Structural Analysis Of Antigenic Epitopes On Envelope Glycoprotein Of Japanese Encephalitis Virus

Japanese encephalitis (JE), caused by Japanese encephalitis virus (JEV) that belongs to the family Flaviviridae,is a zoonose transmitted by mosquitoes and seriously threatens the health of human beings and animals. As the the main structure protein of JEV, the envelope glycoprotein (E) is closely related to viral attachment and fusion with host cell, and determines the virus's hemagglutination ability, cellular tropism, viral virulence and induction of protective immune response. Referring to the sequence of SA14-14-2, we designed a series of peptides by deleting animo acids one by one from N-terminal and C-terminal of six antigenic epitopes (E1,E11,E19,E33,E39 and E49 on E protein) which were identified by our team. The oligo nucleotides chains corresponding to the peptides were synthesized. After fusion-expressed with GST in a pGEX-6p-1 vector and western blot analysis using JEV positive serum, the core sequences of the six antigenic epitopes were identified as E1(⁷GNRDFIEG¹⁴),E11(⁸³EKRADSS⁸⁹),E19(¹⁵⁰ENHGNYS¹⁵⁶),E33(²⁵⁹EGGLHHA²⁶⁵),E39(³¹⁰FAKNPVD³¹⁶) and E49(³⁸⁵VGRGDK³⁹⁰).The sequence of epitope E19 was included in a reported neutralization epitope (¹⁴⁹SENHGNYSAQVGASQ¹⁶³), so we deduced E19 antigenic epitope may have neutral activity.Flavivirus is the biggest family of Flaviviridae and classified into eight serology subfamilies, including more than 70 kinds of viruses, such as JEV, Dengue virus (DV), West Nile virus (WNV) and Kunjin virus (KUNV), all of which have serious cross reaction with each other. The diseases caused by these viruses always have similar clinical symptoms like fever and encephalitis, and we still have no perfect diagnostic techniques to differentiate them. In this study, bioinformatics software was used to compare the homologous sequences of the identified antigenic epitopes of 24 JEV strains and 12 strains of flavivirus. 47 mutated peptides were designed and expressed with GST. After immunoblot assays, we found that the epitopes E1, E10, E19 and E33 had the potential abilities to distinguish JEV with other flaviviruses.This study will provide important basis for further structural and functional research of JEV envelope protein, development of new epitope vaccine and diagnostic techniques.