OPTIMIZED PRODUCTION OF BACTERIAL AGENT-DEGRADING ENZYMES

A NUMBER OF ORGANOPHOSPHORUS COMPOUNDS ARE HIGHLY TOXIC AS A RESULT OF THEIR ABILITY TO INHIBIT CHOLINESTERASES. THIS TOXICITY HAS LED TO THE WIDESPREAD USE OF THESE COMPOUNDS AS PESTICIDES AND SUGGESTS THEIR POTENTIAL USE AS AGENTS OF CHEMICAL WARFARE. A NUMBER OF BACTERIAL SPECIES ELABORATE ENZYMES, TERMED ORGANOPHOSPHORUS ACID (OPA) ANHYDRASES, CAPABLE OF DETOXIFYING THESE COMPOUNDS. THE GOALS OF THE PROPOSED RESEARCH WILL BE TO DETERMINE UNDER WHAT CONDITIONS OF GROWTH MAXIMUM OPA ANHYDRASE ACTIVITY CAN BE OBTAINED WITH SELECTED BACTERIAL STRAINS AND TO DEVELOP PRELIMIARY METHODS FOR THE ISOLATION OF ENZYMES POSSESSING THIS ACTIVITY FROM THESE STRAINS. THESE GOALS WILL BE ACCOMPLISHED BY: 1) DEVELOPING METHODS FOR THE RAPID AND EFFICIENT SCREENING OF STRAINS FOR THEIR LEVELS OF OPA ANHYDRASE ACTIVITY 2) PERFORMING LARGE SCALE SCREENING OF CULTURES TO DETERMINE THE OPTIMUM GROWTH CONDITIONS FOR HIGH LEVEL ENZYME ACTIVITY, 3) DETERMINING WHETHER THE CONDITIONS SO ESTABLISHED WILL SCALE-UP FROM TUBE OR SHAKE FLASK CULTURES TO SMALL AND MEDIUM SCALE FERMENTORS, AND 4) DEVELOPING PRELIMINARY PROTOCOLS FOR THE PURIFICATION OF OPA ANHYDRASEES FROM THESE ORGANISMS. SUCCESSFUL COMPLETION OF THIS WORK WILL AID IN ESTABLISHING THE FEASIBILITY OF USING THESE ORGANISMS OR GENES FROM THESE ORGANISMS FOR THE LARGE SCALE PRODUCTION OF ENZYMES FOR DETOXIFICATION OF ORGANOPHORPHORUS COMPOUNDS.