TY - JOUR
AB - The hepatitis E virus (HEV) capsid protein pORF2 comprises three potential N‑linked glycosylation sites. One site, N562, is located at the cell attachment and neutralizing antigenic regions. The present study performed detailed analyses of the effects of specific amino acid substitutions at position 562 in the homodimerization, glycosylation, antigenicity, immunogenicity and neutralization activities of HEV pORF2. Recombinant HEV pORF2 glycoprotein E1 (amino acids 439‑617) and three mutant variants (N562L, N562C and N562K) were expressed in Pichia pastoris (P. pastoris) and SDS‑PAGE, Western blot analysis, tunicamycin assay, double‑antibody sandwich ELISA and in vitro PCR‑based neutralization assay were performed to characterize the different constructs. All proteins were indicated to be secreted by P. pastoris and formed homodimers. Tunicamycin assay revealed the glycosylated status of the wild‑type protein, but the mutants were indicated to be non‑glycosylated. All proteins were immunoreactive with a neutralizing monoclonal antibody but were not recognized by the antibody after denaturation into monomers. An in vitro PCR‑based neutralization assay using mouse antibodies indicated efficient neutralization against N562L, whereas antibodies against N562C and N562K were revealed to be non‑neutralizing. Collectively, the present study indicated that specific amino acid substitutions at position 562 serve crucial roles in the activity of the HEV neutralizing epitope.
AD - Medical Research and Laboratory Diagnostic Center, Jinan Central Hospital Affiliated to Shandong First Medical University and Shandong Academy of Medical Sciences, Jinan, Shandong 250013, P.R. China
Department of Liver Disease, Jinan Infectious Disease Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China
AU - Xu,Mingjie
AU - Sun,Lizhi
AU - Wang,Yan
AU - Gao,Shuchun
AU - Yang,Weihua
AU - Li,Meng
DA - 2021/02/01
DO - 10.3892/etm.2020.9542
IS - 2
JO - Exp Ther Med
KW - hepatitis E virus
N562
glycosylation
dimerization
neutralization
PY - 2021
SN - 1792-0981
1792-1015
SP - 110
ST - Different mutations at position 562 of the hepatitis E virus capsid proteins exhibit differential effects on viral neutralizing activity
T2 - Experimental and Therapeutic Medicine
TI - Different mutations at position 562 of the hepatitis E virus capsid proteins exhibit differential effects on viral neutralizing activity
UR - https://doi.org/10.3892/etm.2020.9542
VL - 21
ER -