The Biological Properties of the SARS-CoV-2 Cameroon Variant Spike: An Intermediate between the Alpha and Delta Variants.

An analysis of the structural effect of the mutations of the B.1.640.2 (IHU) Spike Receptor Binding Domain (RBD) and N-terminal Domain (NTD) is reported along with a comparison with the sister lineage B.1.640.1. and a selection of variants of concern. The effect of the mutations on the RBD-ACE2 interaction was also assessed. The structural analysis applied computational methods that are able to carry out in silico mutagenesis to calculate energy minimization and the folding energy variation consequent to residue mutations. Tools for electrostatic calculation were applied to quantify and display the protein surface electrostatic potential. Interactions at the RBD-ACE2 interface were scrutinized using computational tools that identify the interactions and predict the contribution of each interface residue to the stability of the complex. The comparison among the RBDs shows that the most evident differences between the variants is in the distribution of the surface electrostatic potential: that of B.1.640.1 is as that of the Alpha RBD, while B.1.640.2 appears to have an intermediate surface potential pattern with characteristics between those of the Alpha and Delta variants. Moreover, the B.1.640.2 Spike includes the mutation E484K that in other variants has been suggested to be involved in immune evasion. These properties may hint at the possibility that B.1.640.2 emerged with a potentially increased infectivity with respect to the sister B.1.640.1 variant, but significantly lower than that of the Delta and Omicron variants. However, the analysis of their NTD domains highlights deletions, destabilizing mutations and charge alterations that can limit the ability of the B.1.640.1 and B.1.640.2 variants to interact with cellular components, such as cell surface receptors.

The SARS-CoV-2 Mu variant should not be left aside: It warrants attention for its immuno-escaping ability.

The COVID-19 pandemic continues to have a threatening impact on a global scale, largely due to the emergence of newly SARS-CoV-2 variants. The Mu (PANGO lineage B.1.621), was first identified in Colombia in January 2021 and was classified as a variant of interest (VOI) in August 2021, due to a constellation of mutations that likely-mediate an unexpectedly enhanced immune resistance to inactivated vaccine-elicited antibodies. Despite recent studies suggesting that the Mu variant appears to have less infectivity than the Delta variant, here we examined the structural effect of the Mu spike protein mutations and predicted the potential impact on infectivity of the Mu variant compared with the Delta and Delta plus spike protein.

SARS-CoV-2 Pandemic Tracing in Italy Highlights Lineages with Mutational Burden in Growing Subsets.

Tracing the appearance and evolution of virus variants is essential in the management of the COVID-19 pandemic. Here, we focus on SARS-CoV-2 spread in Italian patients by using viral sequences deposited in public databases and a tracing procedure which is used to monitor the evolution of the pandemic and detect the spreading, within the infected population of emergent sub-clades with a potential positive selection. Analyses of a collection of monthly samples focused on Italy highlighted the appearance and evolution of all the main viral sub-trees emerging at the end of the first year of the pandemic. It also identified additional expanding subpopulations which spread during the second year (i.e., 2021). Three-dimensional (3D) modelling of the main amino acid changes in mutated viral proteins, including ORF1ab (nsp3, nsp4, 2'-o-ribose methyltransferase, nsp6, helicase, nsp12 [RdRp]), N, ORF3a, ORF8, and spike proteins, shows the potential of the analysed structural variations to result in epistatic modulation and positive/negative selection pressure. These analyzes will be of importance to the early identification of emerging clades, which can develop into new "variants of concern" (i.e., VOC). These analyses and settings will also help SARS-CoV-2 coronet genomic centers in other countries to trace emerging worldwide variants.

SARS-CoV-2 B.1.617 Indian variants: Are electrostatic potential changes responsible for a higher transmission rate?

Lineage B.1.617+, also known as G/452R.V3 and now denoted by WHO with the Greek letters δ and κ, is a recently described SARS-CoV-2 variant under investigation first identified in October 2020 in India. As of May 2021, three sublineages labeled as B.1.617.1 (κ), B.1.617.2 (δ), and B.1.617.3 have been already identified, and their potential impact on the current pandemic is being studied. This variant has 13 amino acid changes, three in its spike protein, which are currently of particular concern: E484Q, L452R, and P681R. Here, we report a major effect of the mutations characterizing this lineage, represented by a marked alteration of the surface electrostatic potential (EP) of the receptor-binding domain (RBD) of the spike protein. Enhanced RBD-EP is particularly noticeable in the B.1.617.2 (δ) sublineage, which shows multiple replacements of neutral or negatively charged amino acids with positively charged amino acids. We here hypothesize that this EP change can favor the interaction between the B.1.617+ RBD and the negatively charged ACE2, thus conferring a potential increase in the virus transmission.

