The Systematic Prediction Of Antigenic Epitopes For Human Enteroviruses 71 And Coxasckievirus A16

Objective:Bioinformatics methods were developed for the prediction of antigenic epitopes of human enteroviruses and applied to predict the epitopes of Enterovirus 71(EV71)and Coxasckievirus A16(CVA16).A routine prediction workflow was established for human enteroviruses' B cellular conformational epitopes and linear epitopes.The conformational epitopes and linear epitopes of EV71 and CVA16 were systematically predicted and compared between these two serotypes of the enteroviruses.The structural differences(including the primary structure,the secondary structure,the tertiary structure and the capsid structure)between EV71 and CVA16 were explored.Finally,these epitopes were focused on during the study of the pathogenic mechanism of the enteroviruses.Methods:1.The structural proteins' tertiary structure files of the representative strains of EV71 and CVA16 were downloaded from RCSB PDB database.The amino acid sequences of these representative strains and other thirty EV71 strains with known genotypes were downloaded from NCBI Nucleotide database.2.The Bioinformatics method for the prediction of human enteroviruses' conformational epitopes was developed and used to predict and compare the epitopes of EV71 and CVA16 systematically.Softwares and online tools such as MUSCLE,Clustal X,ESPript,Pymol,UCSF Chimera and RIVEM were performed to compare the primary structures,the secondary structures,the tertiary structures and the capsid structures.Based on these analyses,the structural differences of the epitopes of EV71 and CVA16 were determined.The conservative analysis for the epitopes was also performed among the subtypes of the serotypes.The reported neutralizing antibodies and the binding areas of the enteroviruses' receptors were obtained via literature retrieval.The predicted accuracy was estimated with the reported data.Furthermore,the focus was put on the role of the epitopes in the pathogenic mechanism of the enteroviruses.3.The Bioinformatics method for the prediction of human enteroviruses' linear epitopes was developed and used to predict and compare the epitopes of EV71 and CVA16 systematically.The relationship between the conformational epitopes and the linear epitopes was explored.The conservative analysis for the epitopes was also performed among the subtypes of the serotypes.Results:1.The conformational epitopes of EV71 and CVA16 were systematically predicted by the developed Bioinformatics method and had a high prediction accuracy.Both the distribution of the conformational epitopes of EV71 and CVA16 had the same pattern.The epitopes consist of three sites: site 1(including sub-site site 1a,site 1b and site 1c),site 2(including sub-site site 2a,site 2b and site 2c),site 3(including sub-site site 3a and site 3b).Each site or part of the site can work as the conformational epitopes independently.2.The secondary structure properties of the conformational epitopes of EV71 and CVA16 were loops between ? sheets,and a few were N-terminal or C-terminal.The positions of the epitopes of EV71 and CVA16 were almost same and the differences were the amino acid residues.The amino acid residues were conservative among the intra-serotypes and different among the inter-serotypes.In a word,the conformational epitope had the serotype specificity.Most conformational epitopes lie around the two sides of canyon.VP1 was the most important structural protein for the conformational epitopes and about half of the residues of the epitopes was from VP1.Most residues of site 1 and part of site 2 were from VP1.3.The amino acid sequences of EV71 and CVA16 had a high homology.The secondary structures of EV71 were also similar to CVA16.The backbones of the tertiary structures of EV71 and CVA16 overlapped to each other,especially for VP2.There were six positions of the backbones of VP1 that did not overlap,five of which were related to the conformational epitopes.According to VP3,four positions did not overlap and two of them were related to the conformational epitopes.Altogether,these positions lie on loops.4.The binding area of the receptors of EV71 overlapped with the conformational epitopes.The binding area of SCARB2 was around the two sides of the canyon and overlapped with site 1a and site 2b while PSGL-1,HS and Anx2 overlapped with site 1.5.The linear epitopes of EV71 and CVA16 distributed widely on the surfaces of the virus caspids.The conformational epitopes and the linear epitopes of the viruses had tight relationship.Each linear epitope had more or less residues from the conformational epitopes.The positions of the linear epitopes of EV71 and CVA16 were almost same.However,the similarity of the sequences were rather low.It hinted that the linear epitopes of EV71 and CVA16 also had the serotypes specificity.Conclusions:1.The developed Bioinformatics methods for the prediction of the conformational and the linear epitopes worked excellently and were helpful for the research on the antigenic epitopes of human enteroviruses.The distribution patterns of the conformational and the linear epitopes of EV71 and CVA16 were simple.The conformational epitopes consist of three sites and the linear epitopes distribute widely on the capsid surfaces.Loops were the most important secondary structure for the epitopes.2.The relationship between the conformational and the linear epitopes of EV71 and CVA16 was quite tight.Each and part of the sites of the conformational epitopes can work as the antigenic epitopes while each linear epitope covered part of the conformational epitopes.That meant the core residues of the conformational epitopes were covered by the linear epitopes.3.Although the distribution patterns of the conformational and the linear epitopes of EV71 and CVA16 were highly similar,the residues of the epitopes were different between EV71 and CVA16 and led to the antigenic difference between the serotypes.