Analysis of the SARS-CoV-2 envelope (E), nucleocapsid (N), and non-structural protein12 (nsp12) genes from COVID-19 patients in West Java

Azzania Fibriani(1*), Irin Annisa Evitayani(2), Gusti Ayu Prani Pradani(3), Rebecca Stephanie(4), Ema Rahmawati(5), Ryan Bayusantika Ristandi(6), Cut Nur Cinthia Alamanda(7), Rifky Waluyajati Rachman(8), Rini Robiani(9), Isak Solihin(10)

(1) Department of Biology, School of Life Sciences and Technology, Bandung Institute of Technology, Bandung 40132, Indonesia
(2) Department of Biology, School of Life Sciences and Technology, Bandung Institute of Technology, Bandung 40132, Indonesia
(3) Department of Microbiology, School of Life Sciences and Technology, Bandung Institute of Technology, Bandung 40132, Indonesia
(4) Department of Microbiology, School of Life Sciences and Technology, Bandung Institute of Technology, Bandung 40132, Indonesia
(5) UPTD Laboratorium Kesehatan Daerah Jawa Barat, Bandung 40161, Indonesia
(6) UPTD Laboratorium Kesehatan Daerah Jawa Barat, Bandung 40161, Indonesia
(7) UPTD Laboratorium Kesehatan Daerah Jawa Barat, Bandung 40161, Indonesia
(8) UPTD Laboratorium Kesehatan Daerah Jawa Barat, Bandung 40161, Indonesia
(9) UPTD Laboratorium Kesehatan Daerah Jawa Barat, Bandung 40161, Indonesia
(10) UPTD Laboratorium Kesehatan Daerah Jawa Barat, Bandung 40161, Indonesia
(*) Corresponding Author

Abstract


According to World Health Organization, as of January 2021, Indonesia is the only Southeast Asian country in which COVID-19 is still occurring in community transmission. West Java is one of the provinces holding the highest positive cases number. With the envelope (E), nucleocapsid (N), and non-structural protein 12 (nsp12) being the target genes of SARS-CoV-2 diagnostic kits and several antiviral drugs, the study of genetic variations has become relevant and greatly important. Out of 267 oro-nasopharyngeal swab specimens that were previously confirmed positive for COVID-19 in qPCR diagnostic test in Laboratorium Kesehatan Provinsi Jawa Barat, ten samples with acceptable qualities were selected and three samples were sequenced using Sanger sequencing. Nonsynonymous mutations were observed in the envelope gene (L21F) and in the nucleocapsid genes (R203K, G204R, A211S, and S193I). Phylogenetic analysis showed that samples were clustered with other sequences carrying identical mutations, but clustered non-discriminatively with all sequences when carrying no mutation. No pattern in geographical areas and clades, except for R203K-G204R for being a marker for the GR clade. Protein structure analysis showed that mutations observed did not change the hydrophobicity and the secondary structure of the nucleocapsid, while stability change (ΔΔG) showed that all mutations, aside from the R203K-G204R, have neutral effect on the protein stability. Therefore, it can be concluded that mutations observed in this experiment did not impart preference to disperse in certain geographical areas or cause any significant structural change in the protein.

Keywords


envelope; nsp12; nucleocapsid; SARS-CoV-2; Sanger sequencing

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References


Azad GK. 2021. The molecular assessment of SARS-CoV-2 Nucleocapsid Phosphoprotein variants among Indian isolates. Heliyon. 7, e06167. DOI: 10.1016/j.heliyon.2021.e06167.

Cortey M et al. 2020. N SARS-CoV-2 amino acid substitutions widely spread in the human population are mainly located in highly conserved segments of the structural proteins. o Title. bioRxiv. DOI: https://doi.org/10.1101/2020.05.16.099499.

Dilucca M, Forcelloni S, Georgakilas AG, Giansanti A, Pavlopoulou A. 2020. Codon Usage and Phenotypic Divergences of SARS-CoV-2 Genes. Viruses. 12, 498. DOI: 10.3390/v12050498.

