In silico comparative analysis of SARS-CoV-2 Nucleocapsid (N) protein using bioinformatics tools

Year 2021, , 1 - 9, 30.04.2021

Mehmet Emin Uras

https://doi.org/10.51753/flsrt.843166

Cited By: 4

Abstract

The world has been encountered to one of the biggest pandemics that causing by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). SARS-CoV-2 is placed in the Beta-CoV genus in the Coronaviridae family. N protein is one of the crucial structural proteins of SARS-CoV-2 that binds to the genome thereby generating helical ribonucleoprotein core. It is involved in viral transcription/replication, translation, and viral assembly after entering the host cell through interacting with host proteins. N protein sequences of SARS-CoV-2 and taxonomically related CoVs are examined using bioinformatics tools and approaches including sequence alignment, sequence and phylogenetic analyzes, and predicting of putative N-Glycosylation and phosphorylation positions and also predictions and comparative analyzes are performed on 3D structures of N proteins from SARS-CoV-2 related CoVs through using of some of applied bioinformatics analyzes. Results of mega BLAST search revealed that the most similar N protein sequence to SARS-CoV-2 is Bat-CoV RaTG13 N protein sequence in the taxonomically related CoVs. SARS-CoV-2 is grouped with SARS, pangolin, civet and bat CoVs (RATG13, SL ZC45 and SL ZXC21) in N protein, nucleotide and protein based ML phylogenetic trees. Some of SARS-CoV-2 N proteins were showed divergence from other SARS-CoV-2 N proteins analyzed due to amino acid substitutions detected in SARS-CoV-2 N proteins samples in phylogenetic trees. The highest amino acid substitutions were detected in Richmont/USA (QJA42209.1) and Greece (QIZ16579.1) samples, with 2 and 3 place substitutions, respectively. By domain analyzes, three domains were detected as Corona_nucleocora (Pfam), N terminal CoV RNA-binding domain (HAMAP) and C terminal N protein dimerization domain (HAMAP). Possible N-glycosylation positions of SARS-CoV-2 N protein were predicted at two positions. Assessments of possible serine, threonine and tyrosine phosphorylations were found to be at 100 positions, 34 of them were higher than 80% possibility. 3D structure analysis based on TM scores revealed that although the results of 3D structure analysis were shown consistency with the taxonomy of the CoVs, the 3D structures of SARS-CoV-2 N protein and taxonomically related CoVs were not at the same fold.

