In Silico Design of Chimeric and Immunogenic Protein-Containing IpaB and IpaD as a Vaccine Candidate against Shigella dysenteriae

2020 ◽  
Vol 17 (4) ◽  
pp. 333-341
Author(s):  
Seyed Akbar Arianzad ◽  
Mehdi Zeinoddini ◽  
Azam Haddadi ◽  
Shahram Nazarian ◽  
Reza Hasan Sajedi
Keyword(s):  
Tertiary Structure ◽  
Vaccine Candidate ◽  
Minimum Energy ◽  
Chimeric Protein ◽  
Severe Form ◽  
Amino Acid Sequences ◽  
Shigella Dysenteriae ◽  
Intestinal Epithelial ◽  
Suitable Target ◽  
Immunogenic Properties

Background: S. dysenteriaeis the causative agent of shigellosis, a severe form of bacillary dysentery and this infectious disease is still a health problem worldwide, especially in children. The most important proteins of the Shigellatype III secretion system are IpaB and IpaD, which attach to the intestinal epithelial cells and provide the possibility of invasion and disease. These two proteins with immunogenic properties can be a suitable target to design and manufacture subunit recombinant vaccines. Objective: The aim of this study is to design an immunogenic chimeric protein against IpaB and IpaD as a subunit vaccine candidate through an in silicostudy. Methods: Firstly, the immunogenic epitopes of amino acid sequences, physico-chemical parameters, and the allergenicity of the chimeric protein were determined. Then the tertiary structure and the potential ability of the chimeric protein were predicted and evaluated in terms of inducing B cells’ immune responses with effective epitopes. Finally, the optimization of the chimeric protein was examined as the index affecting the protein expression. Results: Data showed an instability index of 37.18 and a well-established predicted third structure for the chimeric protein, with a z-score of -6.11. Also, more than 99% of its amino acids were in the optimal range. Minimum energy for mRNA structure increased to -317.9 and the Codon Adaptive Index (CAI) rose to 88%. The designed protein had no IgE specific B cell epitopes. Conclusion: Overall, the results of this study show that the designed protein can be considered as an immunogen vaccine candidate against S. dysenteriae.

Immunogenicity Evaluation of Chimeric Subunit Vaccine Comprising Adhesion Coli Surface Antigens from Enterotoxigenic Escherichia coli

2019 ◽  
Vol 29 (1-6) ◽  
pp. 91-100
Author(s):  
Dorna Khoobbakht ◽  
Shohreh Zare Karizi ◽  
Mohammad Javad  Motamedi ◽  
Rouhollah Kazemi ◽  
Pooneh Roghanian ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Epithelial Cells ◽  
Subunit Vaccine ◽  
Fusion Gene ◽  
Intestinal Epithelial Cells ◽  
High Titer ◽  
Chimeric Protein ◽  
Amino Acid Sequences ◽  
Intestinal Epithelial ◽  
E Coli

Enterotoxigenic <i>Escherichia coli</i> (ETEC) is the most common agent of diarrhea morbidity in developing countries. ETEC adheres to host intestinal epithelial cells via various colonization factors. The CooD and CotD proteins play a significant role in bacteria binding to the intestinal epithelial cells as adhesin tip subunits of CS1 and CS2 pili. The purpose here was to design a new construction containing <i>cooD</i> and <i>cotD</i> genes and use several types of bioinformatics software to predict the structural and immunological properties of the designed antigen. The fusion gene was synthesized with codon bias of <i>E. coli</i> in order to increase the expression level of the protein. The amino acid sequences, protein structure, and immunogenicity properties of potential antigens were analyzed in silico. The chimeric protein was expressed in <i>E. coli</i>BL21 (DE3). The antigenicity of the recombinant proteins was verified by Western blotting and ELISA. In order to assess the induced immunity, the immunized mice were challenged with wild-type ETEC by an intraperitoneal route. Immunological analyses showed the production of a high titer of IgG serum with no sign of serum-mucosal IgA antibody response. The result of the challenge assay showed that 30% of immunized mice survived. The results of this study showed that CooD-CotD recombinant protein can stimulate immunity against ETEC. The designed chimera could be a prototype for the subunit vaccine, which is worthy of further consideration.

scholarly journals A Chimeric Protein That Functions as both an Anthrax Dual-Target Antitoxin and a Trivalent Vaccine

2010 ◽  
Vol 54 (11) ◽  
pp. 4750-4757 ◽  
Author(s):  
Gaobing Wu ◽  
Yuzhi Hong ◽  
Aizhen Guo ◽  
Chunfang Feng ◽  
Sha Cao ◽  
...  
Keyword(s):  
Protective Antigen ◽  
Vaccine Candidate ◽  
Lethal Factor ◽  
Lethal Dose ◽  
Chimeric Protein ◽  
Dominant Negative ◽  
Lethal Challenge ◽  
Edema Factor ◽  
Trivalent Vaccine