Garvin MR et al. 2020. Potentially adaptive SARS-CoV-2 mutations discovered with novel spatiotemporal and explainable AI models. Genome Biol. 21, 1–26. DOI: 10.1186/s13059-020-02191-0.

George RA, Heringa J. 2002. An analysis of protein domain linkers: Their classification and ole in protein folding. Protein Eng. DOI: 10.1093/protein/15.11.871.

Internal Balai Laboratorium Kesehatan Provinsi Jawa Barat. 2020. Analisis Genom SARS- CoV-2 Indonesia. Kilic T, Weissleder R, Lee H. 2020. Molecular and Immunological Diagnostic Tests of COVID-19: Current Status and Challenges. iScience. 23, 101406. DOI: 10.1016/j.isci.2020.101406.

Koyama T, Platt D, Parida L. 2020. Variant analysis of SARS-cov-2 genomes. Bull. World Health Organ. 98, 495–504. DOI: 10.2471/BLT.20.253591.

Kumar A et al. 2020. Characterization of Nucleocapsid (N) Protein from Novel Coronavirus SARS-CoV-2. Preprints. DOI: 10.20944/preprints202005.0413.v1.

Liu DX, Liang JQ, Fung TS. 2020. Human Coronavirus-229E, -OC43, -NL63, and -HKU1. In: Reference Module in Life Sciences. Elsevier. DOI: 10.1016/b978-0-12-809633- 8.21501-x.

Maitra A et al. 2020. Mutations in SARS-CoV-2 viral RNA identified in Eastern India: Possible implications for the ongoing outbreak in India and impact on viral structure and host susceptibility. J. Biosci. 45. DOI: 10.1007/s12038-020-00046-1.

Mercatelli D, Giorgi FM. 2020. Geographic and Genomic Distribution of SARS-CoV-2 Mutations. Front. Microbiol. 11, 1800. DOI: 10.3389/fmicb.2020.01800.

Nieto-Torres JL et al. 2014. Severe Acute Respiratory Syndrome Coronavirus Envelope Protein Ion Channel Activity Promotes Virus Fitness and Pathogenesis Denison, MR, editor. PLoS Pathog. 10, e1004077. DOI: 10.1371/journal.ppat.1004077.

Pachetti M et al. 2020. Emerging SARS-CoV-2 mutation hot spots include a novel RNA- dependent-RNA polymerase variant. J. Transl. Med. 18, 179. DOI: 10.1186/s12967- 020-02344-6.

Rahman MS et al. 2020. Evolutionary dynamics of SARS‐CoV‐2 nucleocapsid (N) protein and its consequences. J. Med. Virol. DOI: 10.1002/jmv.26626.

Robson F et al. 2020. Coronavirus RNA Proofreading: Molecular Basis and Therapeutic Targeting. Mol. Cell. 79, 710–727. DOI: https://doi.org/10.1016/j.molcel.2020.07.027.

Schoeman D, Fielding BC. 2019. Coronavirus envelope protein: Current knowledge. Virol. J. 16, 1–22. DOI: 10.1186/s12985-019-1182-0.

Tung HYL, Limtung P. 2020. Mutations in the phosphorylation sites of SARS-CoV-2 encoded nucleocapsid protein and structure model of sequestration by protein 14-3-3. Biochem. Biophys. Res. Commun. 532, 134–138. DOI: 10.1016/j.bbrc.2020.08.024.

Wang S, Ma J, Xu J. 2016. AUCpreD: proteome-level protein disorder prediction by AUC- maximized deep convolutional neural fields. Bioinformatics. 32, i672–i679. DOI: 10.1093/bioinformatics/btw446.

World Health Organization. 2021. WHO Coronavirus Disease (COVID-19) Dashboard. https://covid19.who.int/.

Zeng W et al. 2020. Biochemical characterization of SARS-CoV-2 nucleocapsid protein. Biochem. Biophys. Res. Commun. 527, 618–623. DOI: 10.1016/j.bbrc.2020.04.136.




DOI: https://doi.org/10.37604/jmsb.v3i1.66

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