Immunoinformatics Approach to Design a Multi-Epitope Nanovaccine against Leishmania Parasite: Elicitation of Cellular Immune Responses
Abstract
:1. Introduction
2. Materials and Methods
2.1. Data Retrieval and Proteome Analysis
2.2. Antigenicity and Allergenicity Evaluation
2.3. T Cell Epitope Prediction, Epitope Selection and Vaccine Construction
2.4. Immunoinformatics Analysis of Chimeric Protein
2.4.1. Physicochemical Properties
2.4.2. Secondary and Tertiary Structure
2.4.3. Refinement and Validation of Tertiary Structure Model
2.4.4. In Silico Prediction of LeishChim’s Docking to MHC Class I and II Molecules
2.4.5. Molecular Dynamics (MD) Simulation of LeishChim-MHCI/MHCII Complexes
2.4.6. CTL, HTL, B Cell Linear and Conformational Epitope Prediction
2.4.7. Immune Simulations
2.5. Expression of Chimeric Protein
2.6. PLGA Vaccine Preparation and Characterization
2.6.1. Materials
2.6.2. Preparation and Characterization of PLGA Nanoparticles
2.6.3. Quantification of Antigen Loading
2.6.4. Quantification of MPLA Loading
2.6.5. In Vitro Release Studies
2.6.6. In Vitro Stability
2.7. Experimental Animals
2.8. Vaccination Schedule
2.9. LeishChim-Specific Spleen Cell Proliferation Assay
2.10. Flow Cytometry Analysis for Determination of LeishChim-Specific Memory and Cytokine-Producing T Cells
2.11. Statistical Analysis
3. Results
3.1. Conserved Leishmania Protein Retrieval and Identification of Cellular Localization
3.2. Antigenicity and Allergenicity Evaluation
3.3. MHC I- and II-Binding Epitope Identification, Selection and Vaccine Construction
3.4. Immunoinformatics Evaluation of Chimeric Protein
3.4.1. Physicochemical Properties, Antigenicity and Allergenicity Evaluation
3.4.2. Secondary and Tertiary Structure
3.4.3. Refinement and Validation of the Tertiary Structure
3.4.4. In Silico Prediction of LeishChim’s Docking onto MHCI and MHCII Molecules
3.4.5. Stability Prediction of the LeishChim and MHCI/MHCII Complexes
3.4.6. B Cell Linear and Conformational Epitope Prediction
3.4.7. CTL and HTL Epitope Prediction
3.4.8. In Silico Immune Simulation Profile of LeishChim
3.5. Immunization with Multi-Epitope Chimeric Protein Encapsulated in PLGA NPs Elicited Antigen-Specific Cellular Immune Responses
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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No | Sequence ID | Function | Cellular Localization 1 | Antigenicity (VaxiJen 2, ANTIGENPro 3) |
---|---|---|---|---|
1 | LinJ.35.1030 | casein kinase, putative | Cytoplasm | 0.5737/0.836 |
2 | LinJ.35.1540 | rieske iron-sulfuric protein, mitochondrion precursor | Cytoplasm | 0.5906/0.785 |
3 | LinJ.08.1290 | beta tubulin | Cytoplasm | 0.5436/0.764 |
4 | LinJ.21.2240 | beta tubulin | Cytoplasm | 0.5349/0.780 |
