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1.
PLoS One ; 17(3): e0264640, 2022.
Article in English | MEDLINE | ID: covidwho-1731599

ABSTRACT

The SARS-CoV-2 is the third coronavirus in addition to SARS-CoV and MERS-CoV that causes severe respiratory syndrome in humans. All of them likely crossed the interspecific barrier between animals and humans and are of zoonotic origin, respectively. The origin and evolution of viruses and their phylogenetic relationships are of great importance for study of their pathogenicity and development of antiviral drugs and vaccines. The main objective of the presented study was to compare two methods for identifying relationships between coronavirus genomes: phylogenetic one based on the whole genome alignment followed by molecular phylogenetic tree inference and alignment-free clustering of triplet frequencies, respectively, using 69 coronavirus genomes selected from two public databases. Both approaches resulted in well-resolved robust classifications. In general, the clusters identified by the first approach were in good agreement with the classes identified by the second using K-means and the elastic map method, but not always, which still needs to be explained. Both approaches demonstrated also a significant divergence of genomes on a taxonomic level, but there was less correspondence between genomes regarding the types of diseases they caused, which may be due to the individual characteristics of the host. This research showed that alignment-free methods are efficient in combination with alignment-based methods. They have a significant advantage in computational complexity and provide valuable additional alternative information on the genomes relationships.


Subject(s)
Comparative Genomic Hybridization/methods , Coronavirus/genetics , Genome, Viral , Chromosome Mapping , Cluster Analysis , Coronavirus/classification , Humans , Phylogeny , SARS-CoV-2/classification , SARS-CoV-2/genetics , Sequence Alignment
2.
J Med Virol ; 94(4): 1627-1632, 2022 04.
Article in English | MEDLINE | ID: covidwho-1718407

ABSTRACT

Following the discovery of the SARS-CoV-2 Omicron variant (B.1.1.529), the global COVID-19 outbreak has resurfaced after appearing to be relentlessly spreading over the past 2 years. This new variant showed marked degree of mutation, compared with the previous SARS-CoV-2 variants. This study investigates the evolutionary links between Omicron variant and recently emerged SARS-CoV-2 variants. The entire genome sequences of SARS-CoV-2 variants were obtained, aligned using Clustal Omega, pairwise comparison was computed, differences, identity percent, gaps, and mutations were noted, and the identity matrix was generated. The phylogenetics of Omicron variants were determined using a variety of evolutionary substitution models. The ultrametric and metric clustering methods, such as UPGMA and neighbor-joining (NJ), using nucleotide substitution models that allowed the inclusion of nucleotide transitions and transversions as Kimura 80 models, revealed that the Omicron variant forms a new monophyletic clade that is distant from other SARS-CoV-2 variants. In contrast, the NJ method using a basic nucleotide substitution model such as Jukes-Cantor revealed a close relationship between the Omicron variant and the recently evolved Alpha variant. Based on the percentage of sequence identity, the closest variants were in the following order: Omicron, Alpha, Gamma, Delta, Beta, Mu, and then the SARS-CoV-2 USA isolate. A genome alignment with other variants indicated the greatest number of gaps in the Omicron variant's genome ranging from 43 to 63 gaps. It is possible, given their close relationship to the Alpha variety, that Omicron has been around for much longer than predicted, even though they created a separate monophyletic group. Sequencing initiatives in a systematic and comprehensive manner is highly recommended to study the evolution and mutations of the virus.


Subject(s)
Evolution, Molecular , Genome, Viral/genetics , Phylogeny , SARS-CoV-2/genetics , Base Sequence , COVID-19/epidemiology , COVID-19/virology , Humans , Mutation , Sequence Alignment
3.
Protein J ; 39(3): 198-216, 2020 06.
Article in English | MEDLINE | ID: covidwho-1718840

ABSTRACT

The devastating effects of the recent global pandemic (termed COVID-19 for "coronavirus disease 2019") caused by the severe acute respiratory syndrome coronavirus-2 (SARS CoV-2) are paramount with new cases and deaths growing at an exponential rate. In order to provide a better understanding of SARS CoV-2, this article will review the proteins found in the SARS CoV-2 that caused this global pandemic.


