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. 2004 Feb 11;182(2):765–773. doi: 10.1016/0042-6822(91)90617-K

Intracellular processing of the porcine coronavirus transmissible gastroenteritis virus spike protein expressed by recombinant vaccinia virus

David J Pulford 1,1, Paul Britton 1,2
PMCID: PMC7131517  PMID: 1850927

Abstract

The Spike (S) protein from a virulent British field isolate of porcine transmissible gastroenteritis virus (TGEV) FS772/70 was constructed from cDNA and inserted into the vaccinia virus (VV) thymidine kinase gene locus under the control of the VV early/late gene P7.5K promoter. Recombinant S protein was synthesized as an endo-β-N-acetylglucosamini-dase H (Endo H)-sensitive glycoprotein with high mannose simple oligosaccharides (gp190) that underwent post-translational modification to an Endo H-resistant glycoprotein with complex oligosaccharides (gp210). Immunofluorescence analysis demonstrated that the majority of recombinant S protein was retained at the Golgi but some S protein was expressed on the plasma membrane. Monoclonal antibodies (mAbs) raised against native S protein reacted with this recombinant S protein; also, mice infected with the recombinant vaccinia virus (rVV) expressing the S protein induced TGEV neutralizing antibodies. A truncated S protein (SΔ) was also expressed in rVV-infected cells by introducing a deletion into the S protein cDNA that removed 292 amino acids from the C-terminus. The SΔ protein (gpl70) was shown to be antigenically similar to TGEV S protein by immunofluorescence and immunoprecipitation tests but was retained in the endoplasmic reticulum and not expressed on the cell surface.

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