scholarly journals Spike protein modeling and single amino acid variant analysis might suggest reduced transmitability of SARS-CoV-2 in Jordan, Middle East

2020 ◽  
Author(s):  
Walid Al-Zyoud ◽  
Hazem Haddad
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Middle East ◽  
Active Sites ◽  
Trna Synthetase ◽  
Surface Glycoprotein ◽  
Spike Protein ◽  
Single Amino Acid ◽  
Spike Glycoprotein ◽  
Amino Acid Variants

Abstract Spike protein (approx. 180 kDa) is the surface glycoprotein of the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) necessary for the interaction of the virus with human endothelial cell receptors on the cell membrane to be engulfed causing COVID-19 disease after binding with the angiotensin-converting enzyme 2 (ACE2) with an evident activation by type II transmembrane protease TMPRSS2 . Therefore, mutations and amino acid variants analysis are essential in characterizing the mechanism of binding of spike protein with its receptor, which totally gives insights on possibilities to design a peptide or nucleotide-based vaccine for COVID-19. Here, we employed Iterative Threading Assembly Refinement (I-TASSER) and Multiple Alignment using Fast Fourier Transform (MAFFT) to predict the three-dimensional structure and to analyze the amino acid variants for spike protein sequences of SARS-CoV-2 from GISAID database of samples collected from Jordan to try to find a justification for low number of confirmed COVID-19 in Jordan, Middle East. Our findings showed the molecules structurally close to the spike glycoprotein from the Enzyme Commission (EC) numbers and active sites included Isoleucyl-tRNA synthetase, Crystal structure of the tricorn protease (hydrolase); Crystal structure of the T. Thermophilus RNA polymerase holoenzyme (transferase); Crystal structure of the complex between pyruvate-ferredoxin oxidoreductase from Desulfovibrio africanus and pyruvate (oxidoreductase); and Reovirus core (virus). Our MAFFT findings showed that Four Amino Acid Variants (SAV) founded in 20 samples of SARS-CoV-2 were not conserved residues in spike glycoprotein. What is equal to 5% of samples showed tyrosine (polar) deletion at Y144 , 62% of samples showed aspartate (polar, acidic) substitution to glycine (nonpolar) at D614G, 5% of samples showed aspartate (polar, acidic) substitution to tyrosine (polar) at D1139Y and 5% of samples showed glycine (nonpolar) substitution to serine (polar) at G1167S respectively. By using Phyre2, our findings have shown lower sensitive mutational that cannot affect the pocket region or alpha and beta-sheet in all mutations except for D614G, which has the highest mutational sensitivity score (5 out of 9) indicating a bigger effect on the function of spike protein. This might suggest, in general, a reduced transmitability of SARS-CoV-2 in Jordan, Middle East. As the crystal structure of spike protein is not revealed yet, it was not possible to compare the predicted modes versus each other.

scholarly journals Single Amino Acid Variant (SAV) Percentage and Monomer Modeling of Spike Protein of SARS-CoV-2 in Jordan

2020 ◽  
Author(s):  
Walid Al-Zyoud ◽  
Hazem Haddad ◽  
Ramzi Foudeh
Keyword(s):  
Amino Acid ◽  
Three Dimensional ◽  
Surface Glycoprotein ◽  
Spike Protein ◽  
Single Amino Acid ◽  
Angiotensin Converting Enzyme 2 ◽  
Sensitivity Score ◽  
Monomer Structure ◽  
Amino Acid Variant ◽  
Amino Acid Variants

