The nucleocapsid protein of SARS-CoV-2: potential target for a vaccine

Posted by Bruno Tillier on 17-Nov-2020 09:12:50
Bruno Tillier

 

What is the N-Protein ?

The nucleocapsid protein (N protein) of SARS-CoV-2 is a structural protein synthesized by the virus, which plays a key role in the RNA packaging into the nucleocapsid. Its function is to mediate viral assembly through interaction with the viral genome and M protein, helping to increase viral RNA transcription and replication. Characterization studies have shown that N protein is very immunogenic and the antibodies against N protein present higher sensitivity and longer persistence than those against other structural proteins of SARS-CoV. These findings allow to consider N protein as a potential target for vaccine development against SARS-CoV-2.

 

Coronaviruses and Diseases

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiological agent of the current coronavirus disease 2019 (COVID-19). It belongs to the family Coronaviridae and shares the features of other coronaviruses, namely it is an enveloped virus, with a positive single-stranded RNA genome and it is pathogenic [1]. Like SARS-CoV (severe acute respiratory syndrome coronavirus) and MERS-CoV (Middle-East respiratory syndrome coronavirus), SARS-Cov-2 is transmitted by zoonotic transmission and spreads among humans through close contact. However, SARS-CoV-2 has shown to be more pathogenic than SARS-CoV and MERS-CoV. The SARS-CoV-2 genome has around 82% sequence similarity with SARS-CoV and MERS-CoV genomes [2].

 

SARS-CoV-2 proteins

The SARS-CoV-2 virion consists of a nucleocapsid core enclosed by an envelope which contains three membrane proteins, namely spike (S), envelope (E) and membrane (M) [2]. The genome, with a size of around 30 kb, codifies for nonstructural replicase polyproteins and for the referred structural proteins [3]. The S protein is responsible for binding to the host cell-surface ACE2 receptor, initiating the infection [4,5]. After, the virus genome attaches to the host cell ribosomes, with the resulting translation of polyproteins, subsequently processed by proteolysis [2]. The virus lifecycle ends with the folding and packing of new virions, which infect other cells. E proteins are a group of small viral proteins that participate in the assembly and release of the virions [6] and are considered potential drug targets [2]. M proteins play a key role in the RNA packaging into the nucleocapsid [7], as well as N proteins [8].

 

The potential drug/vaccine targets

The S, E, M and N proteins, along with two isoforms of replicase polyprotein, namely 1a and 1ab, are considered potential drug/vaccine targets [2], and the S protein is already being used in several studies as a target antigen in vaccine development [9,10]. Nonetheless, the S protein undergoes conformational changes induced after its penetration in the host cell [11], and these changes seem to be provoked by mutations, which cause alterations in the antigenicity of this target. These dynamic changes in the protein may affect immune responses [12] and also may lead to drug resistance, if antiviral drugs are to be developed using S protein as target [13].

Hence, other proteins are being considered as targets for vaccine/drug development against SARS-CoV-2, and the N protein seems promising, as it is not affected by the variability of S protein. In fact, it was observed that N proteins from BAT-CoV, SARS-CoV and SARS-CoV-2 present highly conserved regions (N protein of SARS-CoV-2 shares nearly 90% sequence identity with that of SARS-CoV), so it is assumed that antibodies against the N protein of SARS-CoV would likely recognize the N protein of SARS-CoV-2 [2].

 

N-Protein specificities

N protein is a highly basic protein, containing two well-folded domains, both of them rich in β-strands. It presents highly positively charged regions which may facilitate binding to non-specific nucleic acids [14]. The function of N protein is to mediate viral assembly through interaction with the viral genome and M protein, helping to increase viral RNA transcription and replication [15]. Compared to the other virus proteins, N protein is more stable and fewer mutations occur over time [16].

IgG, IgA and IgM antibodies against the N protein were detected in confirmed COVID-19 patients’ sera [14,17] and were highly expressed during infection [15]. In addition, N protein was found to be very immunogenic, and the antibodies against N protein presented higher sensitivity and longer persistence than those against other structural proteins of SARS-CoV [18,19], eliciting proliferation and cytotoxic activity of SARS-specific T-cells [20,21].

