Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.1.7 (acetylcholinesterase)
 28,390 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A nonspecific phospholipid exchange protein (PLEP) preparation was used to transfer dansyl [3H]phosphatidylethanolamine (DNS-PE), dansyl[3H]phosphatidylserine (DNS-PS), and dehydroergosterol (DHE) from sonicated lipid vesicles to electroplax plasma membrane fragments enriched in Na+,K+-ATPase with retention of 80-90% of Na+,K+-ATPase activity. The transfer of individual fluorescent lipid molecules was distinguished from a nonspecific association of lipid vesicles and membranes by including [14C]triolein, a lipid that is not transferred by PLEPs, in the vesicles. Dansyl 3H-labeled phospholipids (DNS-[3H]PLs) or DHE was considered "incorporated" into the Na+,K+-ATPase membranes when fluorophores pelleted with the Na+,K+-ATPase preparation without the nonexchangeable [14C]triolein. The locations of incorporated DHE and DNS-PLs were also described by iodide quenching experiments. DHE was not accessible to iodide for quenching, while 75% of the DNS-PLs incorporated into Na+,K+-ATPase membrane fragments were accessible to iodide. After a technique was developed for using PLEP to incorporate fluorescent lipids into membranes with the Na+,K+-ATPase preparation, DNS-PE, DNS-PS, and DHE were then analogously incorporated into electroplax plasma membranes enriched in acetylcholinesterase (AChE) and into erythrocyte ghosts in order to evaluate the fluorophores as membrane probes. In the subsequent evaluation, the fluorescent properties of membrane-incorporated DNS-PE, DNS-PS, and DHE were systematically compared to the fluorescent properties of the molecules in lipid vesicles. The fluorescence polarizations of both DNS-PLs were increased by the presence of protein in a bilayer. The fluorescence polarization of DNS-PS was greater than the polarization of DNS-PE in both membranes and vesicles. In contrast, the polarization (and the lifetime) of DHE was the same whether the fluorescent sterol was in a membrane preparation or in vesicles. Fluorescence polarization and intensity of all three fluorophores were measured in the bilayer preparations as a function of temperature. The intensities of all three probes and the polarization of DNS-PE in both membranes and vesicles decreased biphasically with a change in slope occurring at 26.0-27.5 degrees C. DNS-PS in lipid vesicles was depolarized biphasically with increasing temperature, but when incorporated into membranes, DNS-PS was depolarized linearly without a change in slope. The polarization of DHE in either membranous or vesicle bilayers did not change with temperature.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Incorporation of danyslated phospholipids and dehydroergosterol into membranes using a phospholipid exchange protein. 631 6

Carbofuran is a carbamate pesticide used in agricultural practice throughout the world. Its effect as a pesticide is due to