ABSTRACT Effective measures for the prophylaxis and treatment of anthrax are still required for counteracting the threat posed by inhalation anthrax. In this study, we first demonstrated that the chimeric protein LFn-PA, created by fusing the protective antigen (PA)-binding domain of lethal factor (LFn) to PA, retained the functions of the respective molecules. On the basis of this observation, we attempted to develop an antitoxin that targets the binding of lethal factor (LF) and/or edema factor (EF) to PA and the transportation of LF/EF. Therefore, we replaced PA in LFn-PA with a dominant-negative inhibitory PA (DPA), i.e., PAF427D. In in vitro models of anthrax intoxication, the LFn-DPA chimera showed 3-fold and 2-fold higher potencies than DPA in protecting sensitive cells against anthrax lethal toxin (LeTx) and edema toxin (EdTx), respectively. In animal models, LFn-DPA exhibited strong potency in rescuing mice from lethal challenge with LeTx. We also evaluated the immunogenicity and immunoprotective efficacy of LFn-DPA as an anthrax vaccine candidate. In comparison with recombinant PA, LFn-DPA induced significantly higher levels of the anti-PA immune response. Moreover, LFn-DPA elicited an anti-LF antibody response that could cross-react with EF. Mice immunized with LFn-DPA tolerated a LeTx challenge that was 5 times its 50% lethal dose. Thus, LFn-DPA represents a highly effective trivalent vaccine candidate for both preexposure and postexposure vaccination. Overall, we have developed a novel and dually functional reagent for the prophylaxis and treatment of anthrax.

scholarly journals In-silicoprediction and modeling of theEntamoeba histolyticaproteins: Serine-richEntamoeba histolyticaprotein and 29 kDa Cysteine-rich protease

PeerJ ◽  
2017 ◽  
Vol 5 ◽  
pp. e3160 ◽  
Author(s):  
Kumar Manochitra ◽  
Subhash Chandra Parija
Keyword(s):  
Amino Acid ◽  
Structure Prediction ◽  
Tertiary Structure ◽  
Protein Structures ◽  
Amino Acid Sequences ◽  
Treatment Modalities ◽  
Bioinformatic Tools ◽  
Complex Protein ◽  
A Cell ◽  
Quaternary Structures

BackgroundAmoebiasis is the third most common parasitic cause of morbidity and mortality, particularly in countries with poor hygienic settings. There exists an ambiguity in the diagnosis of amoebiasis, and hence there arises a necessity for a better diagnostic approach. Serine-richEntamoeba histolyticaprotein (SREHP), peroxiredoxin and Gal/GalNAc lectin are pivotal inE. histolyticavirulence and are extensively studied as diagnostic and vaccine targets. For elucidating the cellular function of these proteins, details regarding their respective quaternary structures are essential. However, studies in this aspect are scant. Hence, this study was carried out to predict the structure of these target proteins and characterize them structurally as well as functionally using appropriatein-silicomethods.MethodsThe amino acid sequences of the proteins were retrieved from National Centre for Biotechnology Information database and aligned using ClustalW. Bioinformatic tools were employed in the secondary structure and tertiary structure prediction. The predicted structure was validated, and final refinement was carried out.ResultsThe protein structures predicted by i-TASSER were found to be more accurate than Phyre2 based on the validation using SAVES server. The prediction suggests SREHP to be an extracellular protein, peroxiredoxin a peripheral membrane protein while Gal/GalNAc lectin was found to be a cell-wall protein. Signal peptides were found in the amino-acid sequences of SREHP and Gal/GalNAc lectin, whereas they were not present in the peroxiredoxin sequence. Gal/GalNAc lectin showed better antigenicity than the other two proteins studied. All the three proteins exhibited similarity in their structures and were mostly composed of loops.DiscussionThe structures of SREHP and peroxiredoxin were predicted successfully, while the structure of Gal/GalNAc lectin could not be predicted as it was a complex protein composed of sub-units. Also, this protein showed less similarity with the available structural homologs. The quaternary structures of SREHP and peroxiredoxin predicted from this study would provide better structural and functional insights into these proteins and may aid in development of newer diagnostic assays or enhancement of the available treatment modalities.

Chimeric protein consisting of 3M2e and HSP as a universal influenza vaccine candidate: from in silico analysis to preliminary evaluation

Virus Genes ◽  
2018 ◽  
Vol 55 (1) ◽  
pp. 22-32 ◽  
Author(s):  
Behrokh Farahmand ◽  
Najmeh Taheri ◽  
Hadiseh Shokouhi ◽  
Hoorieh Soleimanjahi ◽  
Fatemeh Fotouhi
Keyword(s):  
Influenza Vaccine ◽  
In Silico ◽  
Vaccine Candidate ◽  
Chimeric Protein ◽  
In Silico Analysis ◽  
Preliminary Evaluation ◽  
Universal Influenza Vaccine ◽  
Silico Analysis

scholarly journals A chimeric protein-based malaria vaccine candidate induces robust T cell responses against Plasmodium vivax MSP119

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Jairo Andres Fonseca ◽  
Monica Cabrera-Mora ◽  
Balwan Singh ◽  
Joseli Oliveira-Ferreira ◽  
Josué da Costa Lima-Junior ◽  
...  
Keyword(s):  
T Cell ◽  
Plasmodium Vivax ◽  
Malaria Vaccine ◽  
Vaccine Candidate ◽  
Chimeric Protein ◽  
T Cell Responses ◽  
Cell Responses ◽  
Malaria Vaccine Candidate

scholarly journals Endocytosis of chimeric influenza virus hemagglutinin proteins that lack a cytoplasmic recognition feature for coated pits.

