CHIA-YI CHANGHuang, Chin-ChengChin-ChengHuangDeng, Ming-ChungMing-ChungDengHuang, Yeou-LiangYeou-LiangHuangLin, Yu-JuYu-JuLinLiu, Hsin-MengHsin-MengLiuLin, Yeou-LiangYeou-LiangLinFUN-IN WANG2022-11-112022-11-11201201681702https://www.scopus.com/inward/record.uri?eid=2-s2.0-84864408569&doi=10.1016%2fj.virusres.2012.06.013&partnerID=40&md5=13fb40228755b7554e621e2e750c8086https://scholars.lib.ntu.edu.tw/handle/123456789/624620Envelope glycoprotein E2 of classical swine fever virus (CSFV) is the major antigen that induces neutralizing antibodies in infected pigs. Previous studies revealed that both conformation-dependent and linear epitopes are most present within domains B/C/D/A in the N-terminal half of E2. However, studies of antigenicity beyond the B/C domains remain limited. This study revealed that conformational epitopes were present on the D/A domains as well as the proximal C-terminal of E2, since the mutation of cysteine abrogated their bindings to monoclonal antibodies (mAbs). The residue R845 at domain A and E902 at the C-terminal region were critical for specific binding to mAbs, further supporting the presence of antigenic determinants on these regions. Substitutions of cysteines in domains D/A not only abrogated the binding to mAbs directed to D/A, but also affected the binding of the downstream C-terminal region to its specific mAbs, suggesting a close interaction between the two conformational epitopes. Mutations on the five proximal cysteines at positions 869, 877, 893, 896 and 930 in the C-terminal region only affected the binding to its specific mAbs binding sites. In addition, mutation on the three distal C-terminal cysteines at positions 945, 966, and 983 resulted in loss of E2 homodimerization. This study demonstrates new antigenic epitopes on D/A domains and C-terminal of E2 that have not been reported before, and that the nine cysteines in the C-terminal function differently in either maintaining the antigenic structure or in intermolecular dimerization of E2. © 2012 Elsevier B.V.Antigenic specificity; Classical swine fever virus; Conformational epitope; E2 glycoprotein; Homodimerizationcysteine; epitope; glycoprotein E2; virus envelope protein; amino acid substitution; antigen binding; antigen specificity; article; Baculovirus; binding site; carboxy terminal sequence; conformational transition; controlled study; dimerization; nonhuman; Pestivirus; priority journal; protein domain; protein expression; protein interaction; site directed mutagenesis; virus mutation; Amino Acid Motifs; Amino Acid Sequence; Animals; Antibodies, Viral; Cell Membrane; Classical Swine Fever; Classical swine fever virus; Epitope Mapping; Molecular Sequence Data; Protein Structure, Tertiary; Swine; Viral Envelope Proteins; Classical swine fever virus; Suidaejournal article10.1016/j.virusres.2012.06.013227276852-s2.0-84864408569