Re-engineering butyrylcholinesterase as a cocaine hydrolase

Hong Sun, Yuan Ping Pang, Oksana Lockridge, Stephen Brimijoin

Research output: Contribution to journalArticlepeer-review

125 Scopus citations


To address the problem of acute cocaine overdose, we undertook molecular engineering of butyrylcholinesterase (BChE) as a cocaine hydrolase so that modest doses could be used to accelerate metabolic clearance of this drug. Molecular modeling of BChE complexed with cocaine suggested that the inefficient hydrolysis (kcat = 4 min-1) involves a rotation toward the catalytic triad, hindered by Tyr332. To eliminate rotational hindrance and retain substrate affinity, we introduced two amino acid substitutions (Ala328Trp/Tyr332Ala). The resulting mutant BChE reduced cocaine burden in tissues, accelerated plasma clearance by 20-fold, and prevented cocaine-induced hyperactivity in mice. The enzyme's kinetic properties (kcat = 154 min-1, KM = 18 μM) satisfy criteria suggested previously for treating cocaine overdose (kcat > 120 min-1, KM < 30 μM). This success demonstrates that computationally guided mutagenesis can generate functionally novel enzymes with clinical potential.

Original languageEnglish (US)
Pages (from-to)220-224
Number of pages5
JournalMolecular pharmacology
Issue number2
StatePublished - 2002

ASJC Scopus subject areas

  • Molecular Medicine
  • Pharmacology


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