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The regions were highly conserved in severe acute respiratory syndrome-CoV-2, unlike other severe acute respiratory syndrome-CoVs

Spike protein binding prediction with neutralizing antibodies of SARS-CoV-2

biorxiv, pp.1-22, (2020)

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摘要

Coronavirus disease 2019 (COVID-19) is a new emerging human infectious disease caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2, also previously known as 2019-nCoV), originated in Wuhan seafood and animal market, China. Since December 2019, more than 69,000 cases of COVID-19 have been confirmed in China and quickly sp...更多

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简介
  • Coronaviridae is a family of enveloped viruses which have a single strand, positive-stranded RNA genome and classified into four genera: ɑ, β, γ, and δ.
  • Coronavirus (CoV) has been identified in human and animals including bats, camels, zpigs, cats, and mice.
  • The viruses usually cause mild to moderate upper-respiratory tract illnesses in human [1].
  • Severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV).
  • Middle East respiratory syndrome (MERS) coronavirus (MERS-CoV), caused severe epidemics during the last two decades.
  • SARS-CoV emerged in November 2002 in Guangdong province, China and affected 29 countries.
  • The epidemic of SARS-CoV resulted in 8,096 human infections and 774 deaths (9.6% fatality rate) by
重点内容
  • Coronaviridae is a family of enveloped viruses which have a single strand, positive-stranded RNA genome and classified into four genera: ɑ, β, γ, and δ
  • To comparing of S gene containing S protein among severe acute respiratory syndrome-CoV-2, severe acute respiratory syndrome-CoV, and Middle East respiratory syndrome-CoV strains, the nucleotide sequences of S gene were retrieved from GISAID [19] and Virus pathogen database and analysis resource [20]
  • The phylogenetic tree showed that the protein gene sequences were clearly clustered into three groups; severe acute respiratory syndrome-
  • Various deletions were observed in severe acute respiratory syndrome-CoV related group
  • Comparing the sequences among the three groups, various deletions were observed in the severe acute respiratory syndrome-CoV related group
  • The regions were highly conserved in severe acute respiratory syndrome-CoV-2, unlike other severe acute respiratory syndrome-CoVs
方法
  • Phylogenetic analysis of SARS-CoV-2 S protein

    To comparing of S gene containing S protein among SARS-CoV-2, SARS-CoV, and MERS-CoV strains, the nucleotide sequences of S gene were retrieved from GISAID [19] and ViPR [20].
  • Phylogenetic analysis of SARS-CoV-2 S protein.
  • To comparing of S gene containing S protein among SARS-CoV-2, SARS-CoV, and MERS-CoV strains, the nucleotide sequences of S gene were retrieved from GISAID [19] and ViPR [20].
  • After removal of identical S gene sequences, 16 genes of S protein were used in the study.
  • The sequence from SARS-CoV-2 Wuhan-Hu-1.
  • (Genbank MN908947.3 or GISAID EPI ISL 402125) was used as a representative sequence of SARS-CoV-2 strains throughout this study.
  • More detail information of the sequences used in this study were listed in Supplementary
结果
  • Results and Discussion

    Phylogenetic analysis and amino acid variation of S protein

    The phylogenetic tree showed that the protein gene sequences were clearly clustered into three groups; SARS-

