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Fusion to green fluorescent protein improves expression levels of Theileria parva sporozoite surface antigen p67 in insect cells

Published online by Cambridge University Press:  11 February 2003

S. A. KABA
Affiliation:
Laboratory of Virology, Wageningen University, Binnenhaven 11, 6709 PD, Wageningen, The Netherlands International Livestock Research Institute, P.O. Box 30709, Nairobi, Kenya
V. NENE
Affiliation:
International Livestock Research Institute, P.O. Box 30709, Nairobi, Kenya Current address: The Institute for Genomic Research, Rockville, USA.
A. J. MUSOKE
Affiliation:
International Livestock Research Institute, P.O. Box 30709, Nairobi, Kenya
J. M. VLAK
Affiliation:
Laboratory of Virology, Wageningen University, Binnenhaven 11, 6709 PD, Wageningen, The Netherlands
M. M. VAN OERS
Affiliation:
Laboratory of Virology, Wageningen University, Binnenhaven 11, 6709 PD, Wageningen, The Netherlands

Abstract

East Coast fever (ECF) is a fatal disease of cattle caused by the protozoan parasite Theileria parva. The development of a subunit vaccine, based on the sporozoite-specific surface antigen p67, has been hampered by difficulties in achieving high-level expression of recombinant p67 in a near-authentic form. Therefore two sets of recombinant baculovirus vectors were constructed. The first set, encoding various regions of p67, produced low levels of the corresponding p67 domains in High Five™ cells, despite the presence of large amounts of p67 RNA. The second, consisting of p67 domains fused to the carboxy-terminus of GFP expressed significantly higher levels of p67 protein. The GFP[ratio ]p67 fusion proteins were recognized by a sporozoite-neutralizing monoclonal antibody (TpM12) raised against native p67 whereas non-fused full length p67 expressed in insect cells was not recognized. GFP-tagging therefore, appeared to enhance the stability of p67 and to conserve its folding. The high-level expression of p67 domains in a more authentic form is an important step towards the development of an effective subunit vaccine against ECF.

Type
Research Article
Copyright
© 2002 Cambridge University Press

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