Subject(s)
Betacoronavirus/chemistry , Betacoronavirus/physiology , Coronavirus Infections/virology , Pneumonia, Viral/virology , Viral Proteins/chemistry , Viral Proteins/metabolism , Amino Acid Sequence , Betacoronavirus/genetics , COVID-19 , Coronavirus Envelope Proteins , Coronavirus Infections/drug therapy , Coronavirus Infections/metabolism , Coronavirus Nucleocapsid Proteins , Drug Discovery/methods , Genome, Viral , Host-Pathogen Interactions/drug effects , Humans , Nucleocapsid Proteins/chemistry , Nucleocapsid Proteins/genetics , Nucleocapsid Proteins/metabolism , Pandemics , Phosphoproteins , Pneumonia, Viral/drug therapy , Pneumonia, Viral/metabolism , Polyproteins , Protein Interaction Maps/drug effects , SARS-CoV-2 , Sequence Alignment , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Viral Envelope Proteins/chemistry , Viral Envelope Proteins/genetics , Viral Envelope Proteins/metabolism , Viral Matrix Proteins/chemistry , Viral Matrix Proteins/genetics , Viral Matrix Proteins/metabolism , Viral Nonstructural Proteins/chemistry , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/metabolism , Viral Proteins/genetics , Viral Regulatory and Accessory Proteins/chemistry , Viral Regulatory and Accessory Proteins/genetics , Viral Regulatory and Accessory Proteins/metabolism , Viroporin Proteins
4.
Microbiol Spectr ; 10(1): e0278021, 2022 02 23.
Article in English | MEDLINE | ID: covidwho-1700612

ABSTRACT

Understanding the immune response to severe acute respiratory syndrome coronavirus (SARS-CoV-2) is critical to overcome the current coronavirus disease (COVID-19) pandemic. Efforts are being made to understand the potential cross-protective immunity of memory T cells, induced by prior encounters with seasonal coronaviruses, in providing protection against severe COVID-19. In this study we assessed T-cell responses directed against highly conserved regions of SARS-CoV-2. Epitope mapping revealed 16 CD8+ T-cell epitopes across the nucleocapsid (N), spike (S), and open reading frame (ORF)3a proteins of SARS-CoV-2 and five CD8+ T-cell epitopes encoded within the highly conserved regions of the ORF1ab polyprotein of SARS-CoV-2. Comparative sequence analysis showed high conservation of SARS-CoV-2 ORF1ab T-cell epitopes in seasonal coronaviruses. Paradoxically, the immune responses directed against the conserved ORF1ab epitopes were infrequent and subdominant in both convalescent and unexposed participants. This subdominant immune response was consistent with a low abundance of ORF1ab encoded proteins in SARS-CoV-2 infected cells. Overall, these observations suggest that while cross-reactive CD8+ T cells likely exist in unexposed individuals, they are not common and therefore are unlikely to play a significant role in providing broad preexisting immunity in the community. IMPORTANCE T cells play a critical role in protection against SARS-CoV-2. Despite being highly topical, the protective role of preexisting memory CD8+ T cells, induced by prior exposure to circulating common coronavirus strains, remains less clear. In this study, we established a robust approach to specifically assess T cell responses to highly conserved regions within SARS-CoV-2. Consistent with recent observations we demonstrate that recognition of these highly conserved regions is associated with an increased likelihood of milder disease. However, extending these observations we observed that recognition of these conserved regions is rare in both exposed and unexposed volunteers, which we believe is associated with the low abundance of these proteins in SARS-CoV-2 infected cells. These observations have important implications for the likely role preexisting immunity plays in controlling severe disease, further emphasizing the importance of vaccination to generate the immunodominant T cells required for immune protection.


Subject(s)
COVID-19/immunology , Epitopes, T-Lymphocyte/immunology , SARS-CoV-2/immunology , Amino Acid Sequence , CD8-Positive T-Lymphocytes/immunology , COVID-19/genetics , COVID-19/virology , Conserved Sequence , Coronavirus/chemistry , Coronavirus/classification , Coronavirus/genetics , Coronavirus/immunology , Coronavirus Infections/genetics , Coronavirus Infections/immunology , Coronavirus Infections/virology , Cross Reactions , Epitope Mapping , Epitopes, T-Lymphocyte/chemistry , Epitopes, T-Lymphocyte/genetics , Humans , SARS-CoV-2/chemistry , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Sequence Alignment , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/immunology
5.
Nat Commun ; 13(1): 317, 2022 01 14.
Article in English | MEDLINE | ID: covidwho-1671550

ABSTRACT

Activation of the serum-resident complement system begins a cascade that leads to activation of membrane-resident complement receptors on immune cells, thus coordinating serum and cellular immune responses. Whilst many molecules act to control inappropriate activation, Properdin is the only known positive regulator of the human complement system. By stabilising the alternative pathway C3 convertase it promotes complement self-amplification and persistent activation boosting the magnitude of the serum complement response by all triggers. In this work, we identify a family of tick-derived alternative pathway complement inhibitors, hereafter termed CirpA. Functional and structural characterisation reveals that members of the CirpA family directly bind to properdin, inhibiting its ability to promote complement activation, and leading to potent inhibition of the complement response in a species specific manner. We provide a full functional and structural characterisation of a properdin inhibitor, opening avenues for future therapeutic approaches.


