Diacetylmorphine is a narcotic analgesic that may be habit-forming. It is a controlled substance (opium derivative). Diacetyl derivative of the opiate morphine, a naturally occurring alkaloid extracted from the seedpod of the Asian poppy (Papaver sp.). Once administered, diamorphine (or diacetylmorphine) is rapidly hydrolyzed to 6-monoacetylmorphine (6-MAM) and then to the end-product morphine which binds to opiate receptors located throughout the mammalian nervous and gastrointestinal systems
2 moreTitle: Fundamental reaction pathway and free energy profile for butyrylcholinesterase-catalyzed hydrolysis of heroin Qiao Y, Han K, Zhan CG Ref: Biochemistry, 52:6467, 2013 : PubMed
The pharmacological function of heroin requires an activation process that transforms heroin into 6-monoacetylmorphine (6-MAM), which is the most active form. The primary enzyme responsible for this activation process in human plasma is butyrylcholinesterase (BChE). The detailed reaction pathway of the activation process via BChE-catalyzed hydrolysis has been explored computationally, for the first time, in this study via molecular dynamics simulation and first-principles quantum mechanical/molecular mechanical free energy calculations. It has been demonstrated that the whole reaction process includes acylation and deacylation stages. The acylation consists of two reaction steps, i.e., the nucleophilic attack on the carbonyl carbon of the 3-acetyl group of heroin by the hydroxyl oxygen of the Ser198 side chain and the dissociation of 6-MAM. The deacylation also consists of two reaction steps, i.e., the nucleophilic attack on the carbonyl carbon of the acyl-enzyme intermediate by a water molecule and the dissociation of the acetic acid from Ser198. The calculated free energy profile reveals that the second transition state (TS2) should be rate-determining. The structural analysis reveals that the oxyanion hole of BChE plays an important role in the stabilization of rate-determining TS2. The free energy barrier (15.9 +/- 0.2 or 16.1 +/- 0.2 kcal/mol) calculated for the rate-determining step is in good agreement with the experimentally derived activation free energy ( approximately 16.2 kcal/mol), suggesting that the mechanistic insights obtained from this computational study are reliable. The obtained structural and mechanistic insights could be valuable for use in the future rational design of a novel therapeutic treatment of heroin abuse.
Selective inhibition of peripheral esterases by tri-ortho-tolyl phosphate in the mouse resulted in an increase in the analgetic activity of heroin, without affecting the activity of morphine. In vitro inhibition of esterases by paraoxon reduced the affinity of heroin for the opiate receptor, while that of morphine was unaffected. These results suggest that both central and peripheral esterases are involved in the metabolism of heroin and that interference with critical esterases can alter its pharmacologic and toxicologic effects.
The enzyme in human serum that rapidly hydrolyzes diacetylmorphine (heroin) to 6-acetylmorphine is identified in this report as serum cholinesterase (EC 3.1.1.8, acylcholine acylhydrolase; also called pseudocholinesterase or butyrylcholine esterase). The rate of heroin hydrolysis was measured spectrophotometrically at 245 nm using highly purified serum cholinesterase. The turnover number was 500 mumol of heroin hydrolyzed per min per mumol active site. The product was identified spectrophotometrically and by thin-layer chromatography to be 6-acetylmorphine. There appeared to be marked product inhibition of heroin hydrolysis, as 6-acetylmorphine (Ki = 0.015 mM) bound 7 times more tightly than heroin (Ki = 0.11 mM). Purified human serum arylesterase did not hydrolyze heroin. Purified serum cholinesterase accounted for all the observed heroin hydrolysis by whole serum. The genetic variants of human serum cholinesterase, silent and atypical cholinesterase, were also tested. Serum from a person identified as having silent cholinesterase did not hydrolyze heroin. Purified atypical cholinestearase hydrolyzed heroin, but the binding was less tight (Km = 0.45 mM) than with usual cholinesterase (Km = 0.11 mM). The possibility that heroin potency may be influenced by serum cholinesterase genotype or activity level remains to be investigated.
2 lessTitle: Fundamental reaction pathway and free energy profile for butyrylcholinesterase-catalyzed hydrolysis of heroin Qiao Y, Han K, Zhan CG Ref: Biochemistry, 52:6467, 2013 : PubMed
The pharmacological function of heroin requires an activation process that transforms heroin into 6-monoacetylmorphine (6-MAM), which is the most active form. The primary enzyme responsible for this activation process in human plasma is butyrylcholinesterase (BChE). The detailed reaction pathway of the activation process via BChE-catalyzed hydrolysis has been explored computationally, for the first time, in this study via molecular dynamics simulation and first-principles quantum mechanical/molecular mechanical free energy calculations. It has been demonstrated that the whole reaction process includes acylation and deacylation stages. The acylation consists of two reaction steps, i.e., the nucleophilic attack on the carbonyl carbon of the 3-acetyl group of heroin by the hydroxyl oxygen of the Ser198 side chain and the dissociation of 6-MAM. The deacylation also consists of two reaction steps, i.e., the nucleophilic attack on the carbonyl carbon of the acyl-enzyme intermediate by a water molecule and the dissociation of the acetic acid from Ser198. The calculated free energy profile reveals that the second transition state (TS2) should be rate-determining. The structural analysis reveals that the oxyanion hole of BChE plays an important role in the stabilization of rate-determining TS2. The free energy barrier (15.9 +/- 0.2 or 16.1 +/- 0.2 kcal/mol) calculated for the rate-determining step is in good agreement with the experimentally derived activation free energy ( approximately 16.2 kcal/mol), suggesting that the mechanistic insights obtained from this computational study are reliable. The obtained structural and mechanistic insights could be valuable for use in the future rational design of a novel therapeutic treatment of heroin abuse.
