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1.
Int J Mol Sci ; 23(6)2022 Mar 15.
Article in English | MEDLINE | ID: covidwho-1742492

ABSTRACT

SARS-CoV-2 variants surveillance is a worldwide task that has been approached with techniques such as Next Generation Sequencing (NGS); however, this technology is not widely available in developing countries because of the lack of equipment and limited funding in science. An option is to deploy a RT-qPCR screening test which aids in the analysis of a higher number of samples, in a shorter time and at a lower cost. In this study, variants present in samples positive for SARS-CoV-2 were identified with a RT-qPCR mutation screening kit and were later confirmed by NGS. A sample with an abnormal result was found with the screening test, suggesting the simultaneous presence of two viral populations with different mutations. The DRAGEN Lineage analysis identified the Delta variant, but there was no information about the other three mutations previously detected. When the sequenced data was deeply analyzed, there were reads with differential mutation patterns, that could be identified and classified in terms of relative abundance, whereas only the dominant population was reported by DRAGEN software. Since most of the software developed to analyze SARS-CoV-2 sequences was aimed at obtaining the consensus sequence quickly, the information about viral populations within a sample is scarce. Here, we present a faster and deeper SARS-CoV-2 surveillance method, from RT-qPCR screening to NGS analysis.


Subject(s)
COVID-19/diagnosis , DNA Mutational Analysis/methods , Genome, Viral/genetics , Mutation , Reverse Transcriptase Polymerase Chain Reaction/methods , SARS-CoV-2/genetics , COVID-19/epidemiology , COVID-19/virology , High-Throughput Nucleotide Sequencing/methods , Humans , Pandemics/prevention & control , Reproducibility of Results , SARS-CoV-2/physiology , Sensitivity and Specificity
2.
PLoS One ; 17(2): e0264201, 2022.
Article in English | MEDLINE | ID: covidwho-1714776

ABSTRACT

Activating mutations in EGFR predict benefit from tyrosine kinase inhibitor therapy for patients with advanced non-small cell lung cancer. Directing patients to appropriate therapy depends on accurate and timely EGFR assessment in the molecular pathology laboratory. This article describes the analytical design, performance characteristics, and clinical implementation of an assay for the rapid detection of EGFR L858R and exon 19 deletion mutations. A droplet digital polymerase chain reaction (ddPCR) assay was implemented with probe hydrolysis-dependent signal detection. A mutation-specific probe was used to detect EGFR L858R. A loss of signal design was used to detect EGFR exon 19 deletion mutations. Analytical sensitivity was dependent on DNA input and was as low as 0.01% variant allele fraction for the EGFR L858R assay and 0.1% variant allele fraction for the EGFR exon 19 deletion assay. Correlation of 20 clinical specimens tested by ddPCR and next generation sequencing showed 100% concordance. ddPCR showed 53% clinical sensitivity in the detection of EGFR mutations in plasma cell-free DNA from patients with lung cancer. The median clinical turnaround time was 5 days for ddPCR compared to 13 days for next generation sequencing. The findings show that ddPCR is an accurate and rapid method for detecting EGFR mutations in patients with non-small cell lung cancer.


Subject(s)
Carcinoma, Non-Small-Cell Lung/genetics , DNA Mutational Analysis/methods , Lung Neoplasms/genetics , Polymerase Chain Reaction/methods , ErbB Receptors/genetics , Humans , Mutation , Sensitivity and Specificity
3.
Brief Bioinform ; 23(2)2022 03 10.
Article in English | MEDLINE | ID: covidwho-1639367

