Production of Hev b5 as a fluorescent biotin-binding tripartite fusion protein in insect cells
- Department of Biological and Environmental Science, NanoScience Center, P.O. Box 35, FIN-40014 University of Jyvaeskylae (Finland)
- AI Virtanen Institute, Department of Molecular Medicine, University of Kuopio, P.O. Box 1627, FIN-70211 Kuopio (Finland)
- Institute of Biotechnology, University of Helsinki, P.O. Box 56 (Viikinkaari 9), FIN-00014 Helsinki (Finland)
The presented green fluorescent protein and streptavidin core-based tripartite fusion system provides a simple and efficient way for the production of proteins fused to it in insect cells. This fusion protein forms a unique tag, which serves as a multipurpose device enabling easy optimization of production, one-step purification via streptavidin-biotin interaction, and visualization of the fusion protein during downstream processing and in applications. In the present study, we demonstrate the successful production, purification, and detection of a natural rubber latex allergen Hev b5 with this system. We also describe the production of another NRL allergen with the system, Hev b1, which formed large aggregates and gave small yields in purification. The aggregates were detected at early steps by microscopical inspection of the infected insect cells producing this protein. Therefore, this fusion system can also be utilized as a fast indicator of the solubility of the expressed fusion proteins and may therefore be extremely useful in high-throughput expression approaches.
- OSTI ID:
- 20711021
- Journal Information:
- Biochemical and Biophysical Research Communications, Vol. 336, Issue 1; Other Information: DOI: 10.1016/j.bbrc.2005.08.095; PII: S0006-291X(05)01786-9; Copyright (c) 2005 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA); ISSN 0006-291X
- Country of Publication:
- United States
- Language:
- English
Similar Records
Efficient and versatile one-step affinity purification of in vivo biotinylated proteins: Expression, characterization and structure analysis of recombinant human glutamate carboxypeptidase II
Crystallization and identification of the glycosylated moieties of two isoforms of the main allergen Hev b 2 and preliminary X-ray analysis of two polymorphs of isoform II