Subject(s)
Arthropod Proteins/chemistry , Arthropod Proteins/immunology , Complement Inactivating Agents/chemistry , Complement Inactivating Agents/immunology , Properdin/immunology , Rhipicephalus/immunology , Amino Acid Sequence , Animals , Arthropod Proteins/genetics , Complement Activation , Complement C3/chemistry , Complement C3/immunology , Complement Pathway, Alternative , Humans , Kinetics , Properdin/chemistry , Properdin/genetics , Rhipicephalus/chemistry , Rhipicephalus/genetics , Sequence Alignment
6.
Nat Commun ; 12(1): 7325, 2021 12 16.
Article in English | MEDLINE | ID: covidwho-1585854

ABSTRACT

Single-domain Variable New Antigen Receptors (VNARs) from the immune system of sharks are the smallest naturally occurring binding domains found in nature. Possessing flexible paratopes that can recognize protein motifs inaccessible to classical antibodies, VNARs have yet to be exploited for the development of SARS-CoV-2 therapeutics. Here, we detail the identification of a series of VNARs from a VNAR phage display library screened against the SARS-CoV-2 receptor binding domain (RBD). The ability of the VNARs to neutralize pseudotype and authentic live SARS-CoV-2 virus rivalled or exceeded that of full-length immunoglobulins and other single-domain antibodies. Crystallographic analysis of two VNARs found that they recognized separate epitopes on the RBD and had distinctly different mechanisms of virus neutralization unique to VNARs. Structural and biochemical data suggest that VNARs would be effective therapeutic agents against emerging SARS-CoV-2 mutants, including the Delta variant, and coronaviruses across multiple phylogenetic lineages. This study highlights the utility of VNARs as effective therapeutics against coronaviruses and may serve as a critical milestone for nearing a paradigm shift of the greater biologic landscape.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , Crystallography, X-Ray , Receptors, Antigen/chemistry , Receptors, Antigen/immunology , Sharks/immunology , Angiotensin-Converting Enzyme 2 , Animals , COVID-19 , Epitopes , Mutation , Phylogeny , Protein Binding , SARS-CoV-2 , Sequence Alignment , Single-Domain Antibodies , Spike Glycoprotein, Coronavirus/immunology
7.
Proc Natl Acad Sci U S A ; 118(52)2021 12 28.
Article in English | MEDLINE | ID: covidwho-1565770

ABSTRACT

The constant emergence of COVID-19 variants reduces the effectiveness of existing vaccines and test kits. Therefore, it is critical to identify conserved structures in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomes as potential targets for variant-proof diagnostics and therapeutics. However, the algorithms to predict these conserved structures, which simultaneously fold and align multiple RNA homologs, scale at best cubically with sequence length and are thus infeasible for coronaviruses, which possess the longest genomes (∼30,000 nt) among RNA viruses. As a result, existing efforts on modeling SARS-CoV-2 structures resort to single-sequence folding as well as local folding methods with short window sizes, which inevitably neglect long-range interactions that are crucial in RNA functions. Here we present LinearTurboFold, an efficient algorithm for folding RNA homologs that scales linearly with sequence length, enabling unprecedented global structural analysis on SARS-CoV-2. Surprisingly, on a group of SARS-CoV-2 and SARS-related genomes, LinearTurboFold's purely in silico prediction not only is close to experimentally guided models for local structures, but also goes far beyond them by capturing the end-to-end pairs between 5' and 3' untranslated regions (UTRs) (∼29,800 nt apart) that match perfectly with a purely experimental work. Furthermore, LinearTurboFold identifies undiscovered conserved structures and conserved accessible regions as potential targets for designing efficient and mutation-insensitive small-molecule drugs, antisense oligonucleotides, small interfering RNAs (siRNAs), CRISPR-Cas13 guide RNAs, and RT-PCR primers. LinearTurboFold is a general technique that can also be applied to other RNA viruses and full-length genome studies and will be a useful tool in fighting the current and future pandemics.


Subject(s)
Algorithms , RNA, Viral/chemistry , SARS-CoV-2/chemistry , Betacoronavirus/chemistry , Betacoronavirus/genetics , Conserved Sequence , Genome, Viral , Mutation , Nucleic Acid Conformation , RNA Folding , RNA, Viral/genetics , SARS-CoV-2/genetics , Sequence Alignment
8.
J Med Virol ; 94(4): 1627-1632, 2022 04.
Article in English | MEDLINE | ID: covidwho-1565207