Shortening Epitopes to Survive: The Case of SARS-CoV-2 Lambda Variant.

Among the more recently identified SARS-CoV-2 Variants of Interest (VOI) is the Lambda variant, which emerged in Peru and has rapidly spread to South American regions and the US. This variant remains poorly investigated, particularly regarding the effects of mutations on the thermodynamic parameters affecting the stability of the Spike protein and its Receptor Binding Domain. We report here an in silico study on the potential impact of the Spike protein mutations on the immuno-escape ability of the Lambda variant. Bioinformatics analysis suggests that a combination of shortening the immunogenic epitope loops and the generation of potential N-glycosylation sites may be a viable adaptation strategy, potentially allowing this emerging viral variant to escape from host immunity.

Long-chain polyphosphates impair SARS-CoV-2 infection and replication.

Inorganic polyphosphates (polyPs) are linear polymers composed of repeated phosphate (PO4 3-) units linked together by multiple high-energy phosphoanhydride bonds. In addition to being a source of energy, polyPs have cytoprotective and antiviral activities. Here, we investigated the antiviral activities of long-chain polyPs against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. In molecular docking analyses, polyPs interacted with several conserved amino acid residues in angiotensin-converting enzyme 2 (ACE2), the host receptor that facilitates virus entry, and in viral RNA-dependent RNA polymerase (RdRp). ELISA and limited proteolysis assays using nano- LC-MS/MS mapped polyP120 binding to ACE2, and site-directed mutagenesis confirmed interactions between ACE2 and SARS-CoV-2 RdRp and identified the specific amino acid residues involved. PolyP120 enhanced the proteasomal degradation of both ACE2 and RdRp, thus impairing replication of the British B.1.1.7 SARS-CoV-2 variant. We thus tested polyPs for functional interactions with the virus in SARS-CoV-2-infected Vero E6 and Caco2 cells and in primary human nasal epithelial cells. Delivery of a nebulized form of polyP120 reduced the amounts of viral positive-sense genomic and subgenomic RNAs, of RNA transcripts encoding proinflammatory cytokines, and of viral structural proteins, thereby presenting SARS-CoV-2 infection in cells in vitro.

The importance of genomic analysis in cracking the coronavirus pandemic.

Introduction: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has pushed the scientific community to undertake intense research efforts. Understanding SARS-CoV-2 biology is necessary to discover therapeutic or preventive strategies capable of containing the pandemic. Knowledge of the structural characteristics of the virus genome and proteins is essential to find targets for therapies and immunological interventions.Areas covered: This review covers different areas of expertise, genomic analysis of circulating strains, structural biology, viral mutations, molecular diagnostics, disease, and vaccines. In particular, the review is focused on the molecular approaches and modern clinical strategies used in these fields.Expert opinion: Molecular approaches to SARS-CoV-2 pandemic have been critical to shorten time for new diagnostic, therapeutic and prevention strategies. In this perspective, the entire scientific community is moving in the same direction. Vaccines, together with the development of new drugs to treat the disease, represent the most important strategy to protect human from viral disease and prevent further spread. In this regard, new molecular technologies have been successfully implemented. The use of a novel strategy of communication is suggested for a better diffusion to the broader public of new data and results.

SARS-Cov-2 ORF3a: Mutability and function.

In this study, analysis of changes of SARS-CoV-2 ORF3a protein during pandemic is reported. ORF3a, a conserved coronavirus protein, is involved in virus replication and release. A set of 70,752 high-quality SARS-CoV-2 genomes available in GISAID databank at the end of August 2020 have been scanned. All ORF3a mutations in the virus genomes were grouped according to the collection date interval and over the entire data set. The considered intervals were: start of collection-February, March, April, May, June, July and August 2020. The top five most frequent variants were examined within each collection interval. Overall, seventeen variants have been isolated. Ten of the seventeen mutant sites occur within the transmembrane (TM) domain of ORF3a and are in contact with the central pore or side tunnels. The other variant sites are in different places of the ORF3a structure. Within the entire sample, the five most frequent mutations are V13L, Q57H, Q57H + A99V, G196V and G252V. The same analysis identified 28 sites identically conserved in all the genome isolates. These sites are possibly involved in stabilization of monomer, dimer, tetramerization and interaction with other cellular components. The results here reported can be helpful to understand virus biology and to design new therapeutic strategies.

Evidence for mutations in SARS-CoV-2 Italian isolates potentially affecting virus transmission.

