Evolution of Acetylcholinesterase and Butyrylcholinesterase in the Vertebrates: An Atypical Butyrylcholinesterase from the Medaka Oryzias latipes

Abstract

Acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) are thought to be the result of a gene duplication event early in vertebrate evolution. To learn more about the evolution of these enzymes, we expressed in vitro, characterized, and modeled a recombinant cholinesterase (ChE) from a teleost, the medaka Oryzias latipes. In addition to AChE, O. latipes has a ChE that is different from either vertebrate AChE or BChE, which we are classifying as an atypical BChE, and which may resemble a transitional form between the two. Of the fourteen aromatic amino acids in the catalytic gorge of vertebrate AChE, ten are conserved in the atypical BChE of O. latipes; by contrast, only eight are conserved in vertebrate BChE. Notably, the atypical BChE has one phenylalanine in its acyl pocket, while AChE has two and BChE none. These substitutions could account for the intermediate nature of this atypical BChE. Molecular modeling supports this proposal. The atypical BChE hydrolyzes acetylthiocholine (ATCh) and propionylthiocholine (PTCh) preferentially but butyrylthiocholine (BTCh) to a considerable extent, which is different from the substrate specificity of AChE or BChE. The enzyme shows substrate inhibition with the two smaller substrates but not with the larger substrate BTCh. In comparison, AChE exhibits substrate inhibition, while BChE does not, but may instead show substrate activation. The atypical BChE from O. latipes also shows a mixed pattern of inhibition. It is effectively inhibited by physostigmine, typical of all ChEs. However, although the atypical BChE is efficiently inhibited by the BChE-specific inhibitor ethopropazine, it is not by another BChE inhibitor, iso-OMPA, nor by the AChE-specific inhibitor BW284c51. The atypical BChE is found as a glycophosphatidylinositol-anchored (GPI-anchored) amphiphilic dimer (G2 a), which is unusual for any BChE. We classify the enzyme as an atypical BChE and discuss its implications for the evolution of AChE and BChE and for ecotoxicology.

Planarian cholinesterase: in vitro characterization of an evolutionarily ancient enzyme to study organophosphorus pesticide toxicity and reactivation

“…We note several similarities in substrate catalysis and inhibitor profiles for DjChE compared to cholinesterases from other invertebrate species, including Schistosoma (platyhelminths) (Bentley et al 2005), nematodes (Johnson and Russell 1983; Combes et al 2001), teleosts (Pezzementi et al 2011), and jawless fish (Sanders et al 1996). With the high turnover substrates, the acylcholine and acylthiocholine esters, mammalian AChEs exhibit substrate inhibition.…”

Butyrylcholinesterase as a biochemical marker

“…BChE is able to split acetylcholine, too. To distinguish whether the sample contains BChE or AChE, a specifi c inhibitor for BChE, tetraisopropyl pyrophosphoramide known also as iso-OMPA, can be added into the sample (79). BChE activity can be simply assayed by a titrimetric or potentiometric device as butyric acid is formed from a pH-neutral compound (64).…”

Electrochemical Sensors for the Estimation of the Inhibitory Effect of Phenylcarbamates to Cholinesterase

“…Both ChEs belong to a large protein family containing the α/β hydrolase fold. Individual amino acid residues involved in determining the molecular basis of the differences in substrate and inhibitor specificity of AChE and BChE have been identified in the acyl pocket, located at the bottom of a deep catalytic gorge; the peripheral anionic site, located at the lip of the gorge; the oxyanion hole; and the choline-binding site of the hydrophobic patch, also located within the gorge [11]. As described [9], the peripheral anionic site is a very important structural element, responsible for the binding of many inhibitors.…”