Due to its multiple functions in viral lifecycle and infection, and the immunological response that it stimulates, N protein from SARS-CoV-2 is now a potential candidate as a target for a SARS-CoV-2 vaccine or drug.

 

Synthelis

Considering the potential of the N-protein from SARS-CoV-2 to fight COVID-19, Synthelis developed a protocol to produce this target in less than 2 weeks by using its cell-free protein expression system. After validation on rapid test devices from the market, this product is now available in Synthelis’ catalog.

As a service provider, we can also develop and produce almost any kind of proteins you may request for your research activity. Please contact us if you want additional information: 

Request a call back

References

[1] Xu Z, Shi L, Wang Y, Zhang J, Huang L, Zhang C, Liu S, Zhao P, Liu H, Zhu L. Pathological findings of COVID-19 associated with acute respiratory distress syndrome. Lancet Respir. Med. 8 (2020) 420-422. https://doi.org/10.1016/S2213-2600(20)30076-X.

[2] Naqvi AAT, Fatima K, Mohammad T, Fatima U, Singh IK, Singh A, Atif SM, Hariprasad G, Hasan GM, Hassan MI. Insights into SARS-CoV-2 genome, structure, evolution, pathogenesis and therapies: Structural genomics approach. BBA Mol. Basis Dis. 1866 (2020) 165878. https://doi.org/ 10.1016/j.bbadis.2020.165878.

[3] Zhou P, Yang XL, Wang XG, Hu B, Zhang L, Zhang W, Si HR, Zhu Y, Li B, Huang CL, Chen HD, Chen J, Luo Y, Guo H, Jiang RD, Liu MQ, Chen Y, Shen XR, Wang X, Zheng XS, Zhao K, Chen QJ, Deng F, Liu LL, Yan B, Zhan FX, Wang YY, Xiao GF, Shi ZL. A pneumonia outbreak associated with a new coronavirus of probable bat origin. Nature 579 (2020) 270-273. https://doi.org/10.1038/s41586-020-2012-7.

[4] Lau YL, Peiris JM. Pathogenesis of severe acute respiratory syndrome. Curr. Opin. Immunol. 17 (2005) 404-410. https://doi.org/10.1016/j.coi.2005.05.009.

[5] Zhang H, Penninger JM, Li Y, Zhong N, Slutsky AS. Angiotensin-converting enzyme 2 (ACE2) as a SARS-CoV-2 receptor: molecular mechanisms and potential therapeutic target. Intensive Care Med. 46 (2020) 586-590. https://doi.org/10.1007/s00134-020-05985-9.

[6] Fehr AR, Perlman S. Coronaviruses: an overview of their replication and pathogenesis. Methods Mol. Biol. 1282 (2015) 1-23. https://doi.org/10.1007/978-1-4939-2438-7_1.

[7] Tang T, Bidon M, Jaimes JA, Whittaker GR, Daniel S. Coronavirus membrane fusion mechanism offers as a potential target for antiviral development. Antiviral Res. 178 (2020) 104792. https://doi.org/10.1016/j.antiviral.2020.104792.

[8] Neuman BW, Buchmeier MJ. Supramolecular architecture of the coronavirus particle. Adv. Virus Res. 96 (2016) 1-27. https://doi.org/10.1016/bs.aivir.2016.08.005.

[9] Pang J, Wang MX, Ang IYH, Tan SHX, Lewis RF, Chen JI, Gutierrez RA, Gwee SXW, Chua PEY, Yang Q, Ng XY, Yap RK, Tan HY, Teo YY, Tan CC, Cook AR, Yap JC, Hsu LY. Potential rapid diagnostics, vaccine and therapeutics for 2019 novel coronavirus (2019-nCoV): a systematic review. J. Clin. Med. 9 (2020) 623. https://doi.org/10.3390/jcm9030623.

[10] Chen WH, Strych U, Hotez PJ, Bottazzi ME. The SARS-CoV-2 vaccine pipeline: an overview. Curr. Trop. Med. Rep. 7 (2020) 61-64. https://doi.org/10.1007/s40475-020-00201-6.

[11] Wrapp D, Wang N, Corbett KS, Goldsmith JA, Hsieh CL, Abiona O, Graham BS, McLellan JS. Cryo-EM structure of the 2019-nCoV spike in the prefusion conformation. Science 367 (2020) 1260-1263. https://doi.org/10.1126/science.abb2507.

