Crystallographic Studies of Human Acetylcholinesterase Reactivation by Oximes: Towards a Neutron Structure

Abstract

Human acetylcholinesterase (hAChE) is responsible for degrading neurotransmitter acetylcholine at synapses of the nervous system. Organophosphate (OP) nerve agents and pesticides inactivate hAChE through chemical modifications of the enzyme's active site leading to a life‐threatening increase of acetylcholine concentration. Exposure to OPs is fatal if not treated with antidotes. However, the current generation of antidotes is not highly efficient, and the rates of reactivation are far slower than the catalytic rate of acetylcholine hydrolysis.Insights into the molecular structures of Torpedo (1) and mammalian AChEs (2) as well as other cholinesterases reveal the underlying limitations in enhancing reactivation rates but are limited in their ability to resolve positions of protons and hydrogens important for understanding of proton transfer in reactivation of organophosphate inhibited cholinesterases. The only experimental method that can provide this information is macromolecular neutron crystallography. In combination with X‐ray crystallography it is capable of providing insights into the active sites of enzymes with the atomic resolution.In preparation for the neutron diffraction studies we have obtained several new high resolution X‐ray structures of hAChE at room and low temperatures in the apo form and in complex with oxime reactivators and other ligands. The enzyme crystallizes in a novel crystal form with the unit cell amenable to neutron crystallography (space group P31, a=b=124.3, c=129.1 Å). Detailed analysis of the reactivators and ligand interactions at the enzyme active will be presented.Support or Funding InformationThis work was supported by the CounterACT Program, National Institutes of Health Office of the Director, Grant 1U01NS083451 from NINDS.