Exploring And Designing Of Broad-spectrum Antigen Of Hand-foot-mouth Disease Vaccine

Hand,foot and mouth disease(HFMD)is a contagious disease,which is easy to break out and mainly affects infants.It is a serious public health problem.The development of a vaccine against HFMD is of great importance for the prevention and control of the outbreak of HFMD.There are many kinds of viruses that cause hand,foot and mouth disease,enterovirus 71,Coxsackie virus A group 4,5,9,10 and B group 2 and 5.But the most common viruses are enterovirus 71,Coxsackie virus A group 6 and A group 16 type.Peptide vaccines have become a hotspot in the research and development of safe vaccines because of their good biological safety and high immune response.At the same time,the recombinant protein vaccine is favored by researchers because of its high safety and easy production.Firstly,the amino acid homology analysis of VP1 nucleotide sequences of 5616 C V-A6,3969 CV-A16 and 8818 EV71 on the NCBI website was carried out by MEG A,respectively,and the results were presented on the protein structure using Py MOL.Two CV-A6 polypeptides,two CV-A16 polypeptides and three EV71 polypeptides werescreened out,which were CV-A6-PEP1(S143-L150): STTPGMLL,CV-A6-PEP2(K96-T106): KDSGTSLDGYT,CV-A16-PEP3(P96-Y106): PTTGTQNTDGY,CV-A16-PEP4(P142-Q149): PNGELVPQ,EV71-PEP5(Y208-Y222)YPTFGEHKQEKDLEY,EV71-PE P6(A139-Q152): ACTPTGEVVPQLLQ and EV71-PEP7(P96-Y106): PLKGTTNPNG Y.The above amino acid sequences were synthesized into peptides,dissolved in PBS,and the peptide nanoparticles were prepared by means of ethanol desolving.The pepti des were mixed with the same amount of Freund’s incomplete adjuvant to construct th e peptide vaccine.Secondly,constructing plasmids CV-A6-VP1-pFastBac Dual,CV-A16-VP1-pFastBac Dual,and EV71-VP1-pFastBac Dual were transformed into Escherichia coli DH10 Bac for reorganization,respectively.SF9 insect cells were infected with the restructuring baculovirus to induce the expression of VP1 protein.Three kinds of VP1 proteins were purified by Ni column affinity chromatography.The purified proteins were mixed with the same amount of Freund’s incomplete adjuvant,and the recombinant HFMD vaccine was successfully constructed.Finally,BALB/c mice were immunized intraperitoneally with the two vaccines,respectively,to evaluate the immune response induced by CV-A6-VP1,CV-A16-VP1,EV71-VP1 recombinant protein vaccines and peptide nanoparticles.ELISA results showed that mice immunized with VP1 recombinant protein and peptide of the same antigen could induce specific antibodies against VP1 recombinant protein of the same antigen and corresponding peptide.This study will improve understanding of the design of protein and peptide antigens and advance the search for a vaccine for HFMD.