1996 ◽  
Vol 134 (2) ◽  
pp. 339-348 ◽  
Author(s):  
J Lazarovits ◽  
H Y Naim ◽  
A C Rodriguez ◽  
R H Wang ◽  
E Fire ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Cell Surface ◽  
Dna Sequences ◽  
Chimeric Protein ◽  
Amino Acid Sequences ◽  
Surface Proteins ◽  
Wild Type ◽  
Coated Pits ◽  
External Domain ◽  
Simplex Virus

The influenza virus A/Japan/305/57 hemagglutinin (HA) can be converted from a protein that is essentially excluded from coated pits into one that is internalized at approximately the rate of uptake of bulk membrane by replacing the HA transmembrane and cytoplasmic sequences with those of either of two other glycoproteins (Roth et al., 1986. J. Cell Biol. 102:1271-1283). To identify more precisely the foreign amino acid sequences responsible for this change in HA traffic, DNA sequences encoding the transmembrane (TM) or cytoplasmic (CD) domains of either the G glycoprotein of vesicular stomatitis virus (VSV) or the gC glycoprotein of herpes simplex virus were exchanged for those encoding the analogous regions of wild type HA (HA wt). HA-HA-G and HA-HA-gC, chimeras that contain only a foreign CD, resembled HA wt in having a long residence on the cell surface and were internalized very slowly. HA-HA-gC was indistinguishable from HA in our assays, whereas twice as much HA-HA-G was internalized as was HA wt. However, HA-G-HA, containing only a foreign TM, was internalized as efficiently as was HA-G-G, a chimeric protein with transmembrane and cytoplasmic sequences of VSV G protein. Conditions that blocked internalization through coated pits also inhibited endocytosis of the chimeric proteins. Although the external domains of the chimeras were less well folded than that of the wild type HA, denaturation of the wild type HA external domain by treatment with low pH did not increase the interaction of HA with coated pits. However, mutation of four amino acids in the TM of HA allowed the protein to be internalized, indicating that the property that allows HA to escape endocytosis resides in its TM. These results indicate that possession of a cytoplasmic recognition feature is not required for the internalization of all cell surface proteins and suggest that multiple mechanisms for internalization exist that operate at distinctly different rates.

scholarly journals Analysis of a nucleotide-binding site of 5-lipoxygenase by affinity labelling: binding characteristics and amino acid sequences

2000 ◽  
Vol 351 (3) ◽  
pp. 697-707 ◽  
Author(s):  
Ying-Yi ZHANG ◽  
Tove HAMMARBERG ◽  
Olof RADMARK ◽  
Bengt SAMUELSSON ◽  
Carol F. NG ◽  
...  
Keyword(s):  
Amino Acid ◽  
Binding Site ◽  
Tertiary Structure ◽  
Nucleotide Binding ◽  
Peptide Sequencing ◽  
Amino Acid Sequences ◽  
Nucleotide Binding Site ◽  
Atp Binding ◽  
Binding Characteristics ◽  
Atp Binding Site

5-Lipoxygenase (5LO) catalyses the first two steps in the biosynthesis of leukotrienes, which are inflammatory mediators derived from arachidonic acid. 5LO activity is stimulated by ATP; however, a consensus ATP-binding site or nucleotide-binding site has not been found in its protein sequence. In the present study, affinity and photoaffinity labelling of 5LO with 5′-p-fluorosulphonylbenzoyladenosine (FSBA) and 2-azido-ATP showed that 5LO bound to the ATP analogues quantitatively and specifically and that the incorporation of either analogue inhibited ATP stimulation of 5LO activity. The stoichiometry of the labelling was 1.4mol of FSBA/mol of 5LO (of which ATP competed with 1mol/mol) or 0.94mol of 2-azido-ATP/mol of 5LO (of which ATP competed with 0.77mol/mol). Labelling with FSBA prevented further labelling with 2-azido-ATP, indicating that the same binding site was occupied by both analogues. Other nucleotides (ADP, AMP, GTP, CTP and UTP) also competed with 2-azido-ATP labelling, suggesting that the site was a general nucleotide-binding site rather than a strict ATP-binding site. Ca2+, which also stimulates 5LO activity, had no effect on the labelling of the nucleotide-binding site. Digestion with trypsin and peptide sequencing showed that two fragments of 5LO were labelled by 2-azido-ATP. These fragments correspond to residues 73–83 (KYWLNDDWYLK, in single-letter amino acid code) and 193–209 (FMHMFQSSWNDFADFEK) in the 5LO sequence. Trp-75 and Trp-201 in these peptides were modified by the labelling, suggesting that they were immediately adjacent to the C-2 position of the adenine ring of ATP. Given the stoichiometry of the labelling, the two peptide sequences of 5LO were probably near each other in the enzyme's tertiary structure, composing or surrounding the ATP-binding site of 5LO.

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