ABSTRACT

Genomic epidemiology is important to study the COVID-19 pandemic, and more than two million severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) genomic sequences were deposited into public databases. However, the exponential increase of sequences invokes unprecedented bioinformatic challenges. Here, we present the Coronavirus GenBrowser (CGB) based on a highly efficient analysis framework and a node-picking rendering strategy. In total, 1,002,739 high-quality genomic sequences with the transmission-related metadata were analyzed and visualized. The size of the core data file is only 12.20 MB, highly efficient for clean data sharing. Quick visualization modules and rich interactive operations are provided to explore the annotated SARS-CoV-2 evolutionary tree. CGB binary nomenclature is proposed to name each internal lineage. The pre-analyzed data can be filtered out according to the user-defined criteria to explore the transmission of SARS-CoV-2. Different evolutionary analyses can also be easily performed, such as the detection of accelerated evolution and ongoing positive selection. Moreover, the 75 genomic spots conserved in SARS-CoV-2 but non-conserved in other coronaviruses were identified, which may indicate the functional elements specifically important for SARS-CoV-2. The CGB was written in Java and JavaScript. It not only enables users who have no programming skills to analyze millions of genomic sequences, but also offers a panoramic vision of the transmission and evolution of SARS-CoV-2.


Subject(s)
COVID-19/epidemiology , COVID-19/virology , Public Health Surveillance/methods , SARS-CoV-2/genetics , Software , Web Browser , Computational Biology/methods , DNA Mutational Analysis , Databases, Genetic , Genome, Viral , Genomics , Humans , Molecular Epidemiology/methods , Molecular Sequence Annotation , Mutation
4.
Proc Natl Acad Sci U S A ; 119(4)2022 01 25.
Article in English | MEDLINE | ID: covidwho-1621333

ABSTRACT

The emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a major concern given their potential impact on the transmissibility and pathogenicity of the virus as well as the efficacy of therapeutic interventions. Here, we predict the mutability of all positions in SARS-CoV-2 protein domains to forecast the appearance of unseen variants. Using sequence data from other coronaviruses, preexisting to SARS-CoV-2, we build statistical models that not only capture amino acid conservation but also more complex patterns resulting from epistasis. We show that these models are notably superior to conservation profiles in estimating the already observable SARS-CoV-2 variability. In the receptor binding domain of the spike protein, we observe that the predicted mutability correlates well with experimental measures of protein stability and that both are reliable mutability predictors (receiver operating characteristic areas under the curve ∼0.8). Most interestingly, we observe an increasing agreement between our model and the observed variability as more data become available over time, proving the anticipatory capacity of our model. When combined with data concerning the immune response, our approach identifies positions where current variants of concern are highly overrepresented. These results could assist studies on viral evolution and future viral outbreaks and, in particular, guide the exploration and anticipation of potentially harmful future SARS-CoV-2 variants.


Subject(s)
COVID-19/virology , Epistasis, Genetic , Epitopes , Mutation , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Viral Proteins/chemistry , Algorithms , Area Under Curve , Computational Biology/methods , DNA Mutational Analysis , Databases, Protein , Deep Learning , Epitopes/chemistry , Genome, Viral , Humans , Models, Statistical , Mutagenesis , Probability , Protein Domains , ROC Curve
5.
Sci Rep ; 11(1): 21084, 2021 10 26.
Article in English | MEDLINE | ID: covidwho-1493213

ABSTRACT

In contrast to the conventional approach of directly comparing genomic sequences using sequence alignment tools, we propose a computational approach that performs comparisons between sequence generators. These sequence generators are learned via a data-driven approach that empirically computes the state machine generating the genomic sequence of interest. As the state machine based generator of the sequence is independent of the sequence length, it provides us with an efficient method to compute the statistical distance between large sets of genomic sequences. Moreover, our technique provides a fast and efficient method to cluster large datasets of genomic sequences, characterize their temporal and spatial evolution in a continuous manner, get insights into the locality sensitive information about the sequences without any need for alignment. Furthermore, we show that the technique can be used to detect local regions with mutation activity, which can then be applied to aid alignment techniques for the fast discovery of mutations. To demonstrate the efficacy of our technique on real genomic data, we cluster different strains of SARS-CoV-2 viral sequences, characterize their evolution and identify regions of the viral sequence with mutations.


