Expression of the rotavirus SA11 protein VP7 in the simple eukaryote Dictyostelium discoideum
العنوان: | Expression of the rotavirus SA11 protein VP7 in the simple eukaryote Dictyostelium discoideum |
---|---|
المؤلفون: | Keith L. Williams, J M Miller, Harry B. Greenberg, Kerry R. Emslie, P R Dormitzer, Martin B. Slade |
المصدر: | Journal of Virology. 69:1747-1754 |
بيانات النشر: | American Society for Microbiology, 1995. |
سنة النشر: | 1995 |
مصطلحات موضوعية: | Signal peptide, Glycosylation, viruses, Protein subunit, Genetic Vectors, Molecular Sequence Data, Immunology, Biology, Antibodies, Viral, medicine.disease_cause, Microbiology, Dictyostelium discoideum, law.invention, Endoglycosidase H, Capsid, fluids and secretions, law, Virology, Rotavirus, medicine, Animals, Dictyostelium, Cloning, Molecular, Antigens, Viral, DNA Primers, Gel electrophoresis, Vaccines, Synthetic, Base Sequence, Molecular mass, Antibodies, Monoclonal, virus diseases, biology.organism_classification, Molecular biology, Recombinant Proteins, Insect Science, Recombinant DNA, biology.protein, Capsid Proteins, Research Article |
الوصف: | The outer capsid protein of rotavirus, VP7, is a major neutralization antigen and is considered a necessary component of any subunit vaccine developed against rotavirus infection. For this reason, significant effort has been directed towards producing recombinant VP7 that maintains the antigenic characteristics of the native molecule. Using a relatively new expression system, the simple eukaryote Dictyostelium discoideum, we have cloned the portion of simian rotavirus SA11 genome segment 9, encoding the mature VP7 protein, downstream of a native D. discoideum secretion signal sequence in a high-copy-number extrachromosomal vector. The majority of the recombinant VP7 expressed by transformants was intracellular and was detected by Western immunoblot following gel electrophoresis as two or three bands with an apparent molecular mass of 35.5 to 37.5 kDa. A small amount of VP7 having an apparent molecular mass of 37.5 kDa was secreted. Both the intracellular VP7 and the secreted VP7 were N glycosylated and sensitive to endoglycosidase H digestion. Under nonreducing electrophoresis conditions, over half the intracellular VP7 migrated as a monomer while the remainder migrated with an apparent molecular mass approximately twice that of the monomeric form. In an enzyme-linked immunosorbent assay, intracellular VP7 reacted with both nonneutralizing and neutralizing antibodies. The monoclonal antibody recognition pattern paralleled that found with VP7 expressed in either vaccinia virus or herpes simplex virus type 1 and confirms that D. discoideum-expressed VP7 is able to form the major neutralization domains present on viral VP7. Because D. discoideum cells are easy and cheap to grow, this expression system provides a valuable alternative for the large-scale production of recombinant VP7 protein. |
تدمد: | 1098-5514 0022-538X |
الوصول الحر: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::ead35bf8642dca0843d16c003a8def3dTest https://doi.org/10.1128/jvi.69.3.1747-1754.1995Test |
حقوق: | OPEN |
رقم الانضمام: | edsair.doi.dedup.....ead35bf8642dca0843d16c003a8def3d |
قاعدة البيانات: | OpenAIRE |
تدمد: | 10985514 0022538X |
---|