Fig 1.
The experimental process of this study.
Table 1.
Physicochemical properties of amino acids.
Fig 2.
Homologous sequence alignment of proteins.
The blue portions represent similar amino acid sequence (The darker the blue, the more conservative it is).
Fig 3.
(A-B) Protein signal peptide was analyzed by SignalP6.0.
Other: the probability that the sequence does not have any signal peptides.
Table 2.
Physical properties, antigenicity and scores of CTL dominant epitopes.
Table 3.
Physical properties, antigenicity and scores of HTL dominant epitopes.
Fig 4.
B cell conformational epitope predicted by Ellipro.
The yellow spheres represent B cell conformational epitopes, the gray area represents the majority of the polyprotein. (A)Rv0986 conformational epitope residues:V228,N229,R230, E231,N232,Q233,T234,D235,Q236,P237,A238,S239,T240,I241,L242,L243,P244,T245,S246,Y247,E248. (B)PknD conformational epitope residues:D242,S243,D244,R245,T246,T261,S262,L263, E264,H265,H266,H267.
Table 4.
Physical properties, antigenicity and scores of LBEs dominant epitopes.
Table 5.
Physical properties, antigenicity and scores of CBEs dominant epitopes.
Fig 5.
Docking complex display (A)Molecular docking between ITAPWGIAV(CTLs) and HLA-A*02:01(B)Molecular docking between FQFFNLIPTLTVLEN(HTLs) and HLA-DRB1*07:01.
Fig 6.
(A-B)Epitopes and their corresponding MHC allele interaction using the LIGPLOT.
Hydrogen bonds are represented by dotted green lines, and red semicircular circles indicate residues involved in hydrophobic interactions.
Fig 7.
Vaccine construct from N-terminal to C-terminal.
Table 6.
The physiochemical profiling of the mRNA vaccine.
Fig 8.
(A)Immunoglobulins in various states (B)The Helper T Cell Population in various states (C)The Helper T Cell Population in various states (D)The Cytotoxic T Cell Population in various states (E)The B cell population in various states (F)The B Cell Population in various states (G)Dendritic Cell Population in various states (H)Macrophage Population in various states (I)Cytokines and Interleukins Production with Simpson Index of the immune response.
Fig 9.
Codon optimization and plasmid vector construction (A)Sequence after adaptation (B)The cloned MEV was inserted into the PVAX1 vector (C)After amplified(1161bp).
Fig 10.
(A)In silico cloning simulation. Codon-optimized multiepitope sequences (red) inserted between the restriction sites BamHI and XhoI in the PVAX1 expression vector (black). (B)Simulated agarose gel electrophoresis results. “1” stands for MEV-PCR,“2”stand for PVAX1,“3” stand for recombinant plasmid.
Fig 11.
(A)Optimal secondary structure (B)Centroid secondary structure.
Fig 12.
(A-B)Prediction of vaccine secondary structure(C)Optimize the pre-tertiary structure(D)Optimize the post-tertiary structure.
Fig 13.
(A)The Ramachsndran diagram was analyzed using PROCHECK (B)Analyze the Z-score using the Pro-SA server. (C) Energy diagram using the Pro-SA server.
Fig 14.
Molecular docking result (A)Docking complex, blue is the vaccine structure, green is the TLR4 receptor. (B)The interaction of MEV-TLR4 docking complex was demonstrated using PyMol. (C)The interaction of MEV-TLR4 complex and its 2D image were analyzed by Ligplot.
Fig 15.
Molecular dynamics simulation results.
(A) The RMSD locus of the receptor, ligand, and complex. The abscissa is the running time of MD simulation, and the ordinate is RMSD-value. (B-F) The RMSF locus of acceptor-ligand, the horizontal coordinate is the amino acid residue base in the docking complex, the ordinate is the rmsf value. (G-I) The trajectory of complex hydrogen bond, Gyrate and SASA respectively.