Spike protein is the surface glycoprotein of the severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) necessary for the entry of the virus via the transmembrane receptors of the human endothelial cells of the respiratoty system for the virus to be engulfed causing COVID-19 disease after priming by type II transmembrane protease TMPRSS2 and then binding with the angiotensin-converting enzyme 2 (ACE2). Therefore, mutations and amino acid variants analysis are essential in understanding the mechanism of binding of spike protein with its receptor to have an insights on possibilities to design a peptide or nucleotide-based vaccine for COVID-19. Here, we employed Iterative Threading Assembly Refinement (I-TASSER) and Multiple Alignment using Fast Fourier Transform (MAFFT) to predict the three-dimensional monomer structure of spike protein of SARS-CoV-2 and to analyze the amino acid variants for protein sequences from GISAID database for samples collected from Jordan in a try to find an explanation for the low confirmed number of COVID-19 in Jordan. Our Protein Homology/analogY Recognition Engine V 2.0 (Phyre2) findings showed four single amino acid variants (SAV) found in 20 samples of SARS-CoV-2. What is equal to 5% of samples showed tyrosine deletion at Y144 located in the SARS-CoV-like_Spike_S1_NTD (N terminal domain), 62% showed aspartate substitution to glycine at D614G located in the SARS-CoV-2_Spike_S1_RBD (spike recognition binding site), 5% showed aspartate substitution to tyrosine at D1139Y and 5% showed glycine substitution to serine at G1167S both located in the Corona_S2 domain. The findings have shown lower mutational sensitivity in all variants that might not affect the function of spike glycoprotein except for D614G, which has the highest mutational sensitivity score (5 out of 9) indicating a higher likelihood to affect the function of the spike protein. This might suggest, in general, a reduced transmitability of SARS-CoV-2 in Jordan.

scholarly journals The Functional Consequences of the Novel Ribosomal Pausing Site in SARS-CoV-2 Spike Glycoprotein RNA

2021 ◽  
Vol 22 (12) ◽  
pp. 6490
Author(s):  
Olga A. Postnikova ◽  
Sheetal Uppal ◽  
Weiliang Huang ◽  
Maureen A. Kane ◽  
Rafael Villasmil ◽  
...  
Keyword(s):  
Amino Acid ◽  
Insertion Sequence ◽  
Experimental Studies ◽  
Spike Protein ◽  
Spike Glycoprotein ◽  
Furin Cleavage ◽  
New Host ◽  
S Protein ◽  
Furin Cleavage Site ◽  
Ribosome Pausing

The SARS-CoV-2 Spike glycoprotein (S protein) acquired a unique new 4 amino acid -PRRA- insertion sequence at amino acid residues (aa) 681–684 that forms a new furin cleavage site in S protein as well as several new glycosylation sites. We studied various statistical properties of the -PRRA- insertion at the RNA level (CCUCGGCGGGCA). The nucleotide composition and codon usage of this sequence are different from the rest of the SARS-CoV-2 genome. One of such features is two tandem CGG codons, although the CGG codon is the rarest codon in the SARS-CoV-2 genome. This suggests that the insertion sequence could cause ribosome pausing as the result of these rare codons. Due to population variants, the Nextstrain divergence measure of the CCU codon is extremely large. We cannot exclude that this divergence might affect host immune responses/effectiveness of SARS-CoV-2 vaccines, possibilities awaiting further investigation. Our experimental studies show that the expression level of original RNA sequence “wildtype” spike protein is much lower than for codon-optimized spike protein in all studied cell lines. Interestingly, the original spike sequence produces a higher titer of pseudoviral particles and a higher level of infection. Further mutagenesis experiments suggest that this dual-effect insert, comprised of a combination of overlapping translation pausing and furin sites, has allowed SARS-CoV-2 to infect its new host (human) more readily. This underlines the importance of ribosome pausing to allow efficient regulation of protein expression and also of cotranslational subdomain folding.

Covid-19 Mutations and How the Vaccine Enhances Immune Intelligence

2021 ◽  
Author(s):  
Xanya Sofra
Keyword(s):  
Amino Acid ◽  
Viral Entry ◽  
Open Reading Frames ◽  
Spike Protein ◽  
Single Amino Acid ◽  
Immune Memory ◽  
Type I ◽  
Specific Amino Acid ◽  
Adaptive Efficiency ◽  
Acid Exchange

