Free-energy perturbation simulation on transition states and redesign of butyrylcholinesterase

Wenchao Yang, Yongmei Pan, Fang Zheng, Hoon Cho, Hsin Hsiung Tai, Chang Guo Zhan

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55 Scopus citations


It is recognized that an ideal anti-cocaine treatment is to accelerate cocaine metabolism by producing biologically inactive metabolites via a route similar to the primary cocaine-metabolizing pathway, i.e., butyrylcholinesterase (BChE)-catalyzed hydrolysis of cocaine. BChE mutants with a higher catalytic activity against (-)-cocaine are highly desired for use as an exogenous enzyme in humans. To develop a rational design for high-activity mutants, we carried out free-energy perturbation (FEP) simulations on various mutations of the transition-state structures in addition to the corresponding free-enzyme structures by using an extended FEP procedure. The FEP simulations on the mutations of both the free-enzyme and transition-state structures allowed us to calculate the mutation-caused shift of the free-energy change from the free enzyme (BChE) to the transition state, and thus to theoretically predict the mutation-caused shift of the catalytic efficiency (kcat/K M). The computational predictions are supported by the kinetic data obtained from the wet experiments, demonstrating that the FEP-based computational design approach is promising for rational design of high-activity mutants of an enzyme. One of the BChE mutants designed and discovered in this study has an ∼1800-fold improved catalytic efficiency against (-)-cocaine compared to wild-type BChE. The high-activity mutant may be therapeutically valuable.

Original languageEnglish
Pages (from-to)1931-1938
Number of pages8
JournalBiophysical Journal
Issue number5
StatePublished - 2009

Bibliographical note

Funding Information:
This work was supported by a research grant from the National Institutes of Health (R01 DA013930 to C.-G.Z.). The entire work was performed at the University of Kentucky. W. Yang worked in C.-G. Zhan's laboratory at the University of Kentucky as an exchange graduate student from Central China Normal University. The authors also acknowledge the Center for Computational Sciences, University of Kentucky, for providing supercomputing time on the Superdome (an HP shared-memory supercomputer, with four nodes for 256 processors) and the IBM X-series Cluster (with 340 nodes and 1360 processors).

ASJC Scopus subject areas

  • Biophysics


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