ABSTRACT

Following the discovery of the SARS-CoV-2 Omicron variant (B.1.1.529), the global COVID-19 outbreak has resurfaced after appearing to be relentlessly spreading over the past 2 years. This new variant showed marked degree of mutation, compared with the previous SARS-CoV-2 variants. This study investigates the evolutionary links between Omicron variant and recently emerged SARS-CoV-2 variants. The entire genome sequences of SARS-CoV-2 variants were obtained, aligned using Clustal Omega, pairwise comparison was computed, differences, identity percent, gaps, and mutations were noted, and the identity matrix was generated. The phylogenetics of Omicron variants were determined using a variety of evolutionary substitution models. The ultrametric and metric clustering methods, such as UPGMA and neighbor-joining (NJ), using nucleotide substitution models that allowed the inclusion of nucleotide transitions and transversions as Kimura 80 models, revealed that the Omicron variant forms a new monophyletic clade that is distant from other SARS-CoV-2 variants. In contrast, the NJ method using a basic nucleotide substitution model such as Jukes-Cantor revealed a close relationship between the Omicron variant and the recently evolved Alpha variant. Based on the percentage of sequence identity, the closest variants were in the following order: Omicron, Alpha, Gamma, Delta, Beta, Mu, and then the SARS-CoV-2 USA isolate. A genome alignment with other variants indicated the greatest number of gaps in the Omicron variant's genome ranging from 43 to 63 gaps. It is possible, given their close relationship to the Alpha variety, that Omicron has been around for much longer than predicted, even though they created a separate monophyletic group. Sequencing initiatives in a systematic and comprehensive manner is highly recommended to study the evolution and mutations of the virus.


Subject(s)
Evolution, Molecular , Genome, Viral/genetics , Phylogeny , SARS-CoV-2/genetics , Base Sequence , COVID-19/epidemiology , COVID-19/virology , Humans , Mutation , Sequence Alignment
9.
Sci Rep ; 11(1): 23622, 2021 12 08.
Article in English | MEDLINE | ID: covidwho-1559938

ABSTRACT

Spike glycoprotein (Sgp) is liable for binding of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) to the host receptors. Since Sgp is the main target for vaccine and drug designing, elucidating its mutation pattern could help in this regard. This study is aimed at investigating the correspondence of specific residues to the SgpSARS-CoV-2 functionality by explorative interpretation of sequence alignments. Centrality analysis of the Sgp dissects the importance of these residues in the interaction network of the RBD-ACE2 (receptor-binding domain) complex and furin cleavage site. Correspondence of RBD to threonine500 and asparagine501 and furin cleavage site to glutamine675, glutamine677, threonine678, and alanine684 was observed; all residues are exactly located at the interaction interfaces. The harmonious location of residues dictates the RBD binding property and the flexibility, hydrophobicity, and accessibility of the furin cleavage site. These species-specific residues can be assumed as real targets of evolution, while other substitutions tend to support them. Moreover, all these residues are parts of experimentally identified epitopes. Therefore, their substitution may affect vaccine efficacy. Higher rate of RBD maintenance than furin cleavage site was predicted. The accumulation of substitutions reinforces the probability of the multi-host circulation of the virus and emphasizes the enduring evolutionary events.


Subject(s)
SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/genetics , Amino Acid Sequence , Angiotensin-Converting Enzyme 2/chemistry , Angiotensin-Converting Enzyme 2/metabolism , Binding Sites , COVID-19/pathology , COVID-19/virology , Cluster Analysis , Humans , Markov Chains , Mutation , Protein Binding , Protein Domains/genetics , SARS-CoV-2/isolation & purification , Sequence Alignment , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/metabolism
10.
Mol Immunol ; 141: 287-296, 2022 01.
Article in English | MEDLINE | ID: covidwho-1559780

ABSTRACT

As the second wave of COVID-19 launched, various variants of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) have emerged with a dramatic global spread amongst millions of people causing unprecedented case fatalities and economic shut-downs. That initiated a necessity for developing specific diagnostics and therapeutics along with vaccines to control such a pandemic. This endeavor describes generation of murine derived recombinant single-chain fragment variable (scFv) as a monoclonal antibody (MAb) platform targeting the receptor binding domain (RBD) of Spike protein of SARS-CoV-2. A specific synthesized RBD coding sequence was cloned and expressed in Baculovirus expression system. The recombinant RBD (rRBD) was ascertained to be at the proper encoding size of ∼ 600bp and expressed protein of the molecular weight of ∼ 21KDa. Purified rRBD was proved genuinely antigenic and immunogenic, exhibiting specific reactivity to anti-SARS-CoV-2 antibody in an indirect enzyme-linked immunosorbent assay (ELISA), and inducing strong seroconversion in immunized mice. The scFv phage display library against rRBD was successfully constructed, revealing ∼ 90 % recombination frequency, and great enriching factor reaching 88 % and 25 % in polyclonal Ab-based and MAb-based ELISAs, respectively. Typically, three unique scFvs were generated, selected, purified and molecularly identified. That was manifested by their: accurate structure, close relation to the mouse immunoglobulin (Ig) superfamily, right anchored six complementarily-determining regions (CDRs) as three within variable heavy (vH) and variable light (vL) regions each, and proper configuration of the three-dimensional (3D) structure. Besides, their expression downstream in a non-suppressive amber codon of E. coli strain SS32 created a distinct protein band at an apparent molecular weight of ∼ 27KDa. Moreover, the purified scFvs showed authentic immunoreactivity and specificity to both rRBD and SARS-CoV-2 in western blot and ELISA. Accordingly, these developed scFvs platform might be a functional candidate for research, inexpensive diagnostics and therapeutics, mitigating spread of COVID-19.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Monoclonal/immunology , Antibodies, Viral/immunology , COVID-19 Serological Testing , COVID-19/diagnosis , Cell Surface Display Techniques , Epitopes/immunology , Receptors, Virus/metabolism , SARS-CoV-2/immunology , Single-Chain Antibodies/immunology , Spike Glycoprotein, Coronavirus/immunology , Amino Acid Sequence , Animals , Antibodies, Monoclonal/biosynthesis , Antibodies, Viral/blood , Antibody Specificity , Baculoviridae , COVID-19/prevention & control , Escherichia coli , Female , Genetic Vectors , Mice , Mice, Inbred BALB C , Models, Molecular , Peptide Library , Protein Conformation , Protein Domains , Recombinant Proteins/biosynthesis , Recombinant Proteins/chemistry , Recombinant Proteins/immunology , Sequence Alignment , Sequence Homology, Amino Acid , Single-Chain Antibodies/biosynthesis , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism
11.
J Comput Biol ; 28(12): 1228-1247, 2021 12.
Article in English | MEDLINE | ID: covidwho-1545879