We traced the coronavirus classification and evolution, analyzed the Covid-19 composition and its distinguishing characteristics when compared to SARS-CoV and MERS-CoV. Despite their close kinship, SARS-CoV and Covid-19 display significant structural differences, including 380 amino acid substitutions, and variable homology between certain open reading frames that are bound to diversify the pathogenesis and virulence of the two viral compounds. A single amino acid substitution such as replacing Aspartate (D) with Glycine (G) composes the D614G mutation that is around 20% more infectious than its predecessor 614D. The B117 variant, that exhibits a 70% transmissibility rate, harbours 23 mutants, each reflecting one amino acid exchange. We examined several globally spreading mutations, 501.V2, B1351, P1, and others, with respect to the specific amino acid conversions involved. Unlike previous versions of coronavirus, where random mutations eventually precipitate extinction, the multiplicity of over 300,000 mutations appears to have rendered Covid-19 more contagious, facilitating its ability to evade detection, thus challenging the effectiveness of a large variety of emerging vaccines. Vaccination enhances immune memory and intelligence to combat or obstruct viral entry by generating antibodies that will prohibit the cellular binding and fusion with the Spike protein, ultimately debilitating the virus from releasing its contents into the cell. Developing antibodies during the innate response, appears to be the most compelling solution in light of the hypothesis that Covid-19 inhibits the production of Interferon type I, compromising adaptive efficiency to recognize the virus, possibly provoking a cytokine storm that injures vital organs. With respect to that perspective, the safety and effectiveness of different vaccines is evaluated and compared, including the Spike protein mRNA version, the Adenovirus DNA, Spike protein subunits, the deactivated virus genres, or, finally, the live attenuated coronavirus that appears to demonstrate the greatest effectiveness, yet, encompass a relatively higher risk.

scholarly journals Optimized Pseudotyping Conditions for the SARS-COV-2 Spike Glycoprotein

2020 ◽  
Vol 94 (21) ◽  
Author(s):  
Marc C. Johnson ◽  
Terri D. Lyddon ◽  
Reinier Suarez ◽  
Braxton Salcedo ◽  
Mary LePique ◽  
...  
Keyword(s):  
Signal Peptide ◽  
Neutralizing Antibodies ◽  
Particle Production ◽  
Surface Glycoprotein ◽  
Spike Protein ◽  
Target Cells ◽  
Spike Glycoprotein ◽  
Infectious Particle ◽  
Pathogenic Virus ◽  
Hiv 1

ABSTRACT The severe acute respiratory syndrome coronavirus 2 (SARS-COV-2) Spike glycoprotein is solely responsible for binding to the host cell receptor and facilitating fusion between the viral and host membranes. The ability to generate viral particles pseudotyped with SARS-COV-2 Spike is useful for many types of studies, such as characterization of neutralizing antibodies or development of fusion-inhibiting small molecules. Here, we characterized the use of a codon-optimized SARS-COV-2 Spike glycoprotein for the generation of pseudotyped HIV-1, murine leukemia virus (MLV), and vesicular stomatitis virus (VSV) particles. The full-length Spike protein functioned inefficiently with all three systems but was enhanced over 10-fold by deleting the last 19 amino acids of the cytoplasmic tail. Infection of 293FT target cells was possible only if the cells were engineered to stably express the human angiotensin-converting enzyme 2 (ACE2) receptor, but stably introducing an additional copy of this receptor did not further enhance susceptibility. Stable introduction of the Spike-activating protease TMPRSS2 further enhanced susceptibility to infection by 5- to 10-fold. Replacement of the signal peptide of the Spike protein with an optimal signal peptide did not enhance or reduce infectious particle production. However, modifications D614G and R682Q further enhanced infectious particle production. With all enhancing elements combined, the titer of pseudotyped HIV-1 particles reached almost 106 infectious particles/ml. Finally, HIV-1 particles pseudotyped with SARS-COV-2 Spike were successfully used to detect neutralizing antibodies in plasma from coronavirus disease 2019 (COVID-19) patients, but not in plasma from uninfected individuals. IMPORTANCE In work with pathogenic viruses, it is useful to have rapid quantitative tests for viral infectivity that can be performed without strict biocontainment restrictions. A common way of accomplishing this is to generate viral pseudoparticles that contain the surface glycoprotein from the pathogenic virus incorporated into a replication-defective viral particle that contains a sensitive reporter system. These pseudoparticles enter cells using the glycoprotein from the pathogenic virus, leading to a readout for infection. Conditions that block entry of the pathogenic virus, such as neutralizing antibodies, will also block entry of the viral pseudoparticles. However, viral glycoproteins often are not readily suited for generating pseudoparticles. Here, we describe a series of modifications that result in the production of relatively high-titer SARS-COV-2 pseudoparticles that are suitable for the detection of neutralizing antibodies from COVID-19 patients.