Keywords

3D structure, Bioinformatics, Coronavirus, COVID-19, SARS-CoV-2, Viral Proteins

References

• Blom, N., Sicheritz-Pontén, T., Gupta, R., Gammeltoft, S., & Brunak, S. (2004). Prediction of post-translational glycosylation and phosphorylation of proteins from the amino acid sequence. Proteomics, 4(6), 1633-1649. https://doi.org/10.1002/pmic.200300771
• Cagliani, R., Forni, D., Clerici, M., & Sironi, M. (2020). Computational inference of selection underlying the evolution of the novel coronavirus, SARS-CoV-2. Journal of Virology, 94(12), 1-11. https://doi.org/10.1128/jvi.00411-20
• Carlson, C. R., Asfaha, J. B., Ghent, C. M., Howard, C. J., Hartooni, N., Safari, M., ... & Morgan, D. O. (2020). Phosphoregulation of phase separation by the SARS-CoV-2 N protein suggests a biophysical basis for its dual functions. Molecular Cell, 80(6), 1092-1103.
• Ceraolo, C., & Giorgi, F. M. (2020). Genomic variance of the 2019‐nCoV coronavirus. Journal of Medical Virology, 92(5), 522-528. https://doi.org/10.1002/jmv.25700
• Chang, C. K., Sue, S. C., Yu, T. H., Hsieh, C. M., Tsai, C. K., Chiang, Y. C., … Huang, T. H. (2006). Modular organization of SARS coronavirus nucleocapsid protein. Journal of Biomedical Science, 13(1), 59–72. https://doi.org/10.1007/s11373-005-9035-9
• Chen, Y., Liu, Q., & Guo, D. (2020). Emerging coronaviruses: Genome structure, replication, and pathogenesis. Journal of Medical Virology, 92(4), 418-423. https://doi.org/10.1002/jmv.25681
• Cui, J., Li, F., & Shi, Z. L. (2019). Origin and evolution of pathogenic coronaviruses. Nature Reviews Microbiology, 17(3), 181-192. https://doi.org/10.1038/s41579-018-0118-9
• De Wit, E., Van Doremalen, N., Falzarano, D., & Munster, V. J. (2016). SARS and MERS: Recent insights into emerging coronaviruses. Nature Reviews Microbiology, 14(8), 523-534. https://doi.org/10.1038/ nrmicro.2016.81
• El-Gebali, S., Mistry, J., Bateman, A., Eddy, S. R., Luciani, A., Potter, S. C., … Finn, R. D. (2019). The Pfam protein families database in 2019. Nucleic Acids Research, 47(D1), D427-D432. https://doi.org/10.1093/nar/gky995
• Fang, X., Ye, L.-B., Zhang, Y., Li, B., Li, S., Kong, L., … Wu, Z. (2006). Nucleocapsid amino acids 211 to 254, in particular, tetrad glutamines, are essential for the interaction between the nucleocapsid and membrane proteins of SARS-associated coronavirus. Journal of Microbiology, 44(5), 577-580.
• Felsenstein, J. (1985). Confidence limits on phylogenies: an approach using the bootstrap. Evolution, 39(4), 783-791. https://doi.org/10.2307/ 2408678
• Fung, T. S., & Liu, D. X. (2018). Post-translational modifications of coronavirus proteins: Roles and function. Future Virology, 13(6), 405-430. https://doi.org/10.2217/fvl-2018-0008
• Genc, B. N. (2020). Critical management of COVID-19 pandemic in Turkey. Frontiers in Life Sciences and Related Technologies, 1(2), 69-73.
• Gupta, R., Jung, E., & Brunak, S. (2004). NetNGlyc: Prediction of N-glycosylation sites in human proteins.
• Hall, T. A. (1999). BIOEDIT: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/ NT. Nucleic Acids Symposium Series, 41, 95-98.
• He, R., Leeson, A., Ballantine, M., Andonov, A., Baker, L., Dobie, F., … Li, X. (2004). Characterization of protein-protein interactions between the nucleocapsid protein and membrane protein of the SARS coronavirus. Virus Research, 105(2), 121-125. https://doi.org/10.1016/j.virusres.2004.05.002
• Hogue, B. G., & Machamer, C. E. (2008). Coronavirus structural proteins and virus assembly. In: Perlman, S., Gallagher, T., Snijder, E. J. (eds) Nidoviruses (pp. 179-200). ASM Press, Washington, DC. https://doi.org/10.1128/9781555815790.ch12
• Huang, S. Y., Shi, S. P., Qiu, J. D., & Liu, M. C. (2015). Using support vector machines to identify protein phosphorylation sites in viruses. Journal of Molecular Graphics and Modelling, 56, 84-90. https://doi.org/10.1016/j.jmgm.2014.12.005
• Kelley, L. A., Mezulis, S., Yates, C. M., Wass, M. N., & Sternberg, M. J. E. (2015). The Phyre2 web portal for protein modeling, prediction and analysis. Nature Protocols, 10(6), 845-858. https://doi.org/10.1038/nprot.2015.053
• Kumar, S., Stecher, G., Li, M., Knyaz, C., & Tamura, K. (2018). MEGA X: Molecular evolutionary genetics analysis across computing platforms. Molecular Biology and Evolution, 35(6), 1547-1549. https://doi.org/10.1093/molbev/msy096
• Li, S., Lin, L., Wang, H., Yin, J., Ren, Y., Zhao, Z., … Liu, S. (2003). The epitope study on the SARS-CoV nucleocapsid protein. Genomics, Proteomics & Bioinformatics / Beijing Genomics Institute, 1(3), 198-206. https://doi.org/10.1016/S1672-0229(03)01025-8
• Mossenta, M., Marchese, S., Poggianella, M., Slon Campos, J. L., & Burrone, O. R. (2017). Role of N-glycosylation on Zika virus E protein secretion, viral assembly and infectivity. Biochemical and Biophysical Research Communications, 492(4), 579-586. https://doi.org/10.1016/j.bbrc.2017.01.022
• Neuman, B. W., Kiss, G., Kunding, A. H., Bhella, D., Baksh, M. F., Connelly, S., … Buchmeier, M. J. (2011). A structural analysis of M protein in coronavirus assembly and morphology. Journal of Structural Biology, 174(1), 11-22. https://doi.org/10.1016/j.jsb.2010.11.021
