Interaction between the peripheral site residues of human butyrylcholinesterase, D70 and Y332, in binding and hydrolysis of substrates

Masson, Patrick; Xie, Weihua; Froment, Marie Thérèse; Levitsky, Vladislav Yu.; Fortier, Pierre Louis; Albaret, Christine; Lockridge, Oksana

doi:10.1016/s0167-4838(99)00115-6

Cited by 80 publications

(77 citation statements)

References 45 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The simulated structures of the prereactive BChE-(−)-cocaine and BChE-(+)-cocaine complexes are similar to the prereactice enzyme-substrate structure proposed for BChE binding with other positively charged substrates, i.e. butyrylthiocholine and succinyldithiocholine [33,37]; they are all positioned horizontally at the bottom of the substrate-binding gorge of BChE.…”

Section: Catalytic Mechanism For Bche-catalyzed Hydrolysis Of Cocainesupporting

confidence: 55%

Rational design of an enzyme mutant for anti-cocaine therapeutics

Zheng

Chen

2007

J Comput Aided Mol Des

View full text Add to dashboard Cite

Summary(−)-Cocaine is a widely abused drug and there is no available anti-cocaine therapeutic. The disastrous medical and social consequences of cocaine addiction have made the development of an effective pharmacological treatment a high priority. An ideal anti-cocaine medication would be to accelerate (−)-cocaine metabolism producing biologically inactive metabolites. The main metabolic pathway of cocaine in body is the hydrolysis at its benzoyl ester group. Reviewed in this article is the stateof-the-art computational design of high-activity mutants of human butyrylcholinesterase (BChE) against (−)-cocaine. The computational design of BChE mutants has been based on not only the structure of the enzyme, but also the detailed catalytic mechanisms for BChE-catalyzed hydrolysis of (−)-cocaine and (+)-cocaine. Computational studies of the detailed catalytic mechanisms and the structure-and-mechanism-based computational design have been carried out through the combined use of a variety of state-of-the-art techniques of molecular modeling. By using the computational insights into the catalytic mechanisms, a recently developed unique computational design strategy based on the simulation of the rate-determining transition state has been employed to design highactivity mutants of human BChE for hydrolysis of (−)-cocaine, leading to the exciting discovery of BChE mutants with a considerably improved catalytic efficiency against (−)-cocaine. One of the discovered BChE mutants (i.e. A199S/S287G/A328W/Y332G) has a ~456-fold improved catalytic efficiency against (−)-cocaine. The encouraging outcome of the computational design and discovery effort demonstrates that the unique computational design approach based on the transition-state simulation is promising for rational enzyme redesign and drug discovery.

show abstract

Section: Catalytic Mechanism For Bche-catalyzed Hydrolysis Of Cocainesupporting

confidence: 55%

Rational design of an enzyme mutant for anti-cocaine therapeutics

Zheng

Chen

2007

J Comput Aided Mol Des

View full text Add to dashboard Cite

show abstract

“…Site-directed mutagenesis of human BChE cDNA was performed by using the QuikChange method (44). Mutations were generated from WT human BChE in a pRc͞CMV expression plasmid (45)(46)(47)(48)(49). Using plasmid DNA as template and primers with specific base pair alterations, mutations were made by PCR with Pfu DNA polymerase for replication fidelity.…”

Section: Site-directed Mutagenesis Protein Expression and Bche Actimentioning

confidence: 99%

Computational redesign of human butyrylcholinesterase for anticocaine medication

Pan

Gao

Yang

et al. 2005

Proc. Natl. Acad. Sci. U.S.A.

