Cholinesterase Structure Identification of Residues and Domains Affecting Organophosphate Inhibition and Catalysis.

During this year we examined oxime reactivation of organophosphate modified cholinesterases using both achiral and chiral organophosphate inhibitors. The chiral inhibitors were resolved enantiomers of known absolute stereochemistry. This approach has been combined with site specific mutagenesis of the acyl pocket in the enzyme. Our studies clearly point to the importance of an impacted gorge in reducing the efficiency of the oxime reactivation reaction and show that the oximes attack with distinct orientations. Studies on the crystal structure of mouse AChE in the fasciculin complex, in the uncomplexed state and conjugated with organophosphates continue. Finally, we have initiated studies using cysteine mutagenesis to produce modified cholinesterases with altered catalytic characteristics. These studies are probing the dynamics and geometry of the active center of the enzyme while in solution. All of these studies are directed to converting acetylcholinesterase from a stoichiometric to a catalytic antidote for organophosphate toxicity.