Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.1.8 (cholinesterase)
12,691 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human and rabbit paraoxonases/arylesterases were purified to homogeneity by chromatographic and gel electrophoretic/isofocusing procedures coupled with activity stains. N-terminal and peptide sequence analysis suggested retention of the secretion signal sequence and allowed design of oligonucleotide probes. The probes were used to isolate a 1294-bp rabbit paraoxonase cDNA clone, which, in turn, was used to isolate three human cDNA clones. Comparison of rabbit and human protein and cDNA sequences indicated a high degree of sequence conservation (approximately 85% identity) and verified that paraoxonase retains its signal sequence (except for the N-terminal Met). The rabbit cDNA encodes a protein of 359 amino acids and the human a protein of 355 amino acids. In situ hybridization demonstrated, as expected, that the paraoxonase gene maps to the long arm of human chromosome 7. Arginine at position 192 specifies high activity paraoxonase and glutamine low activity human paraoxonase. Variation in protein levels explains the variation of enzyme activity observed within a genetic class. Toxicity studies showed that raising rat plasma paraoxonase levels by i.v. administration of partially purified rabbit paraoxonase protected animals against cholinesterase inhibition by paraoxon and chlorpyrifos oxon. Protection correlated with the relative rates of hydrolysis of these two compounds.
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PMID:Human and rabbit paraoxonases: purification, cloning, sequencing, mapping and role of polymorphism in organophosphate detoxification. 839 45

We characterized the interaction of the prodrug dipivefrin hydrochloride (DPE) with esterase activity in the rabbit cornea. The esterases which were identified included: (1) cholinesterase, (2) acetylcholinesterase, (3) a mixture containing carboxylesterase, acetylesterase and arylesterase, and (4) a non-specific esterase. DPE suppressed all of their activities as well as that of the mixture containing carboxylesterase, acetylesterase and arylesterase, and a nonspecific esterase. However, its effect on cholinesterase was larger than on any of the other activities, suggesting that DPE is a better substrate for cholinesterase than for any of the other esterases. These measurements along with those of substrate-dependent inhibition of 14C-DPE hydrolysis indicated that the DPE-esterase interaction was competitive based on changes in the apparent Km values which were extracted from Lineweaver-Burk plots of esterase activity. The substrate for cholinesterase competed with DPE most strongly among substrates. These results seem to suggest that DPE is hydrolyzed by various corneal esterases, mainly cholinesterase.
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PMID:Characterization of esterases involved in the hydrolysis of dipivefrin hydrochloride. 844 67

Many organophosphorus compounds (OPs) are potent cholinesterase inhibitors, accounting for their use as insecticides and, unfortunately, also as nerve agents. Each year there are approximately 3 million pesticide poisonings world-wide resulting in 220,00 deaths. In 1990, there were 1.36 million kg of chlorpyrifos, 4.67 million kg of diazinon and 1.23 million kg of ethyl parathion manufactured in the USA (data supplied by the USEPA). In addition to exposure risks during pesticide manufacturing, distribution and use, there are risks associated with the major international effort aimed at destroying the arsenals of nerve agents, including soman and sarin. The United States has pledged to destroy approximately 25,000 tons of chemical agents by the end of the decade. The high density lipoprotein (HDL)-associated enzyme paraoxonase (PON1) contributes significantly to the detoxication of several OPs (Fig. 1). The insecticides parathion, chlorpyrifos and diazinon are bioactivated to potent cholinesterase inhibitors by cytochrome P-450 systems. The resulting toxic oxon forms can be hydrolysed by PON1, which also hydrolyses the nerve agents soman and sarin (Fig. 1). PON1 is polymorphic in human populations and different individuals also express widely different levels of this enzyme. The Arg192 (R192) PON1 isoform hydrolyses paraoxon rapidly, while the Gln192 (Q191) isoform hydrolyses paraoxon slowly. Both isoforms hydrolyse chlorpyrifos-oxon and phenylacetate at approximately the same rate. The role of PON1 in OP detoxication is physiologically significant. Injected PON1 protects against OP poisoning in rodent model systems and interspecies differences in PON1 activity correlate well with observed median lethal dose (LD50) values. We report here a simple enzyme analysis that provides a clear resolution of PON1 genotypes and phenotypes allowing for a reasonable assessment of an individual's probable susceptibility or resistance to a given OP, extending earlier studies on this system. We also show that the effect of the PON1 polymorphism is reversed for the hydrolysis of diazoxon, soman and especially sarin, thus changing the view of which PON1 isoform is considered to be protective.
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PMID:The effect of the human serum paraoxonase polymorphism is reversed with diazoxon, soman and sarin. 889 66