172

312

View full text Add to dashboard Cite

Molecular dynamics was used to simulate the transition state for the first chemical reaction step (TS1) of cocaine hydrolysis catalyzed by human butyrylcholinesterase (BChE) and its mutants. The simulated results demonstrate that the overall hydrogen bonding between the carbonyl oxygen of (؊)-cocaine benzoyl ester and the oxyanion hole of BChE in the TS1 structure for (؊)-cocaine hydrolysis catalyzed by A199S͞S287G͞A328W͞Y332G BChE should be significantly stronger than that in the TS1 structure for (؊)-cocaine hydrolysis catalyzed by the WT BChE and other simulated BChE mutants. Thus, the transition-state simulations predict that A199S͞ S287G͞A328W͞Y332G mutant of BChE should have a significantly lower energy barrier for the reaction process and, therefore, a significantly higher catalytic efficiency for (؊)-cocaine hydrolysis. The theoretical prediction has been confirmed by wet experimental tests showing an Ϸ(456 ؎ 41)-fold improved catalytic efficiency of A199S͞S287G͞A328W͞Y332G BChE against (؊)-cocaine. This is a unique study to design an enzyme mutant based on transitionstate simulation. The designed BChE mutant has the highest catalytic efficiency against cocaine of all of the reported BChE mutants, demonstrating that the unique design approach based on transition-state simulation is promising for rational enzyme redesign and drug discovery. molecular dynamics ͉ rational design ͉ transition-state stabilization ͉ cocaine ͉ enzyme-substrate binding C ocaine is recognized as the most reinforcing of all drugs of abuse (1-3). The disastrous medical and social consequences of cocaine addiction have made the development of an effective pharmacological treatment a high priority (4-6). However, cocaine mediates its reinforcing and toxic effects by blocking neurotransmitter reuptake, and the classical pharmacodynamic approach has failed to yield small-molecule receptor antagonists because of the difficulties inherent in blocking a blocker (1-5). An alternative to receptor-based approaches is to interfere with the delivery of cocaine to its receptors or accelerate its metabolism in the body (5,(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17). An ideal molecule for this purpose should be a potent enzyme catalyzing the hydrolysis of cocaine into biologically inactive metabolites. The dominant pathway for cocaine metabolism in primates is butyrylcholinesterase (BChE)-catalyzed hydrolysis at the benzoyl ester group (Fig. 3, which is published as supporting information on the PNAS web site), and the metabolites are all biologically inactive (5, 18). Clearly, BChE-catalyzed hydrolysis of cocaine at the benzoyl ester is the metabolic pathway most suitable for amplification. However, the catalytic activity of this plasma enzyme is Ϸ1,000-fold lower against the naturally occurring (Ϫ)-cocaine than that against the biologically inactive (ϩ)-cocaine enantiomer (19)(20)(21)(22). (ϩ)-cocaine can be cleared from plasma in seconds, before partitioning into the CNS, whereas (Ϫ)-cocaine has a plasma half-life of Ϸ45-90 min, long enough for ...

show abstract

“…Site-directed mutagenesis of human BChE cDNA was performed by using the QuikChange method (Braman et al, 2000). Further mutation(s) required to produce a new BChE mutant cDNA was/were generated from the cDNA corresponding to the A199S/S287G/A328W/Y332G mutant of human BChE in a pRc/CMV expression plasmid (Masson et al, 1999). Using plasmid DNA as template and primers with specific base-pair alterations, mutations were made by polymerase chain reaction with Pfu DNA polymerase for replication fidelity.…”

Section: Introductionmentioning

confidence: 99%

Design, Preparation, and Characterization of High-Activity Mutants of Human Butyrylcholinesterase Specific for Detoxification of Cocaine

Xue¹,

Ko²,

Tang³

et al. 2010

Mol Pharmacol

104

View full text Add to dashboard Cite

Cocaine is a widely abused drug without a U.S. Food and Drug Administration-approved medication. There is a recognized, promising anticocaine medication to accelerate cocaine metabolism, producing biologically inactive metabolites via a route similar to the primary cocaine-metabolizing pathway [i.e., cocaine hydrolysis catalyzed by butyrylcholinesterase (BChE) in plasma]. An ideal, therapeutically valuable mutant of human BChE should have not only a significantly improved catalytic activity against (Ϫ)-cocaine but also certain selectivity for (Ϫ)-cocaine over neurotransmitter acetylcholine (ACh), such that one would not expect systemic administration of the BChE mutant to interrupt cholinergic transmission. The present study accounting for the mutation-caused changes of the catalytic activities of BChE against both (Ϫ)-cocaine and ACh by means of molecular modeling and site-directed mutagenesis has led to identification of three BChE mutants that have not only a considerably improved catalytic efficiency against (Ϫ)-cocaine but also the desirable selectivity for (Ϫ)-cocaine over ACh. Two representative BChE mutants have been confirmed to be potent in actual protection of mice from acute toxicity (convulsion and lethality) of a lethal dose of cocaine (180 mg/kg). Pretreatment with the BChE mutant (i.e., 1 min before cocaine administration) dose-dependently protected mice against cocaineinduced convulsions and lethality. In particular, all mice pretreated with the mutant (e.g., 0.02 mg or more of A199S/F227A/S287G/ A328W/E441D BChE) survived. The in vivo data reveal the primary factor (i.e., the relative catalytic efficiency), determining the efficacy in practical protection of mice from the acute cocaine toxicity and future direction for further improving the efficacy of the enzyme in the cocaine overdose treatment.

show abstract

Interaction between the peripheral site residues of human butyrylcholinesterase, D70 and Y332, in binding and hydrolysis of substrates

Cited by 80 publications

References 45 publications

Rational design of an enzyme mutant for anti-cocaine therapeutics

Rational design of an enzyme mutant for anti-cocaine therapeutics

Computational redesign of human butyrylcholinesterase for anticocaine medication

Design, Preparation, and Characterization of High-Activity Mutants of Human Butyrylcholinesterase Specific for Detoxification of Cocaine

Contact Info

Product

Resources

About