BACKGROUND AND PURPOSE Carboxylesterases (CEs) metabolize a wide range of xenobiotic substrates including heroin, cocaine, meperidine and the anticancer agent CPT-11. In this study, we have purified to homogeneity human liver and intestinal CEs and compared their ability with hydrolyse heroin, cocaine and CPT-11.
EXPERIMENTAL APPROACH:
The hydrolysis of heroin and cocaine by recombinant human CEs was evaluated and the kinetic parameters determined. In addition, microsomal samples prepared from these tissues were subjected to chromatographic separation, and substrate hydrolysis and amounts of different CEs were determined.
KEY RESULTS:
In contrast to previous reports, cocaine was not hydrolysed by the human liver CE, hCE1 (CES1), either as highly active recombinant protein or as CEs isolated from human liver or intestinal extracts. These results correlated well with computer-assisted molecular modelling studies that suggested that hydrolysis of cocaine by hCE1 (CES1), would be unlikely to occur. However, cocaine, heroin and CPT-11 were all substrates for the intestinal CE, hiCE (CES2), as determined using both the recombinant protein and the tissue fractions. Again, these data were in agreement with the modelling results.
CONCLUSIONS AND IMPLICATIONS:
These results indicate that the human liver CE is unlikely to play a role in the metabolism of cocaine and that hydrolysis of this substrate by this class of enzymes is via the human intestinal protein hiCE (CES2). In addition, because no enzyme inhibition is observed at high cocaine concentrations, potentially this route of hydrolysis is important in individuals who overdose on this agent.
Title: Genetic variants of human serum cholinesterase influence metabolism of the muscle relaxant succinylcholine. Lockridge O Ref: Pharmacol Ther, 47:35, 1990 : PubMed
People with genetic variants of cholinesterase respond abnormally to succinylcholine, experiencing substantial prolongation of muscle paralysis with apnea rather than the usual 2-6 min. The structure of usual cholinesterase has been determined including the complete amino acid and nucleotide sequence. This has allowed identification of altered amino acids and nucleotides. The variant most frequently found in patients who respond abnormally to succinylcholine is atypical cholinesterase, which occurs in homozygous form in 1 out of 3500 Caucasians. Atypical cholinesterase has a single substitution at nucleotide 209 which changes aspartic acid 70 to glycine. This suggests that Asp 70 is part of the anionic site, and that the absence of this negatively charged amino acid explains the reduced affinity of atypical cholinesterase for positively charged substrates and inhibitors. The clinical consequence of reduced affinity for succinylcholine is that none of the succinylcholine is hydrolyzed in blood and a large overdose reaches the nerve-muscle junction where it causes prolonged muscle paralysis. Silent cholinesterase has a frame shift mutation at glycine 117 which prematurely terminates protein synthesis and yields no active enzyme. The K variant, named in honor of W. Kalow, has threonine in place of alanine 539. The K variant is associated with 33% lower activity. All variants arise from a single locus as there is only one gene for human cholinesterase (EC 3.1.1.8). Comparison of amino acid sequences of esterases and proteases shows that cholinesterase belongs to a new family of serine esterases which is different from the serine proteases.
Selective inhibition of peripheral esterases by tri-ortho-tolyl phosphate in the mouse resulted in an increase in the analgetic activity of heroin, without affecting the activity of morphine. In vitro inhibition of esterases by paraoxon reduced the affinity of heroin for the opiate receptor, while that of morphine was unaffected. These results suggest that both central and peripheral esterases are involved in the metabolism of heroin and that interference with critical esterases can alter its pharmacologic and toxicologic effects.
The enzyme in human serum that rapidly hydrolyzes diacetylmorphine (heroin) to 6-acetylmorphine is identified in this report as serum cholinesterase (EC 3.1.1.8, acylcholine acylhydrolase; also called pseudocholinesterase or butyrylcholine esterase). The rate of heroin hydrolysis was measured spectrophotometrically at 245 nm using highly purified serum cholinesterase. The turnover number was 500 mumol of heroin hydrolyzed per min per mumol active site. The product was identified spectrophotometrically and by thin-layer chromatography to be 6-acetylmorphine. There appeared to be marked product inhibition of heroin hydrolysis, as 6-acetylmorphine (Ki = 0.015 mM) bound 7 times more tightly than heroin (Ki = 0.11 mM). Purified human serum arylesterase did not hydrolyze heroin. Purified serum cholinesterase accounted for all the observed heroin hydrolysis by whole serum. The genetic variants of human serum cholinesterase, silent and atypical cholinesterase, were also tested. Serum from a person identified as having silent cholinesterase did not hydrolyze heroin. Purified atypical cholinestearase hydrolyzed heroin, but the binding was less tight (Km = 0.45 mM) than with usual cholinesterase (Km = 0.11 mM). The possibility that heroin potency may be influenced by serum cholinesterase genotype or activity level remains to be investigated.
human-BCHE
