Organophosphorus Hydrolases (OPH)

Use of Organophosphorus Hydrolases

Organophosphorous compounds are a broad class of chemicals that have found a variety of uses most commonly as pesticides and chemical warfare agents. These compounds act as neurotoxins upon exposure to humans and other higher organisms. All of these chemicals have similar chemical architectures that include phosphoester bonds of sulfur or oxygen.

In the United States, organophosphorous compounds have become the largest class of pesticides in use ever since organochlorine chemicals were banned. In excess of 5,000 metric tons of over 40 registered organophosphorous pesticides are used annually in the United States alone for applications ranging from commercial agriculture to homes and gardens to prevent infestation by insect pests.

Applications

For military applications, the United States and former Soviet Union have stockpiled large quantities of organophosphorous compounds as chemical warfare agents, with estimates approaching a combined 70,000 metric tons. By international treaty the countries currently in possession of chemical warfare agent stockpiles have a mandate to destroy these materials. Eliminating these chemicals with current technology is estimated to require years to complete and 10's of billions of dollars in cost.

There is considerable interest in developing methods to inactivate biologically active organophosphorous compounds. Of the various methods for decontamination, the most promising utilizes organophosphorous hydrolase (OPH) enzymes to render these toxins harmless. The rationale for a protein based remediation method is that no harsh chemicals or multi-step processes are required to convert organophosphorous agents to nontoxic chemical byproducts.

Recently, the United States Military in conjunction with Genencor announced an agreement whereby Genencor will produce and supply enzymes to the military for use in decontamination of organophosphorous based chemical warfare agents. Few products have been developed for decontaminating and managing non-military applications of organophosphorous agents. In spite of its potential, existing enzyme technology has a number of disadvantages including substrate inactivation, lack of enzyme stability, and narrow enzyme substrate specificity.

There is an ongoing need for new enzyme based systems that can overcome the limitations of current technology and deliver cost effective solutions to rendering organophosphorous compounds harmless for both military and non-military applications. Lybradyn has a number of new enzyme systems at various stages of development designed to address the most critical properties that limit existing enzyme based organophosphorous decontamination products.

Technology Summary of Lybradyn OPH's

Lybradyn, in collaboration with the University of Illinois at Chicago, is developing OPH solutions for military, agricultural, and consumer product applications. The technology of the OPH system involves a combination of novel, proprietary wild-type recombinant and engineered enzymes.

The Native OPH's are derived from soil thermophilic organisms including Bacillus and a thermally stable OPH enzyme from Deinococcus species, which have been cloned into production-ready E. coli systems. The enzymes have been shown to efficiently hydrolyze model organophosphorous compounds as well if not better than currently available enzymes from Pseudomonas. Advantages of these enzymes include resistance to substrate inactivation, thermal stability, solvent stability, long shelf lives, and broad substrate specificity. Lybradyn is currently optimizing these enzymes to increase substrate breadth and an increased and consistent rate of turnover.

Engineered enzymes have been developed based on known OPH enzymes and are being optimized using Lybradyn's expertise in evolutionary engineering, error prone PCR, saturation mutagenesis, and site directed mutagenesis techniques. The modifications to original enzymes are tracked and quantified through a combination of chemical assay and structural biology driven strategies to create improved performance characteristics. When coupled with engineered proprietary systems at Lybradyn, these enhanced enzymes will provide a product capable of decontaminating a broad array of organophosphorous compounds.

Lybradyn's expertise in fermentation process development enables design of microbial systems capable of producing OPH enzymes on manufacturing scale cost effectively. Expression and fermentation systems are optimized to include GRAS hosts, high cell densities, maximized protein yields, cost effective culture media formulations, efficient culture processes and downstream protein processing methods.