Observing The Active Gorge Of AChE By The Atomic Force Microscopy

Acetylcholinesterase (AChE) is an essential enzyme in the cholinergic nervous system. Its principal biological role is to terminate the synaptic signal by catalyzing the hydrolysis reaction of the neurotransmitter, acetylcholine (ACh). Understanding the pharmacological mechanism of AChE to hydrolyze ACh rapidly is of great importance in life science. The newly developed atomic force microscopy (AFM) has opened a new window to the microworld of cells, subcellular structures and biomolecules. In the present study, we observed the structure of the native AChE reconstituted in phospholipid membrane and the structure of AChE reacted with ACh by AFM respectively, in order to make clear the relationship of the rapid hydrolyzing mechanism of AChE to its ultrastructure. The results showed that the appearance of the native AChE was elliptic and smooth, the average long-diameter was94nm, the average short-diameter was 53.01nm and the average height was 3.359nm.The appearance of AChE after reacted with ACh was rough and the shape became irregular. Most particles had holes in the middle. These results demonstrated that AFM can be used to study the structure of enzyme and the reaction of enzyme with its substrate. The phenomenon that the particles of AChE had holes after reacted with ACh suggests that the active site gorge of AChE probably has two states: close-open or narrow-broad. The gorge is closed or narrow when there are no ACh and is open or wide when there is ACh released from the presynaptic ending. The substrate ACh has an active counter-reaction on the ultrastructure of AChE. It makes the active gorge open or widened to fascilitates more ACh to enter the active site through the active gorge to meet the needs of the rapid hydrolyzing function of the enzyme.