Optimized Platforms for Proper Glycosylation and Sialylation of Recombinant Human Butyrylcholinesterase (rBChE)

Organophosphorus (OP) toxins and chemical warfare agents (CWA) are active nerve agents posing significant hazard for both soldiers and civilians. Butyrylcholinesterase (BChE) is a safe and effective bioscavenger that detoxifies OP and CWAs when injected either prophylactically or post-exposure; but a major technical problem has been producing a recombinant form of BChE with acceptable properties for human use, namely tetramerization and glycosylation for high activity and long-life in the blood. Our proposal capitalizes on over 30 years of research experience in protein engineering and glycosylation using baculovirus and novel insect hosts. First we will express BChE in an established glycoengineered baculovirus-insect system for production of recombinant proteins with fully humanized glycosylation patterns, including sialylation. Next, we will prepare a polyproline fusion protein with sequences known to induce stable tetramer formation. Finally, we combine these elements to evaluate the tetramerization and activity of BChE after affinity purification. We have assembled a unique set of technologies and experience to establish feasibility during this Phase I project. If successful, our proposal will create a system for commercializing active BChE as a biopharmaceutical treatment for neurotoxin exposure.