ABSTRACT

The detrimental effect of coronavirus disease 2019 (COVID-19) pandemic has manifested itself as a global crisis. Currently, no specific treatment options are available for COVID-19, so therapeutic interventions to tackle the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection must be urgently established. Therefore, cohesive and multidimensional efforts are required to identify new therapies or investigate the efficacy of small molecules and existing drugs against SARS-CoV-2. Since the RNA-dependent RNA Polymerase (RdRP) of SARS-CoV-2 is a promising therapeutic target, this study addresses the identification of antiviral molecules that can specifically target SARS-CoV-2 RdRP. The computational approach of drug development was used to screen the antiviral molecules from two antiviral libraries (Life Chemicals [LC] and ASINEX) against RdRP. Here, we report six antiviral molecules (F3407-4105, F6523-2250, F6559-0746 from LC and BDG 33693278, BDG 33693315, LAS 34156196 from ASINEX), which show substantial interactions with key amino acid residues of the active site of SARS-CoV-2 RdRP and exhibit higher binding affinity (>7.5 kcalmol-1) than Galidesivir, an Food and Drug Administration-approved inhibitor of the same. Further, molecular dynamics simulation and Molecular Mechanics Poisson-Boltzmann Surface Area results confirmed that identified molecules with RdRP formed higher stable RdRP-inhibitor(s) complex than RdRP-Galidesvir complex. Our findings suggest that these molecules could be potential inhibitors of SARS-CoV-2 RdRP. However, further in vitro and preclinical experiments would be required to validate these potential inhibitors of SARS-CoV-2 protein.


Subject(s)
Antiviral Agents/pharmacology , COVID-19/drug therapy , Computational Chemistry/methods , Coronavirus RNA-Dependent RNA Polymerase/antagonists & inhibitors , Drug Discovery/methods , Drug Evaluation, Preclinical/methods , Pandemics , SARS-CoV-2/drug effects , Amino Acid Motifs , Amino Acid Sequence , Antiviral Agents/chemistry , Antiviral Agents/pharmacokinetics , Catalytic Domain/drug effects , Coronavirus RNA-Dependent RNA Polymerase/chemistry , Databases, Chemical , Molecular Docking Simulation , Molecular Dynamics Simulation , Molecular Structure , Protein Binding , Protein Conformation , SARS-CoV-2/enzymology , Sequence Alignment , Sequence Homology, Amino Acid , Small Molecule Libraries
12.
J Med Virol ; 93(12): 6595-6604, 2021 12.
Article in English | MEDLINE | ID: covidwho-1544310

ABSTRACT

As a kind of human betacoronavirus, SARS-CoV-2 has endangered globally public health. As of January 2021, the virus had resulted in 2,209,195 deaths. By studying the evolution trend and characteristics of 265 SARS-CoV-2 strains in the United States from January to March, it is found that the strains can be divided into six clades, USA clade-1, USA clade-2, USA clade-3, USA clade-4, USA clade-5, and USA clade-6, in which US clade-1 may be the most ancestral clade, USA clade-2 is an interim clade of USA clade-1 and USA clade-3, the other three clades have similar codon usage pattern, while USA clade-6 is the newest and most adaptable clade. Mismatch analysis and protein alignment showed that the evolution of the clades arises from some special mutations in viral proteins, which may help the strain to invade, replicate, transcribe and so on. Compared with previous research and classifications, we suggest that clade O in GISAID should not be an independent clade and Wuhan-Hu-1 (EPI_ISL_402125) should not be an ancestral reference sequence. Our study decoded the evolutionary dynamic of SARS-CoV-2 in the early stage from the United States, which give some clues to infer the current evolution trend of SARS-CoV-2 and study the function of viral mutational protein.