• Ng, P. C., & Henikoff, S. (2006). Predicting the Effects of Amino Acid Substitutions on Protein Function. Annual Review of Genomics and Human Genetics, 7(1), 61-80. https://doi.org/10.1146/annurev.genom.7.080505.115630
• Nieto-Torres, J. L., DeDiego, M. L., Álvarez, E., Jiménez-Guardeño, J. M., Regla-Nava, J. A., Llorente, M., … Enjuanes, L. (2011). Subcellular location and topology of severe acute respiratory syndrome coronavirus envelope protein. Virology, 415(2), 69-82. https://doi.org/10.1016/j.virol.2011.03.029
• Pedruzzi, I., Rivoire, C., Auchincloss, A. H., Coudert, E., Keller, G., De Castro, E., … Bridge, A. (2015). HAMAP in 2015: updates to the protein family classification and annotation system. Nucleic Acids Research, 43(D1), D1064-D1070. https://doi.org/10.1093/nar/gku1002
• Perera, K. D., Rathnayake, A., Liu, H., Pedersen, N. C., Groutas, W. C., Chang, K. O., & Kim, Y. (2019). Characterization of amino acid substitutions in feline coronavirus 3C-like protease from a cat with feline infectious peritonitis treated with a protease inhibitor. Veterinary Microbiology, 237, 108398. https://doi.org/10.1016/j.vetmic.2019.108398
• Satija, N., & Lal, S. K. (2007). The molecular biology of SARS coronavirus. Annals of the New York Academy of Sciences, 1102(1), 26-38. https://doi.org/10.1196/annals.1408.002
• Schrauwen, E. J. A., Richard, M., Burke, D. F., Rimmelzwaan, G. F., Herfst, S., & Fouchier, R. A. M. (2016). Amino acid substitutions that affect receptor binding and stability of the hemagglutinin of influenza A/H7N9 Virus. Journal of Virology, 90(7), 3794-3799. https://doi.org/10.1128/jvi.03052-15
• Supekar, N. T., Shajahan, A., Gleinich, A., Rouhani, D., Heiss, C., & Azadi, P. (2020). SARS-CoV-2 Nucleocapsid protein is decorated with multiple N-and O-glycans. BioRxiv, 1-32. https://doi.org/10.1101/2020.08.26.269043
• Tajima, F. (1989). Statistical method for testing the neutral mutation hypothesis by DNA polymorphism. Genetics, 123(3), 585-595.
• Teng, S., Srivastava, A. K., Schwartz, C. E., Alexov, E., & Wang, L. (2010). Structural assessment of the effects of amino acid substitutions on protein stability and protein-protein interaction. International Journal of Computational Biology and Drug Design, 3(4), 334-349. https://doi.org/10.1504/IJCBDD.2010.038396
• Thompson, J. D., Higgins, D. G., & Gibson, T. J. (1994). CLUSTAL W: Improving the sensitivity of progressive multiple sequence alignment through sequence weighting, position-specific gap penalties and weight matrix choice. Nucleic Acids Research, 22(22), 4673-4680. https://doi.org/10.1093/nar/22.22.4673
• Tilocca, B., Soggiu, A., Sanguinetti, M., Musella, V., Britti, D., Bonizzi, L., … Roncada, P. (2020). Comparative computational analysis of SARS-CoV-2 nucleocapsid protein epitopes in taxonomically related coronaviruses. Microbes and Infection, 22(4-5), 188-194. https://doi.org/10.1016/j.micinf.2020.04.002
• WHO. (2020a). Coronavirus Disease (COVID-19) Situation Report - 104. Retrieved from https://www.who.int/docs/default-source/coronaviruse/situation-reports/20200503-covid-19-sitrep-104.pdf?sfvrsn=53328f46_2
• WHO. (2020b). Novel Coronavirus (2019-nCoV) Situation Report 1. Retrieved from https://www.who.int/docs/default-source/coronaviruse/situation-reports/20200121-sitrep-1-2019-ncov.pdf?sfvrsn=20a99c10_4
• WHO. (2020c). WHO Coronavirus Disease (COVID-19) Dashboard. Retrieved from https://covid19.who.int/?gclid=Cj0KCQiA_qD_BRDiARIsANjZ2LBM_HykD99cBBzOqxepXsh8SGA4ZONQUk8baXeAXC2B8_DKBEzGaWQaAh2CEALw_wcB
• Williams, C. J., Headd, J. J., Moriarty, N. W., Prisant, M. G., Videau, L. L., Deis, L. N., … Richardson, D. C. (2018). MolProbity: More and better reference data for improved all-atom structure validation. Protein Science, 27(1), 293-315. https://doi.org/10.1002/pro.3330
• Wu, A., Peng, Y., Huang, B., Ding, X., Wang, X., Niu, P., … Jiang, T. (2020). Genome composition and divergence of the novel coronavirus (2019-nCoV) originating in China. Cell Host and Microbe, 27(3), 325-328. https://doi.org/10.1016/j.chom.2020.02.001
• Wu, F., Zhao, S., Yu, B., Chen, Y. M., Wang, W., Song, Z. G., … Zhang, Y. Z. (2020). A new coronavirus associated with human respiratory disease in China. Nature, 579(7798), 265-269. https://doi.org/10.1038/s41586-020-2008-3
• Ye, Z. W., Yuan, S., Yuen, K. S., Fung, S. Y., Chan, C. P., & Jin, D. Y. (2020). Zoonotic origins of human coronaviruses. International Journal of Biological Sciences, 16(10), 1686-1697. https://doi.org/10.7150/ijbs.45472
• Zhang, T., Wu, Q., & Zhang, Z. (2020). Probable pangolin origin of SARS-CoV-2 associated with the COVID-19 outbreak. Current Biology, 30(7), 1346-1351. https://doi.org/10.1016/j.cub.2020.03.022
• Zhang, Y., & Skolnick, J. (2004). Scoring function for automated assessment of protein structure template quality. Proteins: Structure, Function, and Bioinformatics, 57(4), 702-710. https://doi.org/10.1002/prot.20264
• Zuckerksndl, E., & Pauling, L. (1965). Evolutionary divergence and convergence in proteins. In: Bryson, V., Vogel, H. J. (eds) Evolving Genes and Proteins (pp. 97-166). Academic Press. https://doi.org/10.1016/b978-1-4832-2734-4.50017-6.