Origin and Structural Biology of Novel Coronavirus (SARS-CoV-2).

INTRODUCTION: A recent rapid outbreak of infection around the globe has been caused by a novel coronavirus, now known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which was first identified in December 2019 in Wuhan city of Hubei province, People's Republic of China. METHODS: We reviewed the currently available literature on coronaviruses. RESULTS: Coronaviruses are a group of enveloped viruses with non-segmented, single-stranded, and positive-sense RNA genomes. Although 13 variation sites in open reading frames have been identified among SARS-CoV-2 strains, no mutation has been observed so far in envelop protein. The origin and structural biology of SARS-CoV-2 in details are discussed. CONCLUSIONS: Origin and structural biology will help the researchers identify the virus's mechanism in the host and drug design. Currently, no clinical treatments or prevention strategies are available for any human coronavirus.

Insights on the Structural Variations of the Furin-Like Cleavage Site Found Among the December 2019-July 2020 SARS-CoV-2 Spike Glycoprotein: A Computational Study Linking Viral Evolution and Infection.

The SARS-CoV-2 (SARS2) is the cause of the coronavirus disease 2019 (COVID-19) pandemic. One unique structural feature of the SARS2 spike protein is the presence of a furin-like cleavage site (FLC) which is associated with both viral pathogenesis and host tropism. Specifically, SARS2 spike protein binds to the host ACE-2 receptor which in-turn is cleaved by furin proteases at the FLC site, suggesting that SARS2 FLC structural variations may have an impact on viral infectivity. However, this has not yet been fully elucidated. This study designed and analyzed a COVID-19 genomic epidemiology network for December 2019 to July 2020, and subsequently generated and analyzed representative SARS2 spike protein models from significant node clusters within the network. To distinguish possible structural variations, a model quality assessment was performed before further protein model analyses and superimposition of the protein models, particularly in both the receptor-binding domain (RBD) and FLC. Mutant spike models were generated with the unique 681PRRA684 amino acid sequence found within the deleted FLC. We found 9 SARS2 FLC structural patterns that could potentially correspond to nine node clusters encompassing various countries found within the COVID-19 genomic epidemiology network. Similarly, we associated this with the rapid evolution of the SARS2 genome. Furthermore, we observed that either in the presence or absence of the unique 681PRRA684 amino acid sequence no structural changes occurred within the SARS2 RBD, which we believe would mean that the SARS2 FLC has no structural influence on SARS2 RBD and may explain why host tropism was maintained.

Multiple Recombination Events and Strong Purifying Selection at the Origin of SARS-CoV-2 Spike Glycoprotein Increased Correlated Dynamic Movements.

Our evolutionary and structural analyses revealed that the severe acute respiratory syndrome (SARS) coronavirus 2 (SARS-CoV-2) spike gene is a complex mosaic resulting from several recombination events. Additionally, the fixation of variants has mainly been driven by purifying selection, suggesting the presence of conserved structural features. Our dynamic simulations identified two main long-range covariant dynamic movements of the novel glycoprotein, and showed that, as a result of the evolutionary duality, they are preserved. The first movement involves the receptor binding domain with the N-terminal domain and the C-terminal domain 2 and is maintained across human, bat and pangolin coronaviruses. The second is a complex network of long-range dynamics specific to SARS-CoV-2 involving the novel PRRA and the conserved KR*SF cleavage sites, as well as conserved segments in C-terminal domain 3. These movements, essential for host cell binding, are maintained by hinges conserved across human, bat, and pangolin coronaviruses glycoproteins. The hinges, located around Threonine 333 and Proline 527 within the N-terminal domain and C-terminal domain 2, represent candidate targets for the future development of novel pan-coronavirus inhibitors. In summary, we show that while recombination created a new configuration that increased the covariant dynamic movements of the SARS-CoV-2 glycoprotein, negative selection preserved its inter-domain structure throughout evolution in different hosts and inter-species transmissions.

SARS-coronavirus-2 replication in Vero E6 cells: replication kinetics, rapid adaptation and cytopathology.

The sudden emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the end of 2019 from the Chinese province of Hubei and its subsequent pandemic spread highlight the importance of understanding the full molecular details of coronavirus infection and pathogenesis. Here, we compared a variety of replication features of SARS-CoV-2 and SARS-CoV and analysed the cytopathology caused by the two closely related viruses in the commonly used Vero E6 cell line. Compared to SARS-CoV, SARS-CoV-2 generated higher levels of intracellular viral RNA, but strikingly about 50-fold less infectious viral progeny was recovered from the culture medium. Immunofluorescence microscopy of SARS-CoV-2-infected cells established extensive cross-reactivity of antisera previously raised against a variety of non-structural proteins, membrane and nucleocapsid protein of SARS-CoV. Electron microscopy revealed that the ultrastructural changes induced by the two SARS viruses are very similar and occur within comparable time frames after infection. Furthermore, we determined that the sensitivity of the two viruses to three established inhibitors of coronavirus replication (remdesivir, alisporivir and chloroquine) is very similar, but that SARS-CoV-2 infection was substantially more sensitive to pre-treatment of cells with pegylated interferon alpha. An important difference between the two viruses is the fact that - upon passaging in Vero E6 cells - SARS-CoV-2 apparently is under strong selection pressure to acquire adaptive mutations in its spike protein gene. These mutations change or delete a putative furin-like cleavage site in the region connecting the S1 and S2 domains and result in a very prominent phenotypic change in plaque assays.