Subject(s)
COVID-19/virology , Computational Biology/methods , Genomics , Mutation , SARS-CoV-2/genetics , Algorithms , Cluster Analysis , DNA Mutational Analysis , Genome, Viral , Humans , Machine Learning , Models, Theoretical , Probability , Stochastic Processes
6.
Cell Rep ; 37(3): 109841, 2021 10 19.
Article in English | MEDLINE | ID: covidwho-1439922

ABSTRACT

Nonstructural protein 1 (nsp1) is a coronavirus (CoV) virulence factor that restricts cellular gene expression by inhibiting translation through blocking the mRNA entry channel of the 40S ribosomal subunit and by promoting mRNA degradation. We perform a detailed structure-guided mutational analysis of severe acute respiratory syndrome (SARS)-CoV-2 nsp1, revealing insights into how it coordinates these activities against host but not viral mRNA. We find that residues in the N-terminal and central regions of nsp1 not involved in docking into the 40S mRNA entry channel nonetheless stabilize its association with the ribosome and mRNA, both enhancing its restriction of host gene expression and enabling mRNA containing the SARS-CoV-2 leader sequence to escape translational repression. These data support a model in which viral mRNA binding functionally alters the association of nsp1 with the ribosome, which has implications for drug targeting and understanding how engineered or emerging mutations in SARS-CoV-2 nsp1 could attenuate the virus.


Subject(s)
COVID-19/genetics , Gene Expression Regulation, Viral , SARS-CoV-2/genetics , Viral Nonstructural Proteins/metabolism , Anisotropy , COVID-19/immunology , DNA Mutational Analysis , Female , Gene Expression Profiling , Green Fluorescent Proteins/metabolism , HEK293 Cells , Humans , Kinetics , Mutation , Phenotype , Point Mutation , Protein Biosynthesis , Protein Domains , RNA Stability , Ribosome Subunits, Small, Eukaryotic/metabolism , Ribosomes/metabolism
7.
STAR Protoc ; 2(4): 100869, 2021 12 17.
Article in English | MEDLINE | ID: covidwho-1433914

ABSTRACT

Here, we describe a protocol to identify escape mutants on the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) receptor-binding domain (RBD) using a yeast screen combined with deep mutational scanning. Over 90% of all potential single S RBD escape mutants can be identified for monoclonal antibodies that directly compete with angiotensin-converting enzyme 2 for binding. Six to 10 antibodies can be assessed in parallel. This approach has been shown to determine escape mutants that are consistent with more laborious SARS-CoV-2 pseudoneutralization assays. For complete details on the use and execution of this protocol, please refer to Francino-Urdaniz et al. (2021).


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , COVID-19/genetics , DNA Mutational Analysis/methods , Mutation , SARS-CoV-2/genetics , Saccharomyces cerevisiae/metabolism , Spike Glycoprotein, Coronavirus/genetics , Angiotensin-Converting Enzyme 2/metabolism , Binding Sites , COVID-19/metabolism , COVID-19/virology , Humans , Saccharomyces cerevisiae/genetics , Spike Glycoprotein, Coronavirus/metabolism
9.
Epidemiol Infect ; 149: e103, 2021 04 28.
Article in English | MEDLINE | ID: covidwho-1279757

ABSTRACT

Although vaccines have become available, emergence and rapid transmission of new variants have added new paradigm in the coronavirus disease-2019 (COVID-19) pandemic. Weather, population and host immunity have been detected as the regulatory elements of COVID-19. This study aims to investigate the effects of weather, population and host factors on the outcome of COVID-19 and mutation frequency in Japan. Data were collected during January 2020 to February 2021. About 92% isolates were form GR clades. Variants 501Y.V1 (53%) and 452R.V1 (24%) were most prevalent in Japan. The strongest correlation was detected between fatalities and population density (rs = 0.81) followed by total population (rs = 0.72). Relative humidity had the highest correlation (rs = -0.71) with the case fatality rate. Cluster mutations namely N501Y (45%), E484K (30%), N439K (16%), K417N (6%) and T478I (3%) at spike protein have increased during January to February 2021. Above 90% fatality was detected in patients aged >60 years. The ratio of male to female patients of COVID-19 was 1.35:1. This study will help to understand the seasonality of COVID-19 and impact of weather on the outcome which will add knowledge to reduce the health burden of COVID-19 by the international organisations and policy makers.