The purpose of these experiments was to determine the reversibility of alpha-chaconine and alpha-solanine inhibition of human plasma butyrylcholinesterase (BuChE). For the substrate alpha-naphthylacetate, optimal assay conditions were 0.50 M sodium phosphate buffer and a substrate concentration of 3-5 x 10(-4) M. Dibucaine (1 x 10(-5) M) indicated the usual phenotype for all subjects; alpha-chaconine and alpha-solanine at 2.88 x 10(-6) M inhibited BuChE about 70 and 50%, respectively. One- and 24-hr incubations at 1 x 10(-5) M with alpha-chaconine, alpha-solanine, paraoxon, eserine, and ethanol yielded reversible inhibition with dilution except for paraoxon. Twenty-four-hour dialyses of incubations showed no inhibition except for paraoxon. PAGE enzyme activity gels of 1- and 24-hr incubations also showed no inhibition except for paraoxon. alpha-Chaconine and alpha-solanine are reversible inhibitors of human butyrylcholinesterase. At estimated tissue levels, alpha-chaconine, alpha-solanine, and solanidine inhibited BuChE 10-86%. In assays which combined alpha-chaconine, alpha-solanine, and solanidine, inhibition of BuChE was less than additive. No inhibition of albumin alpha-naphthylacetate esterase (an arylesterase) was noted with any inhibitor. The importance of these data to adverse toxicological effects of potato alkaloids is discussed.
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PMID:Inhibition of human plasma and serum butyrylcholinesterase (EC 3.1.1.8) by alpha-chaconine and alpha-solanine. 892 46

The extensive international use of organophosphorus compounds (OP) results in numerous acute intoxications each year. OPs inhibit acetylcholinesterase, the enzyme responsible for breaking down the neurotransmitter acetylcholine. The World Health Organization recognizes cholinesterase (ChE) biomonitoring as a preventive measure against OP overexposure. The aim of this study was to determine if dermal OP contamination could interfere with current field ChE biomonitoring assays, which use a fingerstick blood sample. In this study we also sought to determine if high levels of a plasma enzyme, A-esterase, could protect ChE from inhibition by hydrolyzing environmentally generated oxons potentially present in a fingerstick sample. A heparinized venous blood sample was collected from a volunteer. Erythrocyte acetylcholinesterase (AChE) and plasma butyrylcholinesterase (PChE) activities were measured using a field-based colorimetric cholinesterase kit. ChE dose-response curves were constructed by allowing 10-microliters blood samples to contact environmentally realistic levels of OP thioate and oxon for 10 s. An inhibition threshold could not be established for PChE when exposed to oxon within the time necessary to perform a fingerstick analysis. AChE was also inhibited by trace amounts of oxon consistent with previously reported environmental levels. These findings suggest that the reliability of field-based biomonitoring results is limited if OP residues remain on a skin surface at the time of sample collection. A-esterase's role in protecting ChE activity was investigated using capillary and venous blood from 30 unexposed individuals. Baseline ChE activities were measured, as were individual A-esterase activities using paraoxon, diazoxon, and phenylacetate as substrates. Results were then compared to ChE activities measured after 10 s of contact with an environmentally realistic amount of OP, containing 1% oxon. Both ChE activities were significantly inhibited, with capillary values being significantly more inhibited than their venous counterparts. However, no protective effect could be associated between the degree of A-esterase activity and the subsequent level of ChE inhibition observed in an individual's blood. These results suggest that (1) if there is any uncertainty about OP skin contamination, venous blood would be a more appropriate specimen to employ when using field ChE biomonitoring kits--it is collected in larger volumes and has essentially no direct contact to dermal surfaces; and (2) A-esterase activity demonstrates no protective effect against ChE inhibition upon a blood droplet's brief contact with an OP residue containing traces of oxon.
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PMID:Simulated dermal contamination with capillary samples and field cholinesterase biomonitoring. 916 60