5 | LinJ.36.5660 | branch point binding protein, putative | Cytoplasm | 0.7933/0.917 |
6 | LinJ.08.1280 | beta tubulin | Cytoplasm | 0.5334/0.784 |
7 | LinJ.28.0780 | hypothetical protein | Cytoplasm | 0.5545/0.652 |
8 | LinJ.28.2940 | receptor for activated C kinase 1 | Extracellular | 0.6190/0.814 |
9 | LinJ.22.1300 | cyclophilin 6, putative | Extracellular | 0.5135/0.775 |
10 | LinJ.16.0470 | 60S ribosomal protein L21, putative | Mitochondria | 0.5437/0.665 |
11 | LinJ.20.0600 | conserved protein, unknown function | Mitochondria | 0.6737/0.735 |
12 | LinJ.34.0420 | hypothetical protein, conserved | Mitochondria | 0.5455/0.779 |
13 | LinJ.34.3620 | ribosomal protein L14, putative | Mitochondria | 0.8221/0.624 |
14 | LinJ.35.3810 | 60S ribosomal protein L27A/L29, putative | Mitochondria | 0.7317/0.754 |
15 | LinJ.24.0040 | 60S ribosomal protein L17, putative | Mitochondria | 0.6032/0.850 |
16 | LinJ.20.0250 | transmembrane protein, putative | Nucleus | 0.5086/0.576 |
17 | LinJ.18.0650 | serine/threonine kinase-like protein, putative | Nucleus | 0.5320/0.546 |
18 | LinJ.33.1560 | RNA-binding protein, putative | Nucleus | 0.5448/0.809 |
19 | LinJ.28.2200 | DNA-directed RNA polymerase-like protein | Nucleus | 0.5213/0.918 |
20 | LinJ.10.0050 | ribosomal protein l35a, putative | Nucleus | 0.6450/0.869 |
21 | LinJ.28.0210 | Histone H2B variant V | Nucleus | 0.5420/0.616 |
22 | LinJ.32.0930 | 60S ribosomal protein L18a, putative | Nucleus | 0.5872/0.509 |
23 | LinJ.33.3340 | small nuclear ribonucleoprotein SmD2 | Nucleus | 0.5678/0.534 |
24 | LinJ.36.6680 | 40S ribosomal protein S8, putative | Nucleus | 0.8525/0.616 |
25 | LinJ.21.0440 | Protein of unknown function (DUF667) | Nucleus | 0.5903/0.840 |
26 | LinJ.27.1450 | DNA-directed RNA polymerase II-like protein | Nucleus | 0.5001/0.731 |
No | Protein (Code) | No of MHCI Epitopes | No of MHCII Epitopes | ||
---|---|---|---|---|---|
Human | Mouse | Human | Mouse | ||
1 | LinJ.08.1290 | 2 | 13 | 4 | 38 |
2 | LinJ.08.1280 | 2 | 13 | 3 | 38 |
3 | LinJ.21.2240 | 2 | 13 | 3 | 40 |
4 | LinJ.22.1300 | 2 | 6 | 4 | 18 |
5 | LinJ.21.0440 | 3 | 6 | 5 | 24 |
6 | LinJ.36.6680 | 3 | 6 | 3 | 61 |
7 | LinJ.16.0470 | 0 | 5 | 0 | 2 |
8 | LinJ.32.0930 | 1 | 5 | 0 | 5 |
9 | LinJ.24.0040 | 2 | 4 | 0 | 9 |
10 | LinJ.34.0420 | 2 | 4 | 1 | 29 |
11 | LinJ.18.0650 | 0 | 4 | 2 | 35 |
12 | LinJ.28.2200 | 3 | 3 | 0 | 21 |
13 | LinJ.28.0780 | 1 | 3 | 0 | 50 |
14 | LinJ.35.1540 | 6 | 3 | 2 | 64 |
15 | LinJ.20.0250 | 5 | 3 | 1 | 37 |
16 | LinJ.36.5660 | 4 | 2 | 3 | 38 |
17 | LinJ.34.3620 | 1 | 2 | 3 | 55 |
18 | LinJ.33.1560 | 3 | 2 | 1 | 10 |
19 | LinJ.33.3340 | 1 | 2 | 5 | 3 |
20 | LinJ.35.3810 | 1 | 1 | 3 | 3 |
21 | LinJ.28.0210 | 1 | 1 | 4 | 44 |
22 | LinJ.10.0050 | 4 | 1 | 0 | 39 |
