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<p>The spike protein is a structural protein of coronaviruses and can be divided into an amino terminal S1 domain and a carboxyl terminal S2 domain. The S1 domain binds to receptors on the cell surface, so it contains major epitopes for neutralizing antibodies and is an important immunogen for vaccine development. Vaccine research and protein structure—function studies require pure protein preparations, ideally with no tag appended. In this study, we designed constructs encoding porcine epidemic diarrhea virus (PEDV) S1 protein with tissue plasminogen activator signal peptide and a Protein Select™ tag at the amino terminus. We showed the expression of PEDV S1 protein in the supernatants of mammalian cell cultures after transfection. We further showed the purification of tag-free PEDV S1 protein upon tag cleavage by the Protein Select™ resin from Cytiva. The purified PEDV S1 protein was then characterized by deglycosylation, dynamic light scattering, size-exclusion chromatography with multi-angle light scattering, one- and two-dimensional native-PAGE, and circular dichroism. The immune reactivity of the purified PEDV S1 protein to a hyper-immune serum was demonstrated in an enzyme-linked immunosorbent assay (ELISA). Taken together, we have expressed, purified, and comprehensively characterized PEDV spike S1 protein without a tag.</p>

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Efficient design and production of glycosylated PEDV spike S1 protein

  • 期刊论文

  • 出版模式：

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  • 发表日期：

    2026年6月3日

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  • Eva-Maria Uhlemann,
  • Rinku Patel,
  • Brittany Thivierge,
  • Glenn Hamonic,
  • Wolfgang Köster,
  • Trina Racine,
  • Qiang Liu

摘要

The spike protein is a structural protein of coronaviruses and can be divided into an amino terminal S1 domain and a carboxyl terminal S2 domain. The S1 domain binds to receptors on the cell surface, so it contains major epitopes for neutralizing antibodies and is an important immunogen for vaccine development. Vaccine research and protein structure—function studies require pure protein preparations, ideally with no tag appended. In this study, we designed constructs encoding porcine epidemic diarrhea virus (PEDV) S1 protein with tissue plasminogen activator signal peptide and a Protein Select™ tag at the amino terminus. We showed the expression of PEDV S1 protein in the supernatants of mammalian cell cultures after transfection. We further showed the purification of tag-free PEDV S1 protein upon tag cleavage by the Protein Select™ resin from Cytiva. The purified PEDV S1 protein was then characterized by deglycosylation, dynamic light scattering, size-exclusion chromatography with multi-angle light scattering, one- and two-dimensional native-PAGE, and circular dichroism. The immune reactivity of the purified PEDV S1 protein to a hyper-immune serum was demonstrated in an enzyme-linked immunosorbent assay (ELISA). Taken together, we have expressed, purified, and comprehensively characterized PEDV spike S1 protein without a tag.