Expression of biologically active human butyrylcholinesterase in the cabbage looper (Trichoplusia ni)

This investigation examined the utility of three recombinant protein-expres sion systems (COS cells, insect cells and insect larvae) to cost-effectivel y produce biologically active human butyrylcholinesterase (BuChE), It was d etermined that baculovirus-infected insect cells (Sf9 and High 5) expressed 3.5- and 8.2-fold, respectively, more active enzyme than COS-7 cells. Bacu lovirus-infected cabbage looper (Trichoplusia ni) insert larvae produced ov er 26 times more than High 5 cells; in fact, one baculovirus-infected insec t larva provided more active protein than 100 mi of insect cell culture. An alysis of the larvally expressed proteins revealed that the vast majority o f BuChE expressed was inactive due to extensive degradation that occurred i n vivo. However, the active form of BuChE does have enzyme kinetics similar to those of its human serum counterpart. Cabbage looper larvae were also e xamined for their ability to serve as an in vivo animal model to study prot ection against anti-cholinesterase toxicity. This was unsuccessful due to t heir high tolerance to the very toxic organophosphorus compounds tested. Th is tolerance may be attributed at least in part to a novel endogenous organ ophosphorus acid anhydride hydrolase activity that is capable of hydrolysin g the chemical-warfare nerve agents sarin (isopropyl methylphosphonofluorid ate) and soman (pinacolyl methylphosphonofluoridate). These results show th at cabbage looper larvae can serve as an inexpensive recombinant protein-ex pression system for human BuChE.