Subject(s)
COVID-19/epidemiology , Population Density , SARS-CoV-2 , Weather , DNA Mutational Analysis , Genome, Viral , Humans , Japan/epidemiology , Mutation , SARS-CoV-2/genetics , Spatial Analysis , Time Factors
10.
J Med Virol ; 93(7): 4382-4391, 2021 Jul.
Article in English | MEDLINE | ID: covidwho-1263102

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic has spread around the globe very rapidly. Previously, the evolution pattern and similarity among the COVID-19 causative organism severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and causative organisms of other similar infections have been determined using a single type of genetic marker in different studies. Herein, the SARS-CoV-2 and related ß coronaviruses Middle East respiratory syndrome coronavirus (MERS-CoV), SARS-CoV,  bat coronavirus (BAT-CoV) were comprehensively analyzed using a custom-built pipeline that employed phylogenetic approaches based on multiple types of genetic markers including the whole genome sequences, mutations in nucleotide sequences, mutations in protein sequences, and microsatellites. The whole-genome sequence-based phylogeny revealed that the strains of SARS-CoV-2 are more similar to the BAT-CoV strains. The mutational analysis showed that on average MERS-CoV and BAT-CoV genomes differed at 134.21 and 136.72 sites, respectively, whereas the SARS-CoV genome differed at 26.64 sites from the reference genome of SARS-CoV-2. Furthermore, the microsatellite analysis highlighted a relatively higher number of average microsatellites for MERS-CoV and SARS-CoV-2 (106.8 and 107, respectively), and a lower number for SARS-CoV and BAT-CoV (95.8 and 98.5, respectively). Collectively, the analysis of multiple genetic markers of selected ß viral genomes revealed that the newly born SARS-COV-2 is closely related to BAT-CoV, whereas, MERS-CoV is more distinct from the SARS-CoV-2 than BAT-CoV and SARS-CoV.


Subject(s)
Alphacoronavirus/genetics , Genome, Viral/genetics , Microsatellite Repeats/genetics , Middle East Respiratory Syndrome Coronavirus/genetics , SARS Virus/genetics , SARS-CoV-2/genetics , Animals , Base Sequence/genetics , Chiroptera/virology , DNA Mutational Analysis , Genetic Markers/genetics , Genetic Variation/genetics , Humans , Phylogeny , Sequence Alignment , Sequence Homology, Nucleic Acid , Whole Genome Sequencing
11.
Viruses ; 13(6)2021 05 29.
Article in English | MEDLINE | ID: covidwho-1256668

ABSTRACT

The COVID-19 pandemic, which began in Wuhan (Hubei, China), has been ongoing for about a year and a half. An unprecedented number of people around the world have been infected with SARS-CoV-2, the etiological agent of COVID-19. Despite the fact that the mortality rate for COVID-19 is relatively low, the total number of deaths has currently already reached more than three million and continues to increase due to high incidence. Since the beginning of the pandemic, a large number of sequences have been obtained and many genetic variants have been identified. Some of them bear significant mutations that affect biological properties of the virus. These genetic variants, currently Variants of Concern (VoC), include the so-called United Kingdom variant (20I/501Y), the Brazilian variant (20J/501Y.V3), and the South African variant (20H/501Y.V2). We describe here a novel SARS-CoV-2 variant with distinct spike protein mutations, first obtained at the end of January 2021 in northwest Russia. Therefore, it is necessary to pay attention to the dynamics of its spread among patients with COVID-19, as well as to study in detail its biological properties.


