Abstract
Cholinesterases are among the most efficient enzymes known. They are divided into two groups: acetylcholinesterase, involved in the hydrolysis of the neurotransmitter acetylcholine, and butyrylcholinesterase of unknown function. Several crystal structures of the former have shown that the active site is located at the bottom of a deep and narrow gorge, raising the question of how substrate and products enter and leave. Human butyrylcholinesterase (BChE) has attracted attention because it can hydrolyze toxic esters such as cocaine or scavenge organophosphorus pesticides and nerve agents. Here we report the crystal structures of several recombinant truncated human BChE complexes and conjugates and provide a description for mechanistically relevant non-productive substrate and product binding. As expected, the structure of BChE is similar to a previously published theoretical model of this enzyme and to the structure of Torpedo acetylcholinesterase. The main difference between the experimentally determined BChE structure and its model is found at the acyl binding pocket that is significantly bigger than expected. An electron density peak close to the catalytic Ser(198) has been modeled as bound butyrate.
Highlights
Cholinesterases are divided into two subfamilies according to their substrate and inhibitor specificities: acetylcholinesterase (AChE1; EC 3.1.1.7) and butyrylcholinesterase (BChE; EC 3.1.1.8)
Crystallographic Analysis of Uninhibited Butyrylcholinesterase—The BChE crystal structure was originally solved by the molecular replacement method and subsequently refined to 2.0-Å resolution using CNS
In AChEs, the four helices involved in the subunit-subunit interaction are antiparallel and the active site openings are located on opposite sides of the dimer
Summary
AchE, acetylcholinesterase; BchE, butyrylcholinesterase; TcAChE, Torpedo californica acetylcholinesterase; DmAChE, Drosophila melanogaster acetylcholinesterase; BTC, butyrylthiocholine; Bicine, N,N-bis(2-hydroxyethyl)glycine; MES, 4-morpholineethanesulfonic acid. There was a great deal of controversy concerning the nature of the residues involved in this site Both the crystal structure and labeling experiments showed that positively charged ligands form -cation interactions with Phe330 and Trp (numbering in italics corresponds to that of torpedo AChE) (10). The physiological role of BChE remains unclair (11, 12) It is capable of hydrolyzing ACh and other acylcholines, so far no endogenous natural substrate has been described for this enzyme. We report several crystal structures of BChE complexed with a substrate, products, and conjugated to soman after aging. From these structures we propose alternative substrate and product binding that may be related to the high catalytic efficiency of the choline esterases
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