title: Kinetic characterization of human butyrylcholinesterase mutants for the hydrolysis of cocaethylene.
abstract: It is known that the majority of cocaine users also consume alcohol. Alcohol can react with cocaine to produce a significantly more cytotoxic compound, cocaethylene. Hence a truly valuable cocaine-metabolizing enzyme as treatment for cocaine abuse/overdose should be efficient for not only cocaine itself, but also cocaethylene. The catalytic parameters (kcat and KM) of human BChE (butyrylcholinesterase) and two mutants (known as cocaine hydrolases E14-3 and E12-7) for cocaethylene are characterized in the present study, for the first time, in comparison with those for cocaine. On the basis of the obtained kinetic data, wild-type human BChE has a lower catalytic activity for cocaethylene (kcat=3.3 min(-1), KM=7.5 μM and kcat/KM=4.40 × 10(5) M(-1)·min(-1)) compared with its catalytic activity for (-)-cocaine. E14-3 and E12-7 have a considerably improved catalytic activity against cocaethylene compared with the wild-type BChE. E12-7 is identified as the most efficient enzyme for hydrolysing cocaethylene in addition to its high activity for (-)-cocaine. E12-7 has an 861-fold improved catalytic efficiency for cocaethylene (kcat=3600 min(-1), KM=9.5 μM and kcat/KM=3.79 × 10(8) M(-1)·min(-1)). It has been demonstrated that E12-7 as an exogenous enzyme can indeed rapidly metabolize cocaethylene in rats. Further kinetic modelling has suggested that E12-7 with an identical concentration as that of the endogenous BChE in human plasma can effectively eliminate (-)-cocaine, cocaethylene and norcocaine in simplified kinetic models of cocaine abuse and overdose associated with the concurrent use of cocaine and alcohol.
authors.affiliation: *Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536, U.S.A.
authors.forename: Shurong
authors.initials: S
authors.lastname: Hou
authors.affiliation: *Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536, U.S.A.
authors.forename: Max
authors.initials: M
authors.lastname: Zhan
authors.affiliation: *Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536, U.S.A.
authors.forename: Xirong
authors.initials: X
authors.lastname: Zheng
authors.affiliation: *Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536, U.S.A.
authors.forename: Chang-Guo
authors.initials: CG
authors.lastname: Zhan
authors.affiliation: *Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, KY 40536, U.S.A.
authors.forename: Fang
authors.initials: F
authors.lastname: Zheng
mesh_terms: D000818:Animals; D002091:Butyrylcholinesterase / Q000235:genetics* / Q000378:metabolism*; D016466:CHO Cells; D002384:Catalysis; D003042:Cocaine / Q000031:analogs & derivatives* / Q000378:metabolism / Q000493:pharmacokinetics; D019970:Cocaine-Related Disorders / Q000378:metabolism; D003412:Cricetulus; D000431:Ethanol / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D008297:Male; D062105:Molecular Docking Simulation; D051381:Rats
chemical_list: D000431:Ethanol; C008671:norcocaine; D002091:Butyrylcholinesterase; C066444:cocaethylene; D003042:Cocaine