Subject(s)
SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , DNA Mutational Analysis , DNA, Complementary , Genome, Viral , Humans , Models, Molecular , Mutation , Phylogeny , Protein Conformation , Russia , SARS-CoV-2/classification , SARS-CoV-2/isolation & purification , Spike Glycoprotein, Coronavirus/chemistry
12.
Nature ; 592(7854): 438-443, 2021 04.
Article in English | MEDLINE | ID: covidwho-1164876

ABSTRACT

Continued uncontrolled transmission of SARS-CoV-2 in many parts of the world is creating conditions for substantial evolutionary changes to the virus1,2. Here we describe a newly arisen lineage of SARS-CoV-2 (designated 501Y.V2; also known as B.1.351 or 20H) that is defined by eight mutations in the spike protein, including three substitutions (K417N, E484K and N501Y) at residues in its receptor-binding domain that may have functional importance3-5. This lineage was identified in South Africa after the first wave of the epidemic in a severely affected metropolitan area (Nelson Mandela Bay) that is located on the coast of the Eastern Cape province. This lineage spread rapidly, and became dominant in Eastern Cape, Western Cape and KwaZulu-Natal provinces within weeks. Although the full import of the mutations is yet to be determined, the genomic data-which show rapid expansion and displacement of other lineages in several regions-suggest that this lineage is associated with a selection advantage that most plausibly results from increased transmissibility or immune escape6-8.


Subject(s)
COVID-19/virology , Mutation , Phylogeny , Phylogeography , SARS-CoV-2/genetics , SARS-CoV-2/isolation & purification , COVID-19/epidemiology , COVID-19/immunology , COVID-19/transmission , DNA Mutational Analysis , Evolution, Molecular , Genetic Fitness , Humans , Immune Evasion , Models, Molecular , SARS-CoV-2/immunology , SARS-CoV-2/pathogenicity , Selection, Genetic , South Africa/epidemiology , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Spike Glycoprotein, Coronavirus/metabolism , Time Factors
13.
J Med Virol ; 93(7): 4382-4391, 2021 Jul.
Article in English | MEDLINE | ID: covidwho-1156882

ABSTRACT

The coronavirus disease 2019 (COVID-19) pandemic has spread around the globe very rapidly. Previously, the evolution pattern and similarity among the COVID-19 causative organism severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and causative organisms of other similar infections have been determined using a single type of genetic marker in different studies. Herein, the SARS-CoV-2 and related ß coronaviruses Middle East respiratory syndrome coronavirus (MERS-CoV), SARS-CoV,  bat coronavirus (BAT-CoV) were comprehensively analyzed using a custom-built pipeline that employed phylogenetic approaches based on multiple types of genetic markers including the whole genome sequences, mutations in nucleotide sequences, mutations in protein sequences, and microsatellites. The whole-genome sequence-based phylogeny revealed that the strains of SARS-CoV-2 are more similar to the BAT-CoV strains. The mutational analysis showed that on average MERS-CoV and BAT-CoV genomes differed at 134.21 and 136.72 sites, respectively, whereas the SARS-CoV genome differed at 26.64 sites from the reference genome of SARS-CoV-2. Furthermore, the microsatellite analysis highlighted a relatively higher number of average microsatellites for MERS-CoV and SARS-CoV-2 (106.8 and 107, respectively), and a lower number for SARS-CoV and BAT-CoV (95.8 and 98.5, respectively). Collectively, the analysis of multiple genetic markers of selected ß viral genomes revealed that the newly born SARS-COV-2 is closely related to BAT-CoV, whereas, MERS-CoV is more distinct from the SARS-CoV-2 than BAT-CoV and SARS-CoV.


Subject(s)
Alphacoronavirus/genetics , Genome, Viral/genetics , Microsatellite Repeats/genetics , Middle East Respiratory Syndrome Coronavirus/genetics , SARS Virus/genetics , SARS-CoV-2/genetics , Animals , Base Sequence/genetics , Chiroptera/virology , DNA Mutational Analysis , Genetic Markers/genetics , Genetic Variation/genetics , Humans , Phylogeny , Sequence Alignment , Sequence Homology, Nucleic Acid , Whole Genome Sequencing
14.
Exp Mol Pathol ; 120: 104634, 2021 06.
Article in English | MEDLINE | ID: covidwho-1152690

