key: cord-0864536-gqn1lj6g authors: Hussain, Mushtaq; Shabbir, Sanya; Amanullah, Anusha; Raza, Fozia; Imdad, Muhammad J.; Zahid, Sahar title: Immunoinformatic analysis of structural and epitope variations in the spike and Orf8 proteins of SARS‐CoV‐2/B.1.1.7 date: 2021-03-25 journal: J Med Virol DOI: 10.1002/jmv.26931 sha: b66d80331e85e94709e3260ce03fe80faf9752e5 doc_id: 864536 cord_uid: gqn1lj6g A newly emerged strain of SARS‐CoV‐2 of B.1.1.7 lineage has caused a significant surge in the SARS‐CoV‐2 infections in the UK. In this study, changes in the epitopes of spike and orf8 proteins in SARS‐CoV‐2 of B.1.1.7 lineage were investigated. Genomic alignment of the SARS‐CoV‐2/B.1.1.7 with SARS‐CoV‐2/Wuhan showed the presence of several mutations in orf1a/b, spike, orf8, and N proteins of SARS‐CoV‐2/B.1.1.7. Molecular models of spike and orf8 proteins were constructed by homology modeling. Superimposition between the spike proteins of SARS‐CoV‐2/Wuhan and SARS‐CoV‐2/B.1.1.7 showed noticeable variations in the spatial orientation in Val70‐Asn74 and Thr250‐Ser255 regions. This may have also resulted in the extension of the epitopic region at Ser244‐Gly249 in the SARS‐CoV‐2/B.1.1.7 spike protein. Superimposition of the SARS‐CoV‐2/B.1.1.7 spike protein over Fab‐spike protein complexes of SARS‐CoV‐2/Wuhan also showed subtle variations in the antibody binding affinity targeting the N‐terminal domain of the spike protein. Epitopic variations were also observed between the corresponding orf8 regions of SARS‐CoV‐2/Wuhan and SARS‐CoV‐2/B.1.1.7. Moreover, the presence of a stop codon at position 27 in orf8 connotes the emergence of two frames (orf8a and orf8b) in SARS‐CoV‐2, which further hampers its extracellular secretion, and in turn, immunogenicity. The findings of the present study could further be used to develop targeted immunotherapeutics. Evasion from the host immune response is one of the most common yet profoundly effective strategies that pathogens including viruses employ to win the evolutionary arms race against their hosts. The The variant was first reported in the UK on September 20, 2020, but since then its presence has also been observed in other parts of the world. 4, 5 The transmissibility and virulence of the new SARS-CoV-2/ B.1.1.7 have not been thoroughly compared with the other variants of SARS-CoV-2. However, its emergence parallels the significant rise in the SARS-CoV-2 infections in the UK with reported cases of infection and related deaths spiked 1.4-and 1.3-folds, respectively, from October 2020 to December 2020. 6 Preliminary genomic analysis showed the presence and/or accumulation of significant numbers of non-synonymous mutations in the SARS-CoV-2/B.1.1.7 compared to SARS-CoV-2/Wuhan. 4 The location and span of the residues predicted to be epitopic were found nearly identical in both the proteins ( Figure 2D and Table S2 ). However, in SARS-CoV-2/Wuhan, the sites predicted to be epitopic at the region, Thr250-Gly252, were found extended (Ser244-Gly249) in SARS/CoV-2/B.1.1.7 ( Figure 2D ). The regions were also found to vary in their electrostatic properties where the former was more polar compared to the latter ( Figure 2E ). This could potentially be due to the change in the spatial conformation of the [25] [26] [27] Moreover, it has also been pointed that convalescent plasma actually increases the genetic variations (via selection pressure) in the virus within the host. 25 In summary, this may indicate that the immunogenic potential of SARS-CoV-2/B.1.1.7 spike protein may in part be different than SARS-CoV-2/Wuhan. Orf8 is an accessory protein of SARS-CoV-2 and has not been found in many other coronaviruses that infect humans. However, the protein has been found in both SARS-CoV and SARS-CoV-2 and showed sequence similarity with some of the bat coronaviruses (RaTG13, ZXC21, and ZC45). The functionality of SARS-CoV-2 orf8 is not known to its full extent, however, it has been suggested that the protein may be involved in the immune evasion and cytokine response mimicking. The presence of N-terminal signal peptide (Met1-Ala15) allows orf8 of SARS-CoV-2 to be released from the infected cells and also leads to the immune response. Seropositivity and circulating peptides of orf8 have been observed along with anti orf8 IgG, IgM, and IgA within the infected individuals. 8 However, orf8 of SARS-CoV has not been reported for any seropositivity, this is possibly because of the deletion of 29 nucleotides. This in turn has resulted in the formation of two chains of orf8 during the earlier phase of the SARS-CoV epidemic, potentially impairing its extracellular secretion. 11 The presence of the stop codon mutation in orf8 in SARS-CoV-2/B.1.1.7 could then also lead to the formation of two chains, orf8a and orf8b, where the latter (larger chain) does not harbor the leader peptide required for the extracellular secretion, thus reducing its immunogenicity and/or seropositivity. Therefore, it is possible to conceive that orf8 of SARS-CoV-2/B.1.1.7 may not be as immunogenic as compared to SARS-CoV-2/Wuhan orf8. Moreover, sequence variations may have resulted in the loss of many epitopic sites in the orf8 of SARS-CoV-2/B.1.1.7 compared to SARS-CoV-2/Wuhan. For SARS-CoV, it has been shown that viruses containing orf8b replicate more efficiently in the presence of interferon compared to the virus with intact orf8. 28 Therefore, it is possible that this potential emergence of orf8a and orf8b in SARS-CoV-2/B.1.1.7, may also confer a similar evolutionary advantage. Furthermore, Val77 of SARS-CoV orf8b has been shown critical to induce orf8b intracellular aggregation, lysosomal stress, autophagy and interleukin-mediated inflammatory response via triggering NLRP3. 29 This Val77 is conserved in both the compared SARS-CoV2/Wuhan and SARS-CoV-2/B.1.1.7. Therefore, it is possible that as in SARS-CoV, its (Val77) exclusive presence in a separate and nonsecretory chain in SARS-CoV-2/B.1.1.7 may also contribute to the pathological manifestation of the infection. 29 Taken together, it is likely that SARS-CoV-2/B.1.1.7 may bear an evolutionary advantage over SARS-CoV-2/Wuhan, mediated not only by the antigenic changes in its spike and orf8 proteins but could also by an elevated cytokine-mediated inflammatory response in the host due to the intracellular aggregation. The study in part is supported by the Higher Education Commission, Government of Pakistan (NRPU-3857). WHO. Weekly operational update on COVID-19 -13 GISAID Investigation of novel SARS-COV-2 variant: Variant of Concern 202012/01. 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Hussain, Sanya Shabbir, Anusha Amanullah, Fozia Raza, Muhammad J. Imdad, and Sahar Zahid executed the study and were involved in the analysis and writing of the manuscript. The peer review history for this article is available at https://publons. com/publon/10.1002/jmv.26931 Data available in article supplementary material. http://orcid.org/0000-0001-8862-179X