Subject(s)
Evolution, Molecular , SARS-CoV-2/genetics , Bayes Theorem , COVID-19/virology , Humans , Mutation/genetics , Phylogeny , Principal Component Analysis , Sequence Alignment , United States/epidemiology
13.
Genes (Basel) ; 12(11)2021 11 18.
Article in English | MEDLINE | ID: covidwho-1533884

ABSTRACT

Multiple sequence alignment (MSA) is the basis for almost all sequence comparison and molecular phylogenetic inferences. Large-scale genomic analyses are typically associated with automated progressive MSA without subsequent manual adjustment, which itself is often error-prone because of the lack of a consistent and explicit criterion. Here, I outlined several commonly encountered alignment errors that cannot be avoided by progressive MSA for nucleotide, amino acid, and codon sequences. Methods that could be automated to fix such alignment errors were then presented. I emphasized the utility of position weight matrix as a new tool for MSA refinement and illustrated its usage by refining the MSA of nucleotide and amino acid sequences. The main advantages of the position weight matrix approach include (1) its use of information from all sequences, in contrast to other commonly used methods based on pairwise alignment scores and inconsistency measures, and (2) its speedy computation, making it suitable for a large number of long viral genomic sequences.


Subject(s)
Automation, Laboratory/methods , Genomics/methods , Sequence Alignment/methods , Algorithms , Animals , Automation, Laboratory/standards , Genomics/standards , Humans , Phylogeny , Sensitivity and Specificity , Sequence Alignment/standards , Sequence Analysis, DNA/methods , Sequence Analysis, DNA/standards , Sequence Analysis, Protein/methods , Sequence Analysis, Protein/standards
14.
Biochim Biophys Acta Mol Basis Dis ; 1868(2): 166291, 2022 02 01.
Article in English | MEDLINE | ID: covidwho-1525693

ABSTRACT

OBJECTIVES: To investigate in silico the presence of nucleotide sequence complementarity between the RNA genome of Severe Acute Respiratory Syndrome CoronaVirus-2 (SARS-CoV-2) and human non-coding (nc)RNA genes. METHODS: The FASTA sequence (NC_045512.2) of each of the 11 SARS-CoV-2 isolate Wuhan-Hu-1 genes was retrieved from NCBI.nlm.nih.gov/gene and the Ensembl.org library interrogated for any base-pair match with human ncRNA genes. SARS-CoV-2 gene-matched human ncRNAs were screened for functional activity using bioinformatic analysis. Finally, associations between identified ncRNAs and human diseases were searched in GWAS databases. RESULTS: A total of 252 matches were found between the nucleotide sequence of SARS-CoV-2 genes and human ncRNAs. With the exception of two small nuclear RNAs, all of them were long non-coding (lnc)RNAs expressed mainly in testis and central nervous system under physiological conditions. The percentage of alignment ranged from 91.30% to 100% with a mean nucleotide alignment length of 17.5 ± 2.4. Thirty-three (13.09%) of them contained predicted R-loop forming sequences, but none of these intersected the complementary sequences of SARS-CoV-2. However, in 31 cases matches fell on ncRNA regulatory sites, whose adjacent coding genes are mostly involved in cancer, immunological and neurological pathways. Similarly, several polymorphic variants of detected non-coding genes have been associated with neuropsychiatric and proliferative disorders. CONCLUSION: This pivotal in silico study shows that SARS-CoV-2 genes have Watson-Crick nucleotide complementarity to human ncRNA sequences, potentially disrupting ncRNA epigenetic control of target genes. It remains to be elucidated whether this could result in the development of human disease in the long term.


Subject(s)
COVID-19/genetics , COVID-19/virology , RNA, Untranslated/genetics , SARS-CoV-2/genetics , Base Sequence , Epigenesis, Genetic , Genes, Viral , Humans , Neoplasms/genetics , RNA, Long Noncoding/genetics , Sequence Alignment , Sequence Homology, Nucleic Acid
15.
Curr Issues Mol Biol ; 43(3): 2036-2047, 2021 Nov 16.
Article in English | MEDLINE | ID: covidwho-1523887

ABSTRACT

The study of the interaction between the SARS-CoV-2 spike protein and the angiotensin-converting enzyme 2 (ACE2) receptor is key to understanding binding affinity and stability. In the present report, we sought to investigate the differences between two already sequenced genome variants (Spanish and British) of SARS-CoV-2. Methods: In silico model evaluating the homology, identity and similarity in the genome sequence and the structure and alignment of the predictive spike by computational docking methods. Results: The identity results between the Spanish and British variants of the Spike protein were 28.67%. This close correspondence in the results between the Spanish and British SARS-CoV-2 variants shows that they are very similar (99.99%). The alignment obtained results in four deletions. There were 23 nucleotide substitutions also predicted which could affect the functionality of the proteins produced from this sequence. The interaction between the binding receptor domain from the spike protein and the ACE2 receptor produces some of the mutations found and, therefore, the energy of this ligand varies. However, the estimated antigenicity of the British variant is higher than its Spanish counterpart. Conclusions: Our results indicate that minimal mutations could interfere in the infectivity of the virus due to changes in the fitness between host cell recognition and interaction proteins. In particular, the N501Y substitution, situated in the RBD of the spike of the British variant, might be the reason for its extraordinary infective potential.