    CoV-2 related, SARS-CoV related and HKU3 related groups (Fig. 1A).
  • The phylogenetic tree showed that the protein gene sequences were clearly clustered into three groups; SARS-.
  • CoV-2 related, SARS-CoV related and HKU3 related groups (Fig. 1A).
  • SARS-CoV and SARS-CoV-2 groups formed a rigid monophyletic group with their own closest bat SARS-CoV related strains, respectively.
  • The result suggested that these two human-pathogenic CoV strains were derived from common ancestral bat CoV.
  • The sequence alignments showed that there were insertions and deletion during the divergence among the strains (Fig. 1B and 1C).
  • Various deletions were observed in SARS-CoV related group.
结论
  • The fact that CoVs similar to SARS in Chinese bats is most identical to SARS-CoV-2 suggests that SARS-CoV may have been originated from a common ancestral bat CoV.
  • The regions were highly conserved in SARS-CoV-2, unlike other SARS-CoVs. Among the neutralizing antibodies for SARS-CoV and MERS-CoV, CR3022 was predicted to have better binding affinity to the S-RBD region of SARS-CoV-2 than other antibodies.
  • The comparison of antibody binding region between SARS-CoV-2 and other coronaviruses, such as SARS-CoV and MERS-CoV, was conducted to apply the suitable diagnostic or therapeutic antibodies and vaccines that are mimetics of extremely infectious SARS-CoV-2
表格
  • Table1: Neutralizing antibodies and their epitopes analyzed in the study
  • Table2: Antibody-antigen docking score and experimental affinity
  • Table3: The sequence alignments and phylogenetic analysis were done using MEGA X [21]. The nucleotide sequence were codon aligned using ClustalW with default parameters and the phylogenetic tree was inferred using neighbor-joining [22], maximum-likelihood [23], and maximum-parsimony methods [24]. The distance matrix was calculated based on the Jukes-Cantor methods [25]. The bootstrap values of the phylogenetic tree were derived from 1,000 replicates [26]. 104 Conservation score and epitope mapping of SARS-CoV-2 S protein The conservation score of amino acid positions on S protein in SARS-CoV-2 was calculated by ConSurf program [27]. The multiple sequence alignment of SARS-CoV-2 strain Wuhan-Hu-1and 12 related strains (Supplementary Table S1) was used as an input for ConSurf. In ConSurf, conservation scores and confidence intervals for the conservation scores were calculated using the empirical Bayesian method. The scores were normalized using the number of inputted sequences. Also, the highest score of ConSurf program means the most conserved position among sequences. We additionally checked the epitope positions on the SARS-CoV-2 S protein based on the known epitope information of 11 neutralizing antibodies developing for SARS-CoV and MERS-CoV. Each information of epitope positions was acquired from literatures (Table 1). 114 Structure of SARS-CoV-2 S protein S-RBD protein structure was used cryo-EM structure (Protein Data Bank ID : 6VSB) [15]. To predict the missing region of cryo-EM structure in SARS-CoV-2 S-RBD, we performed homology modeling based on known the three dimensional structure of SARS-CoV (PDB ID: 6NB7) using SWISS-MODEL (https://swissmodel.expasy.org/) [28]. Then, the best homology models were selected according to Qualitative Model Energy ANalysis (QMEAN) statistical parameter. The structures were visualized with UCSF’s Chimera (https://www.cgl.ucsf.edu/chimera/). 122 Neutralizing antibody candidates 123 As neutralizing antibody candidates of SARS-CoV-2, the five antibodies against SARS-CoV and the six
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基金
  • This work was supported by National Research Council of Science and Technology grant by the Ministry of Science and ICT (Grant No CRC‐16‐01‐KRICT)
研究对象与分析
cases: 69000
Coronavirus disease 2019 (COVID-19) is a new emerging human infectious disease caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2, also previously known as 2019-nCoV), originated in Wuhan seafood and animal market, China. Since December 2019, more than 69,000 cases of COVID-19 have been confirmed in China and quickly spreads to other counties. Currently, researchers put their best efforts to identify effective drugs for COVID-19

genomes: 62
Phylogenetic analysis of SARS-CoV-2 S protein

To comparing of S gene containing S protein among SARS-CoV-2, SARS-CoV, and MERS-CoV strains, the nucleotide sequences of S gene were retrieved from GISAID [19] and ViPR [20]. The S genes of SARS-CoV-2 were retrieved from initially sequenced 62 genomes of SARS-CoV-2 strains. After removal of identical S gene sequences, 16 genes of S protein were used in the study

genes: 16
The S genes of SARS-CoV-2 were retrieved from initially sequenced 62 genomes of SARS-CoV-2 strains. After removal of identical S gene sequences, 16 genes of S protein were used in the study. The sequence from SARS-CoV-2 Wuhan-Hu-1

(Genbank MN908947.3 or GISAID EPI ISL 402125) was used as a representative sequence of SARS-CoV-2 strains throughout this study

genomes: 62
More detail information of the sequences used in this study were listed in SupplementaryTo comparing of S gene containing S protein among SARS-CoV-2, SARS-CoV, and MERS-CoV strains, the nucleotide sequences of S gene were retrieved from GISAID [19] and ViPR [20]. The S genes of SARS-CoV-2 were retrieved from initially sequenced 62 genomes of SARS-CoV-2 strains. After removal of identical S gene sequences, 16 genes of S protein were used in the study

genes: 16
The S genes of SARS-CoV-2 were retrieved from initially sequenced 62 genomes of SARS-CoV-2 strains. After removal of identical S gene sequences, 16 genes of S protein were used in the study. The sequence from SARS-CoV-2 Wuhan-Hu-1

related strains: 12
The conservation score of amino acid positions on S protein in SARS-CoV-2 was calculated by ConSurf program [27]. The multiple sequence alignment of SARS-CoV-2 strain Wuhan-Hu-1and 12 related strains (Supplementary Table S1) was used as an input for ConSurf. In ConSurf, conservation scores and confidence intervals for the conservation scores were calculated using the empirical Bayesian method

neutralizing antibodies: 11
Also, the highest score of ConSurf program means the most conserved position among sequences. We additionally checked the epitope positions on the SARS-CoV-2 S protein based on the known epitope information of 11 neutralizing antibodies developing for SARS-CoV and. MERS-CoV

antibodies: 5
The structures were visualized with UCSF’s Chimera (https://www.cgl.ucsf.edu/chimera/). As neutralizing antibody candidates of SARS-CoV-2, the five antibodies against SARS-CoV and the six antibodies to prevent MERS-CoV were selected in the study (Table 1). The complex structure of RBD and ten neutralizing antibodies was retrieved from PDB