SAAVpedia: Identification, Functional Annotation, and Retrieval of Single Amino Acid Variants for Proteogenomic Interpretation

2019 ◽  
Vol 18 (12) ◽  
pp. 4133-4142
Author(s):  
Soo Youn Lee ◽  
Heeyoun Hwang ◽  
Young-Mook Kang ◽  
Hyejin Kim ◽  
Dong Geun Kim ◽  
...  
Keyword(s):  
Amino Acid ◽  
Functional Annotation ◽  
Single Amino Acid ◽  
Amino Acid Variants

Predicting Structural and Serologic Modifications in Blood Group Antigens by Homology Modeling.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 451-451
Author(s):  
Connie M. Westhoff ◽  
Dwane E. Wylie
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Homology Modeling ◽  
Blood Group ◽  
Amino Acid Change ◽  
Single Amino Acid ◽  
Side Chain ◽  
Blood Group Antigens ◽  
Aromatic Side Chain ◽  
Transport Channel

Abstract Homology modeling of blood group proteins offers the possibility of predicting the effect of amino acid changes on serologic phenotype and immunogenicity. The location of an amino acid change within known structural motifs, its phylogenetic conservation, and its proximity to known epitopes give insight into its potential effect on protein structure and, consequently, its clinical significance. We applied this approach to investigate the loss of membrane expression of the Dombrock blood group antigens in a patient with a single amino acid change and to investigate RhD alterations in weak D phenotypes. The Dombrock homology model was derived with rat ART2.2 crystal structure as template. For the RhD model, the crystal structure of the Rh-like-ammonia transporter from Nitrosomonas europaea was used. Protein alignment was derived with Clustal X, adjusted visually, and submitted to the Swiss Modeling server. Models were viewed with Deep View Swiss Pdb Viewer. The Dombrock null containes a Phe62Ser substitution. This Phe (F) residue is located in an FDDQY motif near the COOH terminus. This region of the protein also contains a HYYLT motif. These two motifs are highly conserved in the ART protein family and contribute several aromatic amino acids to this region of the molecule. Aromatic side chain interactions between these residues could contribute to the stability of the Do protein. In support, the distance in the ART2.2 crystal structure between Phe in FDDQY and His in HYYLT is 3.7 Å, which is the appropriate distance for aromatic side chain interactions. This is also the measured distance between these two residues in the Do model. Thus, protein modeling indicates that the Phe62Ser mutation disrupts important stacking interactions between Phe62 and His160. When amino acid changes causing weak D phenotypes were examined, some of those affecting expression of RhD were located near the vestigial transport channel. These include the Trp220Arg mutation (weak D Type 16). This Trp residue is part of the transport channel in Nitrosomonas and is conserved in Rh proteins of almost all species. Its role in maintaining Rh structure is indicated by the dramatic effect its modification has on protein and epitope expression. Additionally, Arg114Trp change (weak D Type 17), which is also near the channel, reduces D expression to only 66 antigen sites/cell. GlyXXXGly motifs stabilize interactions of adjacent alpha helices in membrane proteins. Evidence for a role in stabilization of RhD is revealed by the Gly282Asp mutation (weak D Type 15) which is part of such a motif. In addition, a D-epitope in loop 3 is near the 282Asp residue. Alteration of helical packing accompanied by epitope conformation could explain production of anti-D in patients with weak D Type 15. Homology modeling is an important tool for understanding the structure and serologic bases of blood group proteins and will continue to give important insight as more protein crystal structures become available.

scholarly journals Quantitative characterization of all single amino acid variants of a viral capsid-based drug delivery vehicle

