Preparation and in vivo characterization of a cocaine hydrolase engineered from human butyrylcholinesterase for metabolizing cocaine

Liu Xue, Shurong Hou, Min Tong, Lei Fang, Xiabin Chen, Zhenyu Jin, Hsin Hsiung Tai, Fang Zheng, Chang Guo Zhan

Research output: Contribution to journalArticlepeer-review

30 Scopus citations


Cocaine is a widely abused drug without an FDA (Food and Drug Administration)-approved medication. It has been recognized that an ideal anti-cocaine medication would accelerate cocaine metabolism producing biologically inactive metabolites via a route similar to the primary cocaine-metabolizing pathway, i.e. human BChE (butyrylcholinesterase)-catalysed hydrolysis. However, the native human BChE has a low catalytic activity against cocaine. We recently designed and discovered a BChE mutant (A199S/F227A/S287G/ A328W/Y332G) with a high catalytic activity (kcat = 5700 min -1, Km = 3.1 μM) specifically for cocaine, and the mutant was proven effective in protecting mice from acute cocaine toxicity of a lethal dose of cocaine (180 mg/kg of body weight, LD100). Further characterization in animalmodels requires establishment of a high-efficiency stable cell line for the BChE mutant production at a relatively larger scale. It has been extremely challenging to develop a high-efficiency stable cell line expressing BChE or its mutant. In the present study, we successfully developed a stable cell line efficiently expressing the BChE mutant by using a lentivirus-based repeated-transduction method. The scaled-up protein production enabled us to determine for the first time the in vivo catalytic activity and the biological half-life of this high-activity mutant of human BChE in accelerating cocaine clearance. In particular, it has been demonstrated that the BChE mutant (administered to mice 1 min prior to cocaine) can quickly metabolize cocaine and completely eliminate cocaine-induced hyperactivity in rodents, implying that the BChE mutant may be developed as a promising therapeutic agent for cocaine abuse treatment.

Original languageEnglish
Pages (from-to)447-454
Number of pages8
JournalBiochemical Journal
Issue number3
StatePublished - Aug 1 2013


  • Butyrylcholinesterase
  • Drug abuse
  • Protein drug
  • Protein production
  • Stable cell line

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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