Subject(s)
COVID-19/virology , SARS-CoV-2/genetics , SARS-CoV-2/physiology , Virus Internalization , Amino Acid Sequence , Angiotensin-Converting Enzyme 2/metabolism , Base Sequence , COVID-19/epidemiology , COVID-19/metabolism , COVID-19/pathology , Computational Biology , Humans , Protein Binding , SARS-CoV-2/isolation & purification , SARS-CoV-2/metabolism , Sequence Alignment , Spain/epidemiology , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , United Kingdom/epidemiology
16.
J Biol Chem ; 298(1): 101403, 2022 01.
Article in English | MEDLINE | ID: covidwho-1517316

ABSTRACT

Designed ankyrin repeat proteins (DARPins) are antibody mimetics with high and mostly unexplored potential in drug development. By using in silico analysis and a rationally guided Ala scanning, we identified position 17 of the N-terminal capping repeat to play a key role in overall protein thermostability. The melting temperature of a DARPin domain with a single full-consensus internal repeat was increased by 8 °C to 10 °C when Asp17 was replaced by Leu, Val, Ile, Met, Ala, or Thr. We then transferred the Asp17Leu mutation to various backgrounds, including clinically validated DARPin domains, such as the vascular endothelial growth factor-binding domain of the DARPin abicipar pegol. In all cases, these proteins showed improvements in the thermostability on the order of 8 °C to 16 °C, suggesting the replacement of Asp17 could be generically applicable to this drug class. Molecular dynamics simulations showed that the Asp17Leu mutation reduces electrostatic repulsion and improves van-der-Waals packing, rendering the DARPin domain less flexible and more stable. Interestingly, this beneficial Asp17Leu mutation is present in the N-terminal caps of three of the five DARPin domains of ensovibep, a SARS-CoV-2 entry inhibitor currently in clinical development, indicating this mutation could be partly responsible for the very high melting temperature (>90 °C) of this promising anti-COVID-19 drug. Overall, such N-terminal capping repeats with increased thermostability seem to be beneficial for the development of innovative drugs based on DARPins.


Subject(s)
Antiviral Agents/pharmacology , Temperature , Amino Acid Sequence , Antiviral Agents/chemistry , Antiviral Agents/therapeutic use , COVID-19/drug therapy , COVID-19/virology , Drug Development , Drug Stability , SARS-CoV-2/drug effects , Sequence Alignment
17.
Int J Mol Sci ; 22(22)2021 Nov 09.
Article in English | MEDLINE | ID: covidwho-1512384

ABSTRACT

Coronaviruses cause diseases in humans and livestock. The SARS-CoV-2 is infecting millions of human beings, with high morbidity and mortality worldwide. The main protease (Mpro) of coronavirus plays a pivotal role in viral replication and transcription, which, in theory, is an attractive drug target for antiviral drug development. It has been extensively discussed whether Xanthohumol is able to help COVID-19 patients. Here, we report that Xanthohumol, a small molecule in clinical trials from hops (Humulus lupulus), was a potent pan-inhibitor for various coronaviruses by targeting Mpro, for example, betacoronavirus SARS-CoV-2 (IC50 value of 1.53 µM), and alphacoronavirus PEDV (IC50 value of 7.51 µM). Xanthohumol inhibited Mpro activities in the enzymatical assays, while pretreatment with Xanthohumol restricted the SARS-CoV-2 and PEDV replication in Vero-E6 cells. Therefore, Xanthohumol is a potent pan-inhibitor of coronaviruses and an excellent lead compound for further drug development.


Subject(s)
3C Viral Proteases/antagonists & inhibitors , Flavonoids/chemistry , Propiophenones/chemistry , Protease Inhibitors/chemistry , SARS-CoV-2/enzymology , 3C Viral Proteases/chemistry , 3C Viral Proteases/metabolism , Alphacoronavirus/enzymology , Alphacoronavirus/physiology , Amino Acid Sequence , Animals , Binding Sites , Biological Products/chemistry , Biological Products/metabolism , Biological Products/pharmacology , Biological Products/therapeutic use , COVID-19/drug therapy , COVID-19/virology , Catalytic Domain , Chlorocebus aethiops , Coronavirus/enzymology , Coronavirus/physiology , Flavonoids/metabolism , Flavonoids/pharmacology , Flavonoids/therapeutic use , Humans , Molecular Docking Simulation , Propiophenones/metabolism , Propiophenones/pharmacology , Propiophenones/therapeutic use , Protease Inhibitors/metabolism , Protease Inhibitors/pharmacology , Protease Inhibitors/therapeutic use , SARS-CoV-2/isolation & purification , Sequence Alignment , Vero Cells , Virus Replication/drug effects
18.
Sci Rep ; 11(1): 22042, 2021 11 11.
Article in English | MEDLINE | ID: covidwho-1510622