Chlorpyrifos (CPF), a commonly used cholinesterase-inhibiting insecticide, is lethal at much lower doses to young animals than adults. To explain this higher sensitivity in younger animals, we hypothesized that young rats have less chlorpyrifos-oxonase (CPFOase) activity than adults. To test this hypothesis, CPFOase activity was measured in the brain, plasma, and liver of male, postnatal day 4 (PND4) and adult (PND90) Long-Evans rats. CPFOase is biochemically defined as a Ca(2+)-dependent A-esterase that hydrolyzes chlorpyrifos-oxon (CPFO), the active metabolite of CPE. No brain CPFOase activity was detected at either age. Plasma and liver CPFOase activities were markedly lower at PND4 compared to adult: PND4 plasma and liver CPFOase activities were 1/11 and 1/2 the adult plasma and liver activities, respectively. Because the Km of CPFOase activity was high (i.e., 210-380 microM), it was important to determine if this CPFOase activity could hydrolyze physiologically relevant concentrations (i.e., nM to low microM) of CPFO. This was accomplished by comparing the shifts in the tissue acetylcholinesterase (AChE) IC50 for CPFO in the presence or absence of CPFOase activity. One would expect an increase in the "apparent" IC50 if CPFOase hydrolyzes substantial amounts of CPFO during the 30 minutes the tissue is preincubated with the CPFO. In the adult, both plasma and liver AChE apparent IC50 values were higher in the presence of CPFOase activity, suggesting that the CPFOase in those tissues was capable of hydrolyzing physiologically relevant concentrations of CPFO within 30 minutes. In young animals, however, there was less of a shift in the IC50 curves compared to the adult, confirming that the young animal has less capacity than the adult to detoxify physiologically relevant concentrations of CPFO via CPFOase.
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PMID:Maturational differences in chlorpyrifos-oxonase activity may contribute to age-related sensitivity to chlorpyrifos. 926 78

Tazarotene is a novel acetylenic retinoid for the treatment of psoriasis and acne. We examined (1) the hydrolysis of tazarotene in blood from Japanese-American and Caucasian subjects, (2) the esterases responsible for this hydrolysis in human blood, and (3) tazarotene hydrolysis in rat and human liver microsomes. Tazarotene hydrolysis and enzyme inhibition were assessed by monitoring the disappearance of tazarotene and the appearance of its primary metabolite tazarotenic acid by HPLC. In blood, tazarotene was converted mainly to tazarotenic acid via first-order kinetics, and there was no statistically significant difference in the hydrolytic (metabolic) rate of tazarotene in uninhibited Japanese-American and Caucasian blood. Physostigmine (a cholinesterase inhibitor), bis(p-nitrophenyl) phosphate (a carboxylesterase inhibitor), and EDTA (an aromatic esterase inhibitor) did not significantly affect tazarotene hydrolysis in blood. Paraoxon, an inhibitor of all serine esterases including cholinesterase and carboxylesterase, decreased the hydrolysis of tazarotene to tazarotenic acid by 95% in both blood and liver microsomes. In conclusion, blood and liver esterases play a significant role in the hydrolysis of tazarotene to tazarotenic acid, and paraoxon-inhibitable forms of esterases are involved in this hydrolysis in humans.
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PMID:Metabolic deesterification of tazarotene in human blood and rat and human liver microsomes. 926 78

1. Paraoxon concentration was estimated by means of inhibition kinetics observed with electric eel acetylcholinesterase (AChE) which was determined by a modified Ellman procedure. In human plasma, paraoxon was stabilized by inactivation of paraoxonase with EDTA and aluminon and by inhibition of butyrylcholinesterase with ethopropazine. Paraoxon (1-50 ng) was recovered at 86+/-1.7% (mean+/-s.e.m.) in ether extracts from 0.5 ml samples of spiked stabilized plasma. It could be stored without loss at - 20 degrees C for at least 1 month. 2. The enzyme-based assay was applied to follow the paraoxon plasma concentrations in three suicidal patients with severe parathion poisoning. In poisoning with excessive doses and initial paraoxon concentrations above 500 nM, therapeutic obidoxime concentrations of approximately 10 microM failed to essentially reactivate erythrocyte AChE in vivo, while reactivatability ex vivo was nearly complete. With the plasma concentrations of paraoxon dropping below 100 nM, however, reactivation by obidoxime became significant. Unexpectedly, paraoxon levels occasionally reincreased during treatment and resulted in re-inhibition of AChE, bearing some resemblance to the Intermediate Syndrome. 3. The paraoxon concentrations measured fitted satisfactorily the values calculated from the kinetic constants previously obtained for AChE inhibition and obidoxime-induced reactivation in vitro. This indicates that diethylphosphoryloxime formation during obidoxime-induced reactivation does not markedly contribute to the re-inhibition of AChE as observed in vitro.
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PMID:Enzyme-based assay for quantification of paraoxon in blood of parathion poisoned patients. 998 68

