Coronavirus nucleocapsid protein

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Nucleocapsid protein
Illustration of a SARS-CoV-2 virion
Model of the external structure of the SARS-CoV-2 virion.[1] The N protein, contained entirely within the virion, is not visible.
Blue: envelope
Turquoise: spike glycoprotein (S)
Red: envelope proteins (E)
Green: membrane proteins (M)
Orange: glycans

The nucleocapsid (N) protein is a protein that packages the positive-sense RNA genome of coronaviruses to form ribonucleoprotein structures enclosed within the viral capsid.[2][3] The N protein is the most highly expressed of the four major coronavirus structural proteins.[2] In addition to its interactions with RNA, N forms protein-protein interactions with the coronavirus membrane protein (M) during the process of viral assembly.[2][3] N also has additional functions in manipulating the cell cycle of the host cell.[3][4] The N protein is highly immunogenic and antibodies to N are found in patients recovered from SARS and Covid-19.[5]


X-ray crystallography structure of the dimer formed by two C-terminal domains from the SARS-CoV-2 N protein.[6]

The N protein is composed of two main protein domains connected by an intrinsically disordered region (IDR) known as the linker region, with additional disordered segments at each terminus.[2][3] A third small domain at the C-terminal tail appears to have an ordered alpha helical secondary structure and may be involved in the formation of higher-order oligomeric assemblies.[6] In SARS-CoV, the causative agent of SARS, the N protein is 422 amino acid residues long[2] and in SARS-CoV-2, the causative agent of Covid-19, it is 419 residues long.[6][7]

Both the N-terminal and C-terminal domains are capable of binding RNA. The C-terminal domain forms a dimer that is likely to be the native functional state.[2] Parts of the IDR, particularly a conserved sequence motif rich in serine and arginine residues (the SR-rich region), may also be implicated in dimer formation, though reports on this vary.[2][3] Although higher-order oligomers formed through the C-terminal domain have been observed crystallographically, it is unclear if these structures have a physiological role.[2][8]

The C-terminal dimer has been structurally characterized by X-ray crystallography for several coronaviruses and has a highly conserved structure.[6] The N-terminal domain - sometimes known as the RNA-binding domain, though other parts of the protein also interact with RNA - has also been crystallized and has been studied by nuclear magnetic resonance spectroscopy in the presence of RNA.[9]

Post-translational modifications

The N protein is post-translationally modified by phosphorylation at sites located in the IDR, particularly in the SR-rich region.[2][10] In several coronaviruses, ADP-ribosylation of the N protein has also been reported.[11][10] With unclear functional significance, the SARS-CoV N protein has been observed to be SUMOylated and the N proteins of several coronaviruses including SARS-CoV-2 have been observed to be proteolytically cleaved.[10][12][13]

Expression and localization

The N protein is the most highly expressed in host cells of the four major structural proteins.[2] Like the other structural proteins, the gene encoding the N protein is located toward the 3' end of the genome.[3]

N protein is localized primarily to the cytoplasm.[3] In many coronaviruses, a population of N protein is localized to the nucleolus,[3][4][14] thought to be associated with its effects on the cell cycle.[4]


Genome packaging and viral assembly

Coronavirus virion structure cross-section. The N proteins are represented by the small circles associated with the RNA genome in the virion interior.
NMR structure of the SARS-CoV-2 N protein N-terminal domain (red) in complex with double-stranded RNA (orange and yellow).[9]

The N protein binds to RNA to form ribonucleoprotein (RNP) structures for packaging the genome into the viral capsid.[2][3] The RNP particles formed are roughly spherical and are organized in flexible helical structures inside the virus.[2][3] Formation of RNPs is thought to involve allosteric interactions between RNA and multiple RNA-binding regions of the protein.[2][8] Dimerization of N is important for assembly of RNPs. Encapsidation of the genome occurs through interactions between N and M.[2][3] N is essential for viral assembly.[3] N also serves as a chaperone protein for the formation of RNA structure in the genomic RNA.[3][8]

Genomic and subgenomic RNA synthesis

Synthesis of genomic RNA appears to involve participation by the N protein. N is physically colocalized with the viral RNA-dependent RNA polymerase early in the replication cycle and forms interactions with non-structural protein 3, a component of the replicase-transcriptase complex.[3] Although N appears to facilitate efficient replication of genomic RNA, it is not required for RNA transcription in all coronaviruses.[3][15] In at least one coronavirus, transmissible gastroenteritis virus (TGEV), N is involved in template switching in the production of subgenomic mRNAs, a process that is a distinctive feature of viruses in the order Nidovirales.[3][15][16]

Cell cycle effects

Coronaviruses manipulate the cell cycle of the host cell through various mechanisms. In several coronaviruses, including SARS-CoV, the N protein has been reported to cause cell cycle arrest in S phase through interactions with cyclin-CDK.[3][4] In SARS-CoV, a cyclin box-binding region in the N protein can serve as a cyclin-CDK phosphorylation substrate.[3] Trafficking of N to the nucleolus may also play a role in cell cycle effects.[4] More broadly, N may be involved in reduction of host cell protein translation activity.[3]

Immune system effects

The N protein is involved in viral pathogenesis via its effects on components of the immune system. In SARS-CoV,[3][17][18] MERS-CoV,[19] and SARS-CoV-2,[20] N has been reported as suppressing interferon responses.

Evolution and conservation

The structures of N proteins from different coronaviruses, particularly the C-terminal domains, appear to be well conserved.[2][6] Similarities between the structure and topology of the N proteins of coronaviruses and arteriviruses suggest a common evolutionary origin and supports the classification of these two groups in the common order Nidovirales.[2][3]

Examination of SARS-CoV-2 sequences collected during the Covid-19 pandemic found that missense mutations were most common in the central linker region of the protein, suggesting this relatively unstructured region is more tolerant of mutations than the structured domains.[6] A separate study of SARS-CoV-2 sequences identified at least one site in the N protein under positive selection.[21]


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