Identification of neutralizing epitopes on the D/A domain of the E2 glycoprotein of classical swine fever virus

Yu-Liang Huang; Denise Meyer; Alexander Postel; Kuo-Jung Tsai; Hsin-Meng Liu; Chia-Huei Yang; Yu-Chun Huang; Hui-Wen Chang; Ming-Chung Deng; Fun-In Wang; Paul Becher; Helen Crooke; Chia-Yi Chang

doi:10.1016/j.virusres.2023.199209

Identification of neutralizing epitopes on the D/A domain of the E2 glycoprotein of classical swine fever virus

Virus Res. 2023 Oct 15:336:199209. doi: 10.1016/j.virusres.2023.199209. Epub 2023 Aug 31.

Authors

Affiliations

¹ WOAH Reference Laboratory for Classical Swine Fever, Veterinary Research Institute, Ministry of Agriculture, 376 Chung-Cheng Road, Tamsui, New Taipei City 25158, Taiwan.
² WOAH Reference Laboratory for Classical Swine Fever, Institute of Virology, University of Veterinary Medicine Hannover, 30559 Hannover, Germany.
³ School of Veterinary Medicine, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei 10617, Taiwan.
⁴ WOAH Reference Laboratory for Classical Swine Fever, Animal and Plant Health Agency, New Haw, Surrey, KT15 3NB, UK. Electronic address: Helen.Crooke@apha.gov.uk.
⁵ School of Veterinary Medicine, National Taiwan University, No. 1, Section 4, Roosevelt Road, Taipei 10617, Taiwan. Electronic address: chiayichang@ntu.edu.tw.

Abstract

Classical swine fever virus (CSFV) shares high antigenic homology with other members of the genus Pestivirus. Because several pestivirus species can also infect swine, eliciting cross-reactive antibodies, it is important to define CSFV-specific epitopes for the differential diagnosis of classical swine fever (CSF) by serology. For this purpose, epitope mapping of seven monoclonal antibodies (mAbs), recognizing sites on the D/A domain of glycoprotein E2, was performed using recombinant expressed antigenic domains and mutants of E2, as well as an overlapping peptide library. Three CSFV-specific epitopes, i.e., 780-IEEMGDDFGFGLCPF-794, 810-NGSAFYLVCPIGWTG-824, and 846-REKPF-850, were identified within the D/A domain of E2. Site-directed mutagenesis further confirmed that residues 783-MGD-785, 789-FGLCPF-794, 813-AFYLVCPIGWTG-824, and 846-REK-848 were critical residues in these regions. In addition, a F789S difference within the epitope 780-IEEMGDDFGFGLCPF-794 was responsible for the absence of binding of two mAbs to the E2 protein of the live attenuated CSFV vaccine strain Riems. Structural modeling revealed that, the three epitopes are located near each other, suggesting that they may form a more complex conformational epitope on the D/A domain in vivo. Six of the mAbs neutralized viruses of diverse genotypes, indicating that the target epitopes are involved in virus interaction with cells. The binding of CSFV to cells was significantly reduced after pre-incubation with either truncated E2 proteins comprising the D/A domain or with the CSFV-specific mAbs targeting the domain D/A. These epitopes identified on the D/A domain are important targets for virus neutralization that might be involved in the early steps of CSFV infection. These findings reveal potential candidates for improving the differential diagnosis of pestiviruses by serology.

Keywords: Classical swine fever virus; Conformational epitope; Epitope mapping; Glycoprotein E2; Pestivirus; Virus neutralization.