Young rats are more sensitive than adults to a single oral dose of chlorpyrifos, an organophosphorus pesticide. A direct comparison of chlorpyrifos effects in young (postnatal day 17; PND17), adolescent (PND27), and adult (70 days) Long-Evans rats was conducted to determine quantitative and possibly qualitative differences in sensitivity in terms of behavioral changes and cholinesterase (ChE; total cholinesterase activity) inhibition at these three ages. Male and female rats were administered chlorpyrifos orally at one of two doses (PND17, 5 or 20 mg/kg; PND27, 20 or 50 mg/kg; adult, 20 or 80 mg/kg) and tested at either 3.5 or 6.5 h after dosing. Behavioral testing included observational evaluations and measurements of motor activity and was followed immediately by tissue collection for ChE determination in brain and blood. For both behavioral changes and ChE inhibition, peak effects occurred at 3.5 h in adult male and PND27 rats (both sexes) and at 6.5 h in adult female and PND17 rats (both sexes). Comparisons of the 20 mg/kg dose across ages showed generally less ChE inhibition and fewer behavioral effects with increasing age, except that the adult females were similar to the PND27 rats. The high dose used for each age group produced similar brain ChE inhibition (80-90%) and generally similar behavioral effects. Interestingly, a few end-points in the young rats were less affected than in adults at this level of ChE inhibition. The degree of ChE inhibition in the brain more closely paralleled the blood inhibition in the younger rats, compared to the adults. Carboxylesterase (CaE) and A-esterase are known to play an important role in the detoxification of organophosphates and may be partially responsible for these sensitivity differences. Liver and plasma CaE and A-esterase activities were measured in untreated male rats on PND1, 4, 7, 12, 17, and 21 and in adults of both sexes (82-92 days old). Preweanling rats had considerably less activity of both enzymes, and adult females had less liver CaE activity than males. These differences in detoxifying enzymes correlate with the age-related differences in behavioral and biochemical effects, as well as the gender differences seen in adult rats, and thus may be a major influence on the differential sensitivity to chlorpyrifos.
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PMID:Age- and gender-related differences in sensitivity to chlorpyrifos in the rat reflect developmental profiles of esterase activities. 1004 24

Previously Haley et al. described six possible syndromes identified by factor analysis of symptoms in Gulf War veterans and demonstrated that veterans with these symptom complexes were more neurologically impaired than age-sex-education-matched well controls. They also uncovered strong associations (relative risks 4-8) suggesting that these symptom complexes were related to wartime exposure to combinations of organophosphate pesticides, chemical nerve agents, high concentration DEET insect repellant, and symptoms of advanced acute toxicity after taking pyridostigmine. Here we have shown that compared to controls, ill veterans with the neurologic symptom complexes were more likely to have the R allele (heterozygous QR or homozygous R) than to be homozygous Q for the paraoxonase/arylesterase 1 (PON1) gene. Moreover, low activity of the PON1 type Q (Gln192, formerly designated type A) arylesterase allozyme distinguished ill veterans from controls better than just the PON1 genotype or the activity levels of the type R (Arg192, formerly designated type B) arylesterase allozyme, total arylesterase, total paraoxonase, or butyrylcholinesterase. A history of advanced acute toxicity after taking pyridostigmine was also correlated with low PON1 type Q arylesterase activity. Type Q is the allozyme of paraoxonase/arylesterase that most efficiently hydrolyzes several organophosphates including sarin, soman, and diazinon. These findings further support the proposal that neurologic symptoms in some Gulf War veterans were caused by environmental chemical exposures.
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PMID:Association of low PON1 type Q (type A) arylesterase activity with neurologic symptom complexes in Gulf War veterans. 1037 7


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