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 (k cat ϭ 4 min Ϫ1 ) 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 (k cat ϭ 154 min Ϫ1

Mechanically strong yet rehealable and recyclable thermosets were achieved through hybridization of rigid imide moieties and dynamic imine linkages.

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Cage-templated synthesis of narrowly distributed palladium nanoparticles (1.8 ± 0.2 nm) and their high catalytic activity in Suzuki–Miyaura coupling reactions are reported.

Butyrylcholinesterase (BChE) is important in cocaine metabolism, but it hydrolyzes (؊)-cocaine only one-two thousandth as fast as the unnatural (؉)-stereoisomer. A starting point in engineering BChE mutants that rapidly clear cocaine from the bloodstream, for overdose treatment, is to elucidate structural factors underlying the stereochemical difference in catalysis. Here, we report two three-dimensional Michaelis-Menten complexes of BChE liganded with natural and unnatural cocaine molecules, respectively, that were derived from molecular modeling and supported by experimental studies. Such complexes revealed that the benzoic ester group of both cocaine stereoisomers must rotate toward the catalytic Ser 198 for hydrolysis. Rotation of (؊)-cocaine appears to be hindered by interactions of its phenyl ring with Phe 329 and Trp 430 . These interactions do not occur with (؉)-cocaine. Because the rate of (؊)-cocaine hydrolysis is predicted to be determined mainly by the re-orientation step, it should not be greatly influenced by pH. In fact, measured rates of this reaction were nearly constant over the pH range from 5.5 to 8.5, despite large rate changes in hydrolysis of (؉)-cocaine. Our models can explain why BChE hydrolyzes (؉)-cocaine faster than (؊)-cocaine, and they suggest that mutations of certain residues in the catalytic site could greatly improve catalytic efficiency and the potential for detoxication.Cocaine overdose is a leading cause of death among urbandwelling young adults (1, 2). A new idea for treating cocaine overdose is to accelerate metabolic clearance by administering an appropriate hydrolase. Plasma butyrylcholinesterase (BChE) 1 can hydrolyze cocaine to ecgonine methyl ester and reportedly accounts for all the blood-borne cocaine hydrolysis activity in humans (3-5). Pretreatment with human BChE provides rats with substantial protection against cocaine-induced cardiac arrhythmia, hypertension, and locomotor hyperactivity (6). Exogenous BChE is also able to rescue rats given an overdose of cocaine (7). However, the k cat of BChE for (Ϫ)-cocaine is only 3.9 min Ϫ1, versus 33,900 min Ϫ1 for the optimal substrate, butyrylthiocholine. We estimate that timely detoxication of a cocaine overdose, with serum levels up to 20 mg/l (8), would require at least 100 mg of enzyme. An A328Y mutant of BChE developed by the trial and error approach (9) showed 4 -5-fold improvement in (Ϫ)-cocaine hydrolysis, but it is still not satisfactory for treatment of cocaine overdose. Because human BChE hydrolyzes synthetic (ϩ)-cocaine (see Fig. 1) about 2,000-fold faster than the natural (Ϫ)-cocaine (see Fig. 1) (9), elucidation of the structural factors responsible for the different catalytic rates may offer a rational strategy for engineering BChE mutants that can hydrolyze (Ϫ)-cocaine rapidly enough to treat cocaine overdose.Theoretical 3D structures of BChE (10 -12), of its complex with cocaine (9), and of the cocaine molecule itself have all been reported (13-16). Nevertheless, the literature provides no explanation o...

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Plasma butyrylcholinesterase (BChE) is important in the metabolism of cocaine, but natural human BChE has limited therapeutic potential for detoxication because of low catalytic efficiency with cocaine. Here we report pharmacokinetics of cocaine in rats treated with A328W/Y332A BChE, an excellent cocaine hydrolase designed with the aid of molecular modeling. Compared with wild-type BChE, this enzyme hydrolyzes cocaine with 40-fold improved k cat (154 min Ϫ1 versus 4.1 min Ϫ1 ) and only slightly increased K M (18 M versus 4.5 M). In rats given this hydrolase (3 mg/kg i.v.) 10 min before cocaine challenge (6.8 mg/kg i.v.), cocaine half-life was reduced from 52 min to 18 min. Mirroring the reductions of plasma cocaine were large increases in benzoic acid, a product of BChE-mediated cocaine hydrolysis. All other pharmacokinetic parameters confirmed a large, dose-dependent acceleration of cocaine removal by the injected cocaine hydrolase. These results show that A328W/Y332A, an efficient cocaine hydrolase in vivo as well as in vitro, might promote cocaine detoxication in a clinical setting.

Hierarchical structured FeCo2@APCFs catalyzed PMS, which exhibited ultrafast degradation of organic pollutants, good stability, recyclability, and high mineralization.