ABSTRACT

The mutation of SARS-CoV-2 influences viral function as residue replacements affect both physiochemical properties and folding conformations. Although a large amount of data on SARS-CoV-2 is available, the investigation of how viral functions change in response to mutations is hampered by a lack of effective structural analysis. Here, we exploit the advances of protein structure fingerprint technology to study the folding conformational changes induced by mutations. With integration of both protein sequences and folding conformations, the structures are aligned for SARS-CoV to SARS-CoV-2, including Alpha variant (lineage B.1.1.7) and Delta variant (lineage B.1.617.2). The results showed that the virus evolution with change in mutational positions and physicochemical properties increased the affinity between spike protein and ACE2, which plays a critical role in coronavirus entry into human cells. Additionally, these structural variations impact vaccine effectiveness and drug function over the course of SARS-CoV-2 evolution. The analysis of structural variations revealed how the coronavirus has gradually evolved in both structure and function and how the SARS-CoV-2 variants have contributed to more severe acute disease worldwide.


Subject(s)
COVID-19/virology , Evolution, Molecular , Mutation , SARS-CoV-2/genetics , Amino Acid Sequence , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/metabolism , Humans , Models, Molecular , Protein Conformation , Protein Folding , Protein Interaction Maps , Protein Multimerization , SARS Virus/chemistry , SARS Virus/genetics , SARS Virus/metabolism , SARS-CoV-2/chemistry , SARS-CoV-2/metabolism , Sequence Alignment , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism
19.
Nat Commun ; 12(1): 6563, 2021 11 09.
Article in English | MEDLINE | ID: covidwho-1510593

ABSTRACT

Knowledge of the origin and reservoir of the coronavirus responsible for the ongoing COVID-19 pandemic is still fragmentary. To date, the closest relatives to SARS-CoV-2 have been detected in Rhinolophus bats sampled in the Yunnan province, China. Here we describe the identification of SARS-CoV-2 related coronaviruses in two Rhinolophus shameli bats sampled in Cambodia in 2010. Metagenomic sequencing identifies nearly identical viruses sharing 92.6% nucleotide identity with SARS-CoV-2. Most genomic regions are closely related to SARS-CoV-2, with the exception of a region of the spike, which is not compatible with human ACE2-mediated entry. The discovery of these viruses in a bat species not found in China indicates that SARS-CoV-2 related viruses have a much wider geographic distribution than previously reported, and suggests that Southeast Asia represents a key area to consider for future surveillance for coronaviruses.


Subject(s)
COVID-19/virology , Chiroptera/virology , SARS-CoV-2/genetics , Amino Acid Sequence , Animals , COVID-19/epidemiology , COVID-19/metabolism , Cambodia/epidemiology , Evolution, Molecular , Genome, Viral , Phylogeny , SARS-CoV-2/isolation & purification , SARS-CoV-2/metabolism , Sequence Alignment , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism
20.
Infect Genet Evol ; 96: 105106, 2021 12.
Article in English | MEDLINE | ID: covidwho-1506080

ABSTRACT

Coronaviruses (especially SARS-CoV-2) are characterized by rapid mutation and wide spread. As these characteristics easily lead to global pandemics, studying the evolutionary relationship between viruses is essential for clinical diagnosis. DNA sequencing has played an important role in evolutionary analysis. Recent alignment-free methods can overcome the problems of traditional alignment-based methods, which consume both time and space. This paper proposes a novel alignment-free method called the correlation coefficient feature vector (CCFV), which defines a correlation measure of the L-step delay of a nucleotide location from its location in the original DNA sequence. The numerical feature is a 16×L-dimensional numerical vector describing the distribution characteristics of the nucleotide positions in a DNA sequence. The proposed L-step delay correlation measure is interestingly related to some types of L+1 spaced mers. Unlike traditional gene comparison, our method avoids the computational complexity of multiple sequence alignment, and hence improves the speed of sequence comparison. Our method is applied to evolutionary analysis of the common human viruses including SARS-CoV-2, Dengue virus, Hepatitis B virus, and human rhinovirus and achieves the same or even better results than alignment-based methods. Especially for SARS-CoV-2, our method also confirms that bats are potential intermediate hosts of SARS-CoV-2.


Subject(s)
Genome, Viral/genetics , Phylogeny , Sequence Analysis, DNA/methods , Coronavirus/genetics , Dengue Virus/genetics , Hepatitis B/genetics , Humans , Models, Genetic , Rhinovirus/genetics , SARS-CoV-2/genetics , Sequence Alignment
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