Mutant Proteins--Enzymes to Hydrolyze Toxic Organophosphates

摘要

This work focuses on the creation of new catalytic activities by modification ofexisting enzymes. To this end, techniques for the synthesis of large genes have been developed together with a complementation system for the expression and correct folding of proteins that are normally self-processing but that have mutated so they can no longer process themselves. (Concurrent expression of the pro-sequence and the mature protein sequence encoded on separate but compatible plasmids accomplishes this purpose; the pro-sequence apparently performs the role of a chaperonin in this case). The enzymes studied are two serine hydrolases, alpha protease and beta-lactamase. For a-lytic proteases the new activity sought was the ability to hydrolyze organophosphates rather than being irreversibly inhibited by these substances. For beta lactamase we created a structural variant that had a dramatically different catalytic role on the normal beta lactam substrates.