Italy is the first western country suffering heavy severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission and disease impact after coronavirus disease-2019 pandemia started in China. Even though the presence of mutations on spike glycoprotein and nucleocapsid in Italian isolates has been reported, the potential impact of these mutations on viral transmission has not been evaluated. We have compared SARS-CoV-2 genome sequences from Italian patients with virus sequences from Chinese patients. We focussed upon three nonsynonymous mutations of genes coding for S(one) and N (two) viral proteins present in Italian isolates and absent in Chinese ones, using various bioinformatics tools. Amino acid analysis and changes in three-dimensional protein structure suggests the mutations reduce protein stability and, particularly for S1 mutation, the enhanced torsional ability of the molecule could favor virus binding to cell receptor(s). This theoretical interpretation awaits experimental and clinical confirmation.

COVID-2019: The role of the nsp2 and nsp3 in its pathogenesis.

Last December 2019, a new virus, named novel Coronavirus (COVID-2019) causing many cases of severe pneumonia was reported in Wuhan, China. The virus knowledge is limited and especially about COVID-2019 pathogenesis. The Open Reading Frame 1ab (ORF1ab) of COVID-2019 has been analyzed to evidence the presence of mutation caused by selective pressure on the virus. For selective pressure analysis fast-unconstrained Bayesian approximation (FUBAR) was used. Homology modelling has been performed by SwissModel and HHPred servers. The presence of transmembrane helical segments in Coronavirus ORF1ab non structural protein 2 (nsp2) and nsp3 was tested by TMHMM, MEMSAT, and MEMPACK tools. Three-dimensional structures have been analyzed and displayed using PyMOL. FUBAR analysis revealed the presence of potential sites under positive selective pressure (P < .05). Position 723 in the COVID-2019 has a serine instead a glycine residue, while at aminoacidic position 1010 a proline instead an isoleucine. Significant (P < .05) pervasive negative selection in 2416 sites (55%) was found. The positive selective pressure could account for some clinical features of this virus compared with severe acute respiratory syndrome (SARS) and Bat SARS-like CoV. The stabilizing mutation falling in the endosome-associated-protein-like domain of the nsp2 protein could account for COVID-2019 high ability of contagious, while the destabilizing mutation in nsp3 proteins could suggest a potential mechanism differentiating COVID-2019 from SARS. These data could be helpful for further investigation aimed to identify potential therapeutic targets or vaccine strategy, especially in the actual moment when the epidemic is ongoing and the scientific community is trying to enrich knowledge about this new viral pathogen.

Sars-CoV-2 Envelope and Membrane Proteins: Structural Differences Linked to Virus Characteristics?

The Coronavirus Disease 2019 (COVID-19) is a new viral infection caused by the severe acute respiratory coronavirus 2 (SARS-CoV-2). Genomic analyses have revealed that SARS-CoV-2 is related to Pangolin and Bat coronaviruses. In this report, a structural comparison between the Sars-CoV-2 Envelope and Membrane proteins from different human isolates with homologous proteins from closely related viruses is described. The analyses here reported show the high structural similarity of Envelope and Membrane proteins to the counterparts from Pangolin and Bat coronavirus isolates. However, the comparisons have also highlighted structural differences specific of Sars-CoV-2 proteins which may be correlated to the cross-species transmission and/or to the properties of the virus. Structural modelling has been applied to map the variant sites onto the predicted three-dimensional structure of the Envelope and Membrane proteins.

Evolutionary analysis of SARS-CoV-2: how mutation of Non-Structural Protein 6 (NSP6) could affect viral autophagy.

BACKGROUND: SARS-CoV-2 is a new coronavirus that has spread globally, infecting more than 150000 people, and being declared pandemic by the WHO. We provide here bio-informatic, evolutionary analysis of 351 available sequences of its genome with the aim of mapping genome structural variations and the patterns of selection. METHODS: A Maximum likelihood tree has been built and selective pressure has been investigated in order to find any mutation developed during the SARS-CoV-2 epidemic that could potentially affect clinical evolution of the infection. FINDING: We have found in more recent isolates the presence of two mutations affecting the Non-Structural Protein 6 (NSP6) and the Open Reding Frame10 (ORF 10) adjacent regions. Amino acidic change stability analysis suggests both mutations could confer lower stability of the protein structures. INTERPRETATION: One of the two mutations, likely developed within the genome during virus spread, could affect virus intracellular survival. Genome follow-up of SARS-CoV-2 spread is urgently needed in order to identify mutations that could significantly modify virus pathogenicity.