neutralizing antibodies: 10
As neutralizing antibody candidates of SARS-CoV-2, the five antibodies against SARS-CoV and the six antibodies to prevent MERS-CoV were selected in the study (Table 1). The complex structure of RBD and ten neutralizing antibodies was retrieved from PDB. The complex structures were superimposed to the RBD structure of SARS-CoV-2 which were built by homology modeling

antibodies: 11
To predict possible neutralizing antibody candidates of SARS-CoV-2, the docking results were compared between interface binding scores of SARS-CoV-2 S-RBD (homology modeling) and interface binding scores of. SARS-CoV or MERS-CoV S-RBD (crystal structure) with 11 antibodies for SARS-CoV and MERS-CoV have been developed. The statistical significance was tested using student’s t-test

genomes: 62
To comparing of S gene containing S protein among SARS-CoV-2, SARS-CoV, and MERS-CoV strains, the nucleotide sequences of S gene were retrieved from GISAID [19] and ViPR [20]. The S genes of SARS-CoV-2 were retrieved from initially sequenced 62 genomes of SARS-CoV-2 strains. After removal of identical S gene sequences, 16 genes of S protein were used in the study

genes: 16
The S genes of SARS-CoV-2 were retrieved from initially sequenced 62 genomes of SARS-CoV-2 strains. After removal of identical S gene sequences, 16 genes of S protein were used in the study. The sequence from SARS-CoV-2 Wuhan-Hu-1 (Genbank MN908947.3 or GISAID EPI ISL 402125) was used as a representative sequence of SARS-CoV-2 strains throughout this study

neutralizing antibodies: 11
We additionally checked the epitope positions on the SARS-CoV-2 S. protein based on the known epitope information of 11 neutralizing antibodies developing for SARS-CoV and. MERS-CoV

neutralizing antibodies: 10
Neutralizing antibody candidates antibodies to prevent MERS-CoV were selected in the study (Table 1). The complex structure of RBD and ten neutralizing antibodies was retrieved from PDB. The complex structures were superimposed to the RBD

human-pathogenic CoV strains: 2
SARS-CoV and SARS-CoV-2 groups formed a rigid monophyletic group with their own closest bat SARS-CoV related strains, respectively. The result suggested that these two human-pathogenic CoV strains were derived from common ancestral bat CoV. The sequence alignments showed that there were insertions and deletion during the divergence among the strains (Fig. 1B and 1C)

SARS-CoV neutralizing antibodies: 4
Epitope map showed that the antibody-binding residues of S protein are located within RDB region (Fig. 2). Four SARS-CoV neutralizing antibodies had the epitopes about 5 to 14 residues (total 34 residues, average 9.5 residues) of S-RBD and six MERS-CoV neutralizing antibodies bound to 22 to 33 (total 52 residues, average 25 residues) residues. Distribution of the antibody-binding residues indicates that

antibodies: 5
SARS and MERS (Fig. 3). The structures of five antibodies including m396, 80R, F26G19, S230, and CR3022 developing to prevent SARS-CoV were aligned on SARS-CoV-2 S-RBD successfully (Fig. 3A). The six MERS-

SARS-CoV antibodies: 2
Because the X-ray crystal structure of CR3022 was not revealed, the optimized structure was predicted using antibody homology modeling by 1000 structures generated using Rosetta program. As a results, two SARS-CoV antibodies including CR3022 (-13.91 dG score) and F26G19 (-15.98 dG score) and MERS-CoV. D12 (-14.01 dG score) antibody had higher binding score than other antibodies with SARS-CoV-2 S-RBD

antibodies: 11
Comparison of Antibody-RBD protein binding interaction. Based on antibody-antigen docking simulation, we calculated the binding scores between 11 antibodies and S-. RBD structures

cases: 99
9. Chen N, Zhou M, Dong X, Qu J, Gong F, Han Y, Qiu Y, Wang J, Liu Y, Wei Y et al: Epidemiological and clinical characteristics of 99 cases of 2019 novel coronavirus pneumonia in Wuhan, China: a descriptive study. Lancet 2020

SARS-CoV species: 27
SARS-CoV but also SARS-CoV-2_bat-RaTG13 showed low sequence similarity. The lower panel indicates the conservation score of the protein sequences of 27 SARS-CoV species. (C) Intersertional regions in SARS-CoV-2. S protein

SARS-CoV antibodies: 5
The predicted RBD. structure of SARS-CoV-2 S protein in complex with five SARS-CoV antibodies (B) and six MERS-CoV. antibodies (C)

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Tamina Park
Tamina Park
Sang-Yeop Lee
Sang-Yeop Lee
Seil Kim
Seil Kim
Mi Jeong Kim
Mi Jeong Kim
Hong Gi Kim
Hong Gi Kim
Sangmi Jun
Sangmi Jun
Bum Tae Kim
Bum Tae Kim
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