cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells

Du Kyung Kang, Ki Wan Kim, Pyeung Hyun Kim, Seung Yong Seoung, Yong Hee Kim, Ick Chan Kwon, Seo Young Jeong, Eui Yeol Choi, Kyung Mee Lee, Hyun Sook Kim, Eui Chong Kim, Sai Ick Joo, Jai Myung Yang

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5 Citations (Scopus)

Abstract

Rotavirus is a major cause of severe gastroenteritis in young children and animals throughout the world. The VP7 of rotavirus is thought to induce the synthesis of neutralizing antibodies and to be responsible for determining viral serotypes. The cDNA coding for the VP7 capsid protein of human rotavirus, obtained from Korean patients (HRV-Y14), was cloned and its nucleotide sequence was determined. Comparative analysis of the nucleotide sequences between VP7 of Y14 and that of other foreign isolates showed 92.7%- 95.2% homology to G1 serotypes (RV-4, KU, K8, WA), 74.2% homology to G2 serotype HU-5, 76.4% homology to G3 serotype SA-11, and 77.6% homology to G4 serotype A01321. These data suggest that HRV-Y14 can be classified as a G1 serotype. cDNA coding for VP7 of HRV-Y14 was subcloned into the baculovirus vector and the VP7 glycoprotein was expressed in insect cells. The expressed proteins in Sf9 cell extract and tissue culture fluid were separated on SDS- PAGE, and Western blot analysis with monoclonal antibody raised against the synthetic peptide containing 21 amino acids within the VP7 conserved region was performed. The molecular weight of recombinant VP7 was estimated to be 36 kDa which is about the same size as the native VP7. Addition of tunicamycin in the culture media caused a reduction of the molecular weight of the recombinant VP7 indicating that the expressed protein was glycosylated.

Original languageEnglish
Pages (from-to)369-377
Number of pages9
JournalJournal of microbiology and biotechnology
Volume8
Issue number4
Publication statusPublished - 1998

Keywords

  • Baculovirus
  • Glycosylation
  • Homology
  • Rotavirus
  • VP7

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology

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  • Cite this

    Kang, D. K., Kim, K. W., Kim, P. H., Seoung, S. Y., Kim, Y. H., Kwon, I. C., Jeong, S. Y., Choi, E. Y., Lee, K. M., Kim, H. S., Kim, E. C., Joo, S. I., & Yang, J. M. (1998). cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells. Journal of microbiology and biotechnology, 8(4), 369-377.