Structural representation of JEV E epitopes defined by alanine-scanning mutagenesis and HDX. (A) JEV E-DIII epitopes for each of the eight mouse and human JEV MAbs were defined by alanine-scanning mutagenesis (shaded gray boxes) and HDX (boldface letters). Genotypic differences from the JEV-SA14-14-2 strain (GIII) are highlighted by a star: V315 is A in the JEV-2372/79 (GI), JEV-MAR 859 (GI), JEV-Bennett (GII), and JEV-Nakayama (GIII) strains; S327 is T in the JEV-2372/79 (GI), JEV-MAR 859 (GI), and JEV-Bennett (GIII) strains; K336 is N in the JEV-2372/79 (GI) and JEV-MAR 859 (GI) strains; and A366 is S in the JEV-2372/79 (GI), JEV-MAR 859 (GI), and JEV-Bennett (GIII) strains. For comparison to the JEV E-DIII epitopes, immediately below we show the structurally defined E-DIII epitopes of ZIKV in complex with ZV-2 (green, ABDE epitope), ZV-48 (cyan, C-C') and ZV-67 (magenta, lateral ridge [LR]), WNV E16 (magenta, LR), DV1-E106 (magenta, LR), DV1-E111 (cyan, C-C′ loop), DV2-1A1D-2 (pink, A strand), DV3-2H12 (light green, AB loop), and DV4-4E11 (pink, A strand). (B) JEV E-DIII epitopes defined by alanine-scanning mutagenesis are depicted on the JEV E-DIII structure (based on the full-length JEV E structure, PDB accession no. 3P54). (C) JEV epitopes defined by alanine-scanning mutagenesis, HDX mapping, and surface exposure are shown in the context of the full-length JEV E structure.