Abstract
Enterovirus (EV71) can cause severe neurological diseases, but the underlying pathogenesis remains unclear. The capsid protein, viral protein 1 (VP1), plays a critical role in the pathogenicity of EV71. High level expression and secretion of VP1 protein are necessary for structure, function and immunogenicity in its natural conformation. In our previous studies, 5 codon-optimized VP1 DNA vaccines, including wt-VP1, tPA-VP1, VP1-d, VP1-hFc and VP1-mFc, were constructed and analyzed. They expressed VP1 protein, but the levels of secretion and immunogenicity of these VP1 constructs were significantly different (P<0.05). In this study, we further investigated the protein levels of these constructs and determined that all of these constructs expressed VP1 protein. The secretion level was increased by including a tPA leader sequence, which was further increased by fusing human IgG Fc (hFc) to VP1. VP1-hFc demonstrated the most potent immunogenicity in mice. Furthermore, hFc domain could be used to purify VP1-hFc protein for additional studies.
Highlights
Hand foot and mouth disease (HFMD), which is caused by enterovirus 71 (EV71), is often associated with severe neurological diseases[1,2], including poliomyelitis-like paralysis, fatal encephalitis with cardiopulmonary complications, meningitis and brain stem encephalitis, which occur mainly in infants and young children[3]
When leader sequence from human tissue plasminogen activator was included as a fusion with viral protein 1 (VP1), we observed an increase in the expression of VP1 protein
To compare the protein expression levels of the 4 VP1 DNA vaccines and to determine if the other 2 VP1 constructs were secreted, the 4 VP1 constructs were transfected under identical conditions
Summary
Hand foot and mouth disease (HFMD), which is caused by enterovirus 71 (EV71), is often associated with severe neurological diseases[1,2], including poliomyelitis-like paralysis, fatal encephalitis with cardiopulmonary complications, meningitis and brain stem encephalitis, which occur mainly in infants and young children[3]. EV71 infections in the AsiaPacific region have shown increasing incidence and mortality[4,5]. The family of enteroviruses shares similar morphological characteristics, containing 60 copies of each of the 4 structural proteins viral protein (VP) 1 to 4. Of these 4 proteins, VP1 is more exposed than the other 3 proteins and plays a critical role in the pathogenicity of EV71[9]. VP1 contains the major epitopes recognized by neutralizing antibodies[11,12] and is used to make subunit vaccines[13,14]. The VP1 gene is serotype-specific and considered the most suitable region for sequence analysis[15,16]. The canyon hypothesis[10,17] suggests that a hydrophobic pocket contained within
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