Gnatt_1994_J.Neurochem_62_749

Reference

Title : Site-directed mutagenesis of active site residues reveals plasticity of human butyrylcholinesterase in substrate and inhibitor interactions - Gnatt_1994_J.Neurochem_62_749
Author(s) : Gnatt A , Loewenstein Y , Yaron A , Schwarz M , Soreq H
Ref : Journal of Neurochemistry , 62 :749 , 1994
Abstract : In search of the molecular mechanisms underlying the broad substrate and inhibitor specificities of butyrylcholinesterase (BCHE), we employed site-directed mutagenesis to modify the catalytic triad residue Ser198, the acyl pocket Leu286 and adjacent Phe329 residues, and Met437 and Tyr440 located near the choline binding site. Mutant proteins were produced in microinjected Xenopus oocytes, and Km values towards butyrylthiocholine and IC50 values for the organophosphates diisopropylfluorophosphonate (DFP), diethoxyphosphinylthiocholine iodide (echothiophate), and tetraisopropylpyrophosphoramide (iso-OMPA) were determined. Substitution of Ser198 by cysteine and Met437 by aspartate nearly abolished activity, and other mutations of Ser198 completely abolished it. Tyr440 and Leu286 mutants remained active, but with higher Km and IC50 values. Rates of inhibition by DFP were roughly parallel to IC50 values for several Leu286 mutants. Both Km and IC50 values increased for Leu286 mutants in the order Asp < Gln < Lys. In contrast, cysteine, leucine, and glutamine mutants of Phe329 displayed unmodified Km values toward butyrylthiocholine, but up to 10-fold decreased IC50 values for DFP, iso-OMPA, and echothiophate. These findings add Tyr440 and Phe329 to the list of residues interacting with substrate and ligands, demonstrate plasticity in the active site region of BCHE, and foreshadow the design of recombinant BCHEs with tailored scavenging properties.
ESTHER : Gnatt_1994_J.Neurochem_62_749
PubMedSearch : Gnatt_1994_J.Neurochem_62_749
PubMedID: 8294937

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Citations formats

Gnatt A, Loewenstein Y, Yaron A, Schwarz M, Soreq H (1994)
Site-directed mutagenesis of active site residues reveals plasticity of human butyrylcholinesterase in substrate and inhibitor interactions
Journal of Neurochemistry 62 :749

Gnatt A, Loewenstein Y, Yaron A, Schwarz M, Soreq H (1994)
Journal of Neurochemistry 62 :749