2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Emily C. Hartman ◽  
Christopher M. Jakobson ◽  
Andrew H. Favor ◽  
Marco J. Lobba ◽  
Ester Álvarez-Benedicto ◽  
...  
Keyword(s):  
Drug Delivery ◽  
Amino Acid ◽  
Single Amino Acid ◽  
Viral Capsid ◽  
Delivery Vehicle ◽  
Quantitative Characterization ◽  
Drug Delivery Vehicle ◽  
Amino Acid Variants

Structure of nondiscriminating glutamyl-tRNA synthetase fromThermotoga maritima

2010 ◽  
Vol 66 (7) ◽  
pp. 813-820 ◽  
Author(s):  
Takuhiro Ito ◽  
Noriko Kiyasu ◽  
Risa Matsunaga ◽  
Seizo Takahashi ◽  
Shigeyuki Yokoyama
Keyword(s):  
Crystal Structure ◽  
Amino Acid ◽  
Thermotoga Maritima ◽  
Trna Synthetase ◽  
Characteristic Structure ◽  
Trna Synthetases ◽  
Aminoacyl Trna Synthetases ◽  
Rossmann Fold

Aminoacyl-tRNA synthetases produce aminoacyl-tRNAs from the substrate tRNA and its cognate amino acid with the aid of ATP. Two types of glutamyl-tRNA synthetase (GluRS) have been discovered: discriminating GluRS (D-GluRS) and nondiscriminating GluRS (ND-GluRS). D-GluRS glutamylates tRNAGluonly, while ND-GluRS glutamylates both tRNAGluand tRNAGln. ND-GluRS produces the intermediate Glu-tRNAGln, which is converted to Gln-tRNAGlnby Glu-tRNAGlnamidotransferase. Two GluRS homologues fromThermotoga maritima, TM1875 and TM1351, have been biochemically characterized and it has been clarified that only TM1875 functions as an ND-GluRS. Furthermore, the crystal structure of theT. maritimaND-GluRS, TM1875, was determined in complex with a Glu-AMP analogue at 2.0 Å resolution. TheT. maritimaND-GluRS contains a characteristic structure in the connective-peptide domain, which is inserted into the catalytic Rossmann-fold domain. The glutamylation ability of tRNAGlnby ND-GluRS was measured in the presence of the bacterial Glu-tRNAGlnamidotransferase GatCAB. Interestingly, the glutamylation efficiency was not affected even in the presence of excess GatCAB. Therefore, GluRS avoids competition with GatCAB and glutamylates tRNAGln.

scholarly journals Multiplex Assessment of Protein Variant Abundance by Massively Parallel Sequencing

2018 ◽  
Author(s):  
Kenneth A. Matreyek ◽  
Lea M. Starita ◽  
Jason J. Stephany ◽  
Beth Martin ◽  
Melissa A. Chiasson ◽  
...  
Keyword(s):  
Amino Acid ◽  
Massively Parallel Sequencing ◽  
Dominant Negative ◽  
Massively Parallel ◽  
Single Amino Acid ◽  
Protein Variant ◽  
Parallel Sequencing ◽  
Missense Variants ◽  
Functional Variants ◽  
Amino Acid Variants

ABSTRACTDetermining the pathogenicity of human genetic variants is a critical challenge, and functional assessment is often the only option. Experimentally characterizing millions of possible missense variants in thousands of clinically important genes will likely require generalizable, scalable assays. Here we describe Variant Abundance by Massively Parallel Sequencing (VAMP-seq), which measures the effects of thousands of missense variants of a protein on intracellular abundance in a single experiment. We apply VAMP-seq to quantify the abundance of 7,595 single amino acid variants of two proteins, PTEN and TPMT, in which functional variants are clinically actionable. We identify 1,079 PTEN and 805 TPMT single amino acid variants that result in low protein abundance, and may be pathogenic or alter drug metabolism, respectively. We observe selection for low-abundance PTEN variants in cancer, and our abundance data suggest that a PTEN variant accounting for ~10% of PTEN missense variants in melanomas functions via a dominant negative mechanism. Finally, we demonstrate that VAMP-seq can be applied to other genes, highlighting its potential as a generalizable assay for characterizing missense variants.

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