Rational design, preparation, and characterization of a therapeutic enzyme mutant with improved stability and function for cocaine detoxification.

ABSTRACT: Cocaine esterase (CocE) is known as the most efficient natural enzyme for cocaine hydrolysis. The major obstacle to the clinical application of wild-type CocE is the thermoinstability with a half-life of only ∼12 min at 37 °C. The previously designed T172R/G173Q mutant (denoted as enzyme E172-173) with an improved in vitro half-life of ∼6 h at 37 °C is currently in clinical trial Phase II for cocaine overdose treatment. Through molecular modeling and dynamics simulation, we designed and characterized a promising new mutant of E172-173 with extra L196C/I301C mutations (denoted as enzyme E196-301) to produce cross-subunit disulfide bonds that stabilize the dimer structure. The cross-subunit disulfide bonds were confirmed by X-ray diffraction. The designed L196C/I301C mutations have not only

SUBMITTER: Fang L 

PROVIDER: S-EPMC4136690 | biostudies-literature | 2014 Aug

REPOSITORIES: biostudies-literature