ABSTRACT

Lung and colorectal cancers (CRC) have two of the highest mortality rates among all cancer types, and their occurrence and the need for personalized diagnostics and subsequent therapy were not influenced by the COVID-19 pandemics. However, due to the disruption of established delivery chains, standard assays for in vitro diagnostics of those cancers were temporarily not available, forcing us to implement alternative testing methods that enabled at least basic therapy decision making. For this reason, we evaluated rapid testing on the Biocartis Idylla™ platform (Biocartis, Mechelen, Belgium) for four important genes commonly mutated in lung and colorectal cancers, namely EGFR, NRAS, KRAS, and BRAF. Clinical specimens from which the mutation status has previously been determined using Next Generation Sequencing (NGS), were retested to determine whether Idylla™ can offer accurate results. To compare the results, the sensitivity, specificity, positive predictive values (PPV) and negative predictive values (NPV) are calculated for each of the mutation types and then combined to determine the values of the Idylla™ system in total, while setting NGS as the gold-standard basis the assays were compared with. Idylla testing thereby displayed acceptable sensitivity and specificity and delivered reliable results for initial therapy decisions.


Subject(s)
DNA Mutational Analysis/methods , GTP Phosphohydrolases/genetics , High-Throughput Nucleotide Sequencing/methods , Membrane Proteins/genetics , Mutation , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins p21(ras)/genetics , COVID-19/epidemiology , COVID-19/prevention & control , COVID-19/virology , Colorectal Neoplasms/diagnosis , Colorectal Neoplasms/genetics , ErbB Receptors/genetics , Humans , Lung Neoplasms/diagnosis , Lung Neoplasms/genetics , Pandemics , Reproducibility of Results , SARS-CoV-2/isolation & purification , SARS-CoV-2/physiology , Sensitivity and Specificity
16.
Expert Rev Mol Diagn ; 21(1): 101-107, 2021 01.
Article in English | MEDLINE | ID: covidwho-962282

ABSTRACT

Background: The SARS-CoV-2 pandemic introduced a global distraction effect in cancer patients' care. The aim of this study was to explore the effect of the pandemic on the largest molecular diagnostics center for cancer patients and high-risk individuals in Serbia.Research design and methods: EGFR, KRAS/NRAS, BRAF, and BRCA1/2 mutation testing were performed by qPCR and NGS. NGS was used for panel testing of hereditary breast/ovarian cancer and cancers associated with Lynch syndrome. The analytical output during the state of emergency (SoE) was compared to the period before and after the outbreak using one-way ANOVA. Statistical significance was set at p < 0.05.Results: A 38% reduction in the number of analysis was detected during the SoE. After the SoE, a 19% reduction was noted compared to SoE and 50% compared to the period before the SoE (p = 0.038). Three of the 48 scheduled appointments for pretest genetic counseling were carried out during the SoE, but the number of NGS tests increased by 50%.Conclusions: The SARS-CoV-2 pandemic had a profound negative effect on the diagnostic output of our centralized molecular diagnostics center. The only positive effect was shortening of waiting lists for hereditary cancer patients and high-risk individuals.


Subject(s)
Breast Neoplasms/diagnosis , Colorectal Neoplasms, Hereditary Nonpolyposis/diagnosis , Mutation , Ovarian Neoplasms/diagnosis , BRCA1 Protein/genetics , BRCA2 Protein/genetics , Breast Neoplasms/genetics , COVID-19 , Colorectal Neoplasms, Hereditary Nonpolyposis/genetics , DNA Mutational Analysis , ErbB Receptors/genetics , Female , GTP Phosphohydrolases/genetics , Genetic Counseling , Genetic Predisposition to Disease , Humans , Liquid Biopsy , Membrane Proteins/genetics , Ovarian Neoplasms/genetics , Pandemics , Pathology, Molecular , Pharmacogenetics , Proto-Oncogene Proteins B-raf/genetics , Proto-Oncogene Proteins p21(ras)/genetics , Serbia/epidemiology
17.
Proteins ; 89(4): 389-398, 2021 04.
Article in English | MEDLINE | ID: covidwho-932034