[12] Walls AC, Park YJ, Tortorici MA, Wall A, McGuire AT, Veesler D. Structure, function, and antigenicity of the SARS-CoV-2 spike glycoprotein. Cell 181 (2020) 281-292. https://doi.org/ 10.1016/j.cell.2020.02.058.

[13] Wang XW, Li JS, Jin M, Zhen B, Kong QX, Song N, Xiao WJ, Yin J, Wei W, Wang GJ, Si BY, Guo BZ, Liu C, Ou GR, Wang MN, Fang TY, Chao FH, Li JW. Study on the resistance of severe acute respiratory syndrome-associated coronavirus. J. Virol. Methods 126 (2005) 171-177. https://doi.org/ 10.1016/j.jviromet.2005.02.005.

[14] Zeng W, Liu G, Ma H, Zhao D, Yang Y, Liu M, Mohammed A, Zhao C, Yang Y, Xie J, Ding C, Ma X, Weng J, Gao Y, He H, Jin T. Biochemical characterization of SARS-CoV-2 nucleocapsid protein. Biochem. Biophys. Res. Commun. 527 (2020) 618-623. https://doi.org/ 10.1016/j.bbrc.2020.04.136.

[15] Cong Y, Ulasli M, Schepers H, Mauthe M, V’kovski P, Kriegenburg F, Thiel V, de Haan CA, Reggiori F. Nucleocapsid Protein recruitment to replication-transcription complexes plays a crucial role in coronaviral life cycle. J. Virol. 94 (2020) e01925-19. https://doi.org/10.1128/JVI.01925-19.

[16] Grifoni A, Sidney J, Zhang Y, Scheuermann RH, Peters B, Sette A. A sequence homology and bioinformatic approach can predict candidate targets for immune responses to SARS-CoV-2. Cell Host Microbe 27 (2020) 671-680. https://doi.org/10.1016/j.chom.2020.03.002.

[17] Leung DT, Tam FC, Ma CH, Chan PK, Cheung JL, Niu H, Tam JS, Lim PL. Antibody response of patients with severe acute respiratory syndrome (SARS) targets the viral nucleocapsid. J. Infect. Dis. 190 (2004) 379-386. https://doi.org/10.1086/422040.

[18] Tan Y, Goh P, Fielding BC, Shen S, Chou C, Fu J, Leong HN, Leo YS, Ooi EE, Ling AE. Profiles of antibody responses against severe acute respiratory syndrome coronavirus recombinant proteins and their potential use as diagnostic markers. Clin. Diagn. Lab. Immunol. 11 (2004) 362-371. https://doi.org/10.1128/cdli.11.2.362-371.2004.

[19] Shi Y, Yi Y, Li P, Kuang T, Li L, Dong M, Ma Q, Cao C. Diagnosis of severe acute respiratory syndrome (SARS) by detection of SARS coronavirus nucleocapsid antibodies in an antigen-capturing enzyme-linked immunosorbent assay. J. Clin. Microbiol. 41 (2003) 5781-5782. https://doi.org/10.1128/jcm.41.12.5781-5782.2003.

[20] Gao W, Tamin A, Soloff A, D’Aiuto L, Nwanegbo E, Robbins PD, Bellini WJ, Barratt-Boyes S, Gambotto A. Effects of a SARS-associated coronavirus vaccine in monkeys. Lancet 362 (2003) 1895-1896. https://doi.org/10.1016/S0140-6736(03)14962-8.

[21] Okada M, Takemoto Y, Okuno Y, Hashimoto S, Yoshida S, Fukunaga Y, Tanaka T, Kita Y, Kuwayama S, Muraki Y, Kanamaru N, Takai H, Okada C, Sakaguchi Y, Furukawa I, Yamada K, Matsumoto M, Kase T, Demello DE, Peiris JS, Chen PJ, Yamamoto N, Yoshinaka Y, Nomura T, Ishida I, Morikawa S, Tashiro M, Sakatani M. The development of vaccines against SARS corona virus in mice and SCID-PBL/hu mice. Vaccine 23 (2005) 2269-2272. https://doi.org/10.1016/j.vaccine.2005.01.036.

 

mythbustings about cell free technology

Latest articles

Subscribe to our blog