many reports on the SARS-CoV-2 NP protein, and it is necessary to continue to study SARS-CoV-2 NP.

N protein
Nucleoprotein
Nucleocapsid protein
Nucleocapsid (N) protein