ABSTRACT

Coronaviruses are responsible for several epidemics, including the 2002 SARS, 2012 MERS, and COVID-19. The emergence of recent COVID-19 pandemic due to SARS-CoV-2 virus in December 2019 has resulted in considerable research efforts to design antiviral drugs and other therapeutics against coronaviruses. In this context, it is crucial to understand the biophysical and structural features of the major proteins that are involved in virus-host interactions. In the current study, we have compared spike proteins from three strains of coronaviruses NL63, SARS-CoV, and SARS-CoV, known to bind human angiotensin-converting enzyme 2 (ACE2), in terms of sequence/structure conservation, hydrophobic cluster formation and importance of binding site residues. The study reveals that the severity of coronavirus strains correlates positively with the interaction area, surrounding hydrophobicity and interaction energy and inversely correlate with the flexibility of the binding interface. Also, we identify the conserved residues in the binding interface of spike proteins in all three strains. The systematic point mutations show that these conserved residues in the respective strains are evolutionarily favored at their respective positions. The similarities and differences in the spike proteins of the three viruses indicated in this study may help researchers to deeply understand the structural behavior, binding site properties and etiology of ACE2 binding, accelerating the screening of potential lead molecules and the development/repurposing of therapeutic drugs.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , COVID-19/virology , Coronavirus NL63, Human , SARS Virus , SARS-CoV-2 , Spike Glycoprotein, Coronavirus/chemistry , Antiviral Agents/pharmacology , Coronavirus Infections/virology , DNA Mutational Analysis , Humans , Hydrophobic and Hydrophilic Interactions , Ligands , Models, Statistical , Mutation , Protein Binding , Protein Conformation , Species Specificity , Spike Glycoprotein, Coronavirus/genetics
18.
Proc Natl Acad Sci U S A ; 117(48): 30610-30618, 2020 12 01.
Article in English | MEDLINE | ID: covidwho-922309

ABSTRACT

Peptide binding to major histocompatibility complexes (MHCs) is a central component of the immune system, and understanding the mechanism behind stable peptide-MHC binding will aid the development of immunotherapies. While MHC binding is mostly influenced by the identity of the so-called anchor positions of the peptide, secondary interactions from nonanchor positions are known to play a role in complex stability. However, current MHC-binding prediction methods lack an analysis of the major conformational states and might underestimate the impact of secondary interactions. In this work, we present an atomically detailed analysis of peptide-MHC binding that can reveal the contributions of any interaction toward stability. We propose a simulation framework that uses both umbrella sampling and adaptive sampling to generate a Markov state model (MSM) for a coronavirus-derived peptide (QFKDNVILL), bound to one of the most prevalent MHC receptors in humans (HLA-A24:02). While our model reaffirms the importance of the anchor positions of the peptide in establishing stable interactions, our model also reveals the underestimated importance of position 4 (p4), a nonanchor position. We confirmed our results by simulating the impact of specific peptide mutations and validated these predictions through competitive binding assays. By comparing the MSM of the wild-type system with those of the D4A and D4P mutations, our modeling reveals stark differences in unbinding pathways. The analysis presented here can be applied to any peptide-MHC complex of interest with a structural model as input, representing an important step toward comprehensive modeling of the MHC class I pathway.


Subject(s)
Major Histocompatibility Complex , Markov Chains , Models, Molecular , Peptides/metabolism , Alanine/genetics , Binding, Competitive , Computer Simulation , DNA Mutational Analysis , Mutation/genetics , Proline/metabolism , Protein Binding
19.
Int J Biol Sci ; 16(15): 3028-3036, 2020.
Article in English | MEDLINE | ID: covidwho-874840

