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

Research output: Contribution to journalArticle

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 Jan 1
Externally publishedYes

Fingerprint

Rotavirus
Cloning
Capsid Proteins
Insects
Organism Cloning
Nucleotides
Complementary DNA
Molecular weight
Proteins
Tunicamycin
Tissue culture
Neutralizing Antibodies
Culture Media
Glycoproteins
Animals
Monoclonal antibodies
Monoclonal Antibodies
Amino Acids
Antibodies
Peptides

Keywords

  • Baculovirus
  • Glycosylation
  • Homology
  • Rotavirus
  • VP7

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology
  • Microbiology

Cite this

Kang, D. K., Kim, K. W., Kim, P. H., Seoung, S. Y., Kim, Y. H., Kwon, I. C., ... 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.

cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells. / Kang, Du Kyung; Kim, Ki Wan; Kim, Pyeung Hyun; Seoung, Seung Yong; Kim, Yong Hee; Kwon, Ick Chan; Jeong, Seo Young; Choi, Eui Yeol; Lee, Kyung Mee; Kim, Hyun Sook; Kim, Eui Chong; Joo, Sai Ick; Yang, Jai Myung.

In: Journal of Microbiology and Biotechnology, Vol. 8, No. 4, 01.01.1998, p. 369-377.

Research output: Contribution to journalArticle

Kang, DK, Kim, KW, Kim, PH, Seoung, SY, Kim, YH, Kwon, IC, Jeong, SY, Choi, EY, Lee, KM, Kim, HS, Kim, EC, Joo, SI & Yang, JM 1998, 'cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells', Journal of Microbiology and Biotechnology, vol. 8, no. 4, pp. 369-377.
Kang DK, Kim KW, Kim PH, Seoung SY, Kim YH, Kwon IC et al. cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells. Journal of Microbiology and Biotechnology. 1998 Jan 1;8(4):369-377.
Kang, Du Kyung ; Kim, Ki Wan ; Kim, Pyeung Hyun ; Seoung, Seung Yong ; Kim, Yong Hee ; Kwon, Ick Chan ; Jeong, Seo Young ; Choi, Eui Yeol ; Lee, Kyung Mee ; Kim, Hyun Sook ; Kim, Eui Chong ; Joo, Sai Ick ; Yang, Jai Myung. / cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells. In: Journal of Microbiology and Biotechnology. 1998 ; Vol. 8, No. 4. pp. 369-377.
@article{eb30224a91c345669f121c8011173eb8,
title = "cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells",
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.",
keywords = "Baculovirus, Glycosylation, Homology, Rotavirus, VP7",
author = "Kang, {Du Kyung} and Kim, {Ki Wan} and Kim, {Pyeung Hyun} and Seoung, {Seung Yong} and Kim, {Yong Hee} and Kwon, {Ick Chan} and Jeong, {Seo Young} and Choi, {Eui Yeol} and Lee, {Kyung Mee} and Kim, {Hyun Sook} and Kim, {Eui Chong} and Joo, {Sai Ick} and Yang, {Jai Myung}",
year = "1998",
month = "1",
day = "1",
language = "English",
volume = "8",
pages = "369--377",
journal = "Journal of Microbiology and Biotechnology",
issn = "1017-7825",
publisher = "Korean Society for Microbiolog and Biotechnology",
number = "4",

}

TY - JOUR

T1 - cDNA cloning and expression of human rotavirus outer capsid protein VP7 in insect cells

AU - Kang, Du Kyung

AU - Kim, Ki Wan

AU - Kim, Pyeung Hyun

AU - Seoung, Seung Yong

AU - Kim, Yong Hee

AU - Kwon, Ick Chan

AU - Jeong, Seo Young

AU - Choi, Eui Yeol

AU - Lee, Kyung Mee

AU - Kim, Hyun Sook

AU - Kim, Eui Chong

AU - Joo, Sai Ick

AU - Yang, Jai Myung

PY - 1998/1/1

Y1 - 1998/1/1

N2 - 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.

AB - 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.

KW - Baculovirus

KW - Glycosylation

KW - Homology

KW - Rotavirus

KW - VP7

UR - http://www.scopus.com/inward/record.url?scp=7344257738&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=7344257738&partnerID=8YFLogxK

M3 - Article

VL - 8

SP - 369

EP - 377

JO - Journal of Microbiology and Biotechnology

JF - Journal of Microbiology and Biotechnology

SN - 1017-7825

IS - 4

ER -