23 | LinJ.20.0600 | 0 | 1 | 6 | 25 |
24 | LinJ.27.1450 | 3 | 0 | 0 | 6 |
25 | LinJ.35.1030 | 2 | 11 | 4 | 26 |
26 | LinJ.28.2940 | 2 | 5 | 0 | 35 |
No | Protein (Code) | Number of MHCI Epitopes | Number of MHCII Epitopes | (%) Coverage of Amino Acid Sequence | Homology Human/Mouse |
---|---|---|---|---|---|
1 | LinJ.34.0420 | 4 | 2 | 30.8% | no |
2 | LinJ.18.0650 | 4 | 9 | 23.6% | 41.29/42.97 |
3 | LinJ.28.2200 | 3 | 4 | 46.7% | 40.26/29.92 |
4 | LinJ.28.0780 | 3 | 50 | 81.1% | no |
5 | LinJ.20.0600 | 1 | 14 | 38.2% | no |
Physicochemical Characteristic | Amino Acid Number | Molecular Weight | Instability Index | GRAVY | Half Life | Aliphatic Index | Theoretical pI |
---|---|---|---|---|---|---|---|
Score | 407 | 46,757.76 Da | 45.65 | −0.338 | >10 h (E. coli, in vivo) | 85.11 | 9.05 |
Protein | HLA Alleles (PDB ID) | Global Energy 1 | aVdW 2 | rVdW 3 | ACE 4 | HB 5 |
---|---|---|---|---|---|---|
LeishChim | HLA-A*0201 (PDB: 1I4F) | 2.37 | −4.43 | 0.79 | −0.01 | −0.22 |
HLA-A*0101 (PDB: 6AT9) | −12.54 | −19.50 | 10.78 | 3.91 | −2.85 | |
HLA-B*0702 (PDB: 5EO0) | 0.00 | 0.00 | 0.00 | 0.00 | 0.00 | |
HLA-B*3501 (PDB:2FYY) | 5.58 | −21.45 | 13.08 | 11.14 | −0.52 | |
HLA-DRB1*03:01 (PDB: 1A6A) | −22.06 | −40.07 | 21.56 | 11.65 | −3.75 | |
HLA-DRB5*01:01 (PDB: 1H15) | −2.69 | −34.44 | 38.98 | 1.97 | −4.19 | |
HLA-DRB1*01:01 (PDB: 2FSE) | −17.35 | −26.01 | 11.39 | 6.73 | −0.35 | |
HLA-DRB3*02:02 (PDB: 3C5J) | 0.43 | −6.65 | 3.13 | 2.47 | 0.00 |
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Margaroni, M.; Agallou, M.; Tsanaktsidou, E.; Kammona, O.; Kiparissides, C.; Karagouni, E. Immunoinformatics Approach to Design a Multi-Epitope Nanovaccine against Leishmania Parasite: Elicitation of Cellular Immune Responses. Vaccines 2023, 11, 304. https://doi.org/10.3390/vaccines11020304
Margaroni M, Agallou M, Tsanaktsidou E, Kammona O, Kiparissides C, Karagouni E. Immunoinformatics Approach to Design a Multi-Epitope Nanovaccine against Leishmania Parasite: Elicitation of Cellular Immune Responses. Vaccines. 2023; 11(2):304. https://doi.org/10.3390/vaccines11020304
Chicago/Turabian StyleMargaroni, Maritsa, Maria Agallou, Evgenia Tsanaktsidou, Olga Kammona, Costas Kiparissides, and Evdokia Karagouni. 2023. "Immunoinformatics Approach to Design a Multi-Epitope Nanovaccine against Leishmania Parasite: Elicitation of Cellular Immune Responses" Vaccines 11, no. 2: 304. https://doi.org/10.3390/vaccines11020304
APA StyleMargaroni, M., Agallou, M., Tsanaktsidou, E., Kammona, O., Kiparissides, C., & Karagouni, E. (2023). Immunoinformatics Approach to Design a Multi-Epitope Nanovaccine against Leishmania Parasite: Elicitation of Cellular Immune Responses. Vaccines, 11(2), 304. https://doi.org/10.3390/vaccines11020304