ABSTRACT

Coronavirus disease 2019 (COVID-19) caused by SARS-CoV-2, with acute respiratory failure as the most significant symptom, has led to a global pandemic. Angiotensin-converting enzyme 2 (ACE2) is considered as the most important receptor of SARS-CoV-2 and wildly expressed in human tissues. Whereas, the extremely low expression of ACE2 in lung could hardly interpret the severe symptom of pneumonia in COVID-19 patients. Here we profiled two SARS-CoV-2 infection related genes, the transmembrane serine protease 2 (TMPRSS2) and the interferon-inducible transmembrane protein 3 (IFITM3), in human tissues and organs. Consistent with the expression and distribution of ACE2, TMPRSS2 was also highly expressed in digestive, urinary and reproductive systems, but low expressed in lung. Notably, the anti-virus protein IFITM3 also expressed much lower in lung than other tissues, which might be related to the severe lung symptoms of COVID-19. In addition, the low expression of IFITM3 in immune cells suggested that SARS-CoV-2 might attack lymphocytes and induce the cytokine release syndrome (CRS). Furthermore, cancer patients were considered as more susceptible to SARS-CoV-2 infection. Our data supposed that fourteen types of tumors might have different susceptibility to the virus according to ACE2, TMPRSS2 and IFITM3 expression patterns. Interestingly the prognosis of six types of cancers including breast carcinoma (BRCA), lung adenocarcinoma (LUAD), uterine corpus endometrial carcinoma (UCEC), renal clear cell carcinoma (KIRC), prostate adenocarcinoma (PRAD), and hepatocellular carcinoma (LIHC) were closely related to these gene expressions. Our study explored the expression and distribution profiles of two potential novel molecules that might participate in SARS-CoV-2 infection and involved in immunity, which may provide a functional basis for preventing infection of SARS-CoV-2.


Subject(s)
Gene Expression Regulation, Neoplastic , Membrane Proteins/physiology , Neoplasms/metabolism , RNA-Binding Proteins/physiology , Receptors, Virus/physiology , Serine Endopeptidases/physiology , Angiotensin-Converting Enzyme 2 , Betacoronavirus , COVID-19 , Coronavirus Infections/genetics , Coronavirus Infections/metabolism , DNA Mutational Analysis , Gene Expression Regulation , Healthy Volunteers , Humans , Membrane Proteins/genetics , Neoplasms/diagnosis , Neoplasms/genetics , Pandemics , Peptidyl-Dipeptidase A/metabolism , Pneumonia, Viral/genetics , Pneumonia, Viral/metabolism , Prognosis , RNA-Binding Proteins/genetics , Receptors, Virus/genetics , SARS-CoV-2 , Serine Endopeptidases/genetics , Tissue Distribution
20.
Mol Biol Evol ; 38(2): 575-588, 2021 01 23.
Article in English | MEDLINE | ID: covidwho-799461

ABSTRACT

RNA viruses are responsible for some of the worst pandemics known to mankind, including outbreaks of Influenza, Ebola, and COVID-19. One major challenge in tackling RNA viruses is the fact they are extremely genetically diverse. Nevertheless, they share common features that include their dependence on host cells for replication, and high mutation rates. We set out to search for shared evolutionary characteristics that may aid in gaining a broader understanding of RNA virus evolution, and constructed a phylogeny-based data set spanning thousands of sequences from diverse single-stranded RNA viruses of animals. Strikingly, we found that the vast majority of these viruses have a skewed nucleotide composition, manifested as adenine rich (A-rich) coding sequences. In order to test whether A-richness is driven by selection or by biased mutation processes, we harnessed the effects of incomplete purifying selection at the tips of virus phylogenies. Our results revealed consistent mutational biases toward U rather than A in genomes of all viruses. In +ssRNA viruses, we found that this bias is compensated by selection against U and selection for A, which leads to A-rich genomes. In -ssRNA viruses, the genomic mutational bias toward U on the negative strand manifests as A-rich coding sequences, on the positive strand. We investigated possible reasons for the advantage of A-rich sequences including weakened RNA secondary structures, codon usage bias, and selection for a particular amino acid composition, and conclude that host immune pressures may have led to similar biases in coding sequence composition across very divergent RNA viruses.


Subject(s)
Mutation , RNA Viruses/genetics , RNA, Viral/genetics , Selection, Genetic , Animals , Codon , DNA Mutational Analysis , Databases, Factual , Evolution, Molecular , Genome, Viral , Humans , Nucleotides , Phylogeny , SARS-CoV-2/genetics
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