Characterization of a high-activity mutant of human butyrylcholinesterase against (-)-cocaine

Wenchao Yang, Liu Xue, Lei Fang, Xi Chen, Chang Guo Zhan

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53 Scopus citations

Abstract

Cocaine addiction and overdose are a well-known public health problem. There is no approved medication available for cocaine abuse treatment. Our recently designed and discovered high-activity mutant (A199S/S287G/A328W/Y332G) of human butyrylcholinesterase (BChE) has been recognized to be worth exploring for clinical application in humans as a potential anti-cocaine medication. The catalytic rate constant (kcat) and Michaelis-Menten constant (KM) for (-)-cocaine hydrolysis catalyzed by A199S/S287G/A328W/Y332G BChE (without fusion with any other peptide) have been determined to be 3060min-1 and 3.1μM, respectively, in the present study. The determined kinetic parameters reveal that the un-fused A199S/S287G/A328W/Y332G mutant has a ∼1080-fold improved catalytic efficiency (kcat/KM) against (-)-cocaine compared to the wild-type BChE. The ∼1080-fold improvement in the catalytic efficiency of the un-fused A199S/S287G/A328W/Y332G mutant is very close to the previously reported the ∼1000-fold improvement in the catalytic efficiency of the A199S/S287G/A328W/Y332G mutant fused with human serum albumin. These results suggest that the albumin fusion did not significantly change the catalytic efficiency of the BChE mutant while extending the plasma half-life. In addition, we have also examined the catalytic activities of the A199S/S287G/A328W/Y332G mutant against two other substrates, acetylthiocholine (ATC) and butyrylthiocholine (BTC). It has been shown that the A199S/S287G/A328W/Y332G mutations actually decreased the catalytic efficiencies of BChE against ATC and BTC, while considerably improving the catalytic efficiency of BChE against (-)-cocaine.

Original languageEnglish
Pages (from-to)148-152
Number of pages5
JournalChemico-Biological Interactions
Volume187
Issue number1-3
DOIs
StatePublished - Sep 2010

Bibliographical note

Funding Information:
This work was supported by NIH (grants R01 DA021416 , R01 DA025100 , and R01 DA013930 ). The authors also acknowledge the Center for Computational Sciences (CCS) at University of Kentucky for supercomputing time on an IBM X-series Cluster with 1360 processors.

Keywords

  • Cocaine addiction
  • Drug overdose
  • Enzyme therapy
  • Hydrolase

ASJC Scopus subject areas

  • Toxicology

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