Development of a chitinase and v-cathepsin negative bacmid for improved integrity of secreted recombinant proteins

The application of the baculovirus-in sect cell expression system for the production of integral membrane and secreted proteins is often more troublesome than for cytoplasmic proteins. One protein expressed at low levels in insect cells is the Theileria parva sporozoite surface protein p67. Theileria parva is a protozoan parasite, which causes the tick-transmitted disease East Coast fever in cattle. Baculovirus vectors were engineered to produce a secreted form of p67 by replacing the signal peptide of p67 with the honeybee mellitin signal sequence and deleting a putative membrane anchor from the C-terminus. Furthermore, the chitinase and v-cathepsin genes were deleted from the baculovirus expression vector in a bacmid setup, allowing broad scale application of this novel vector. Deletion of the chitinase and v-cathepsin gene had a positive effect on the integrity of both the intracellular and secreted recombinant protein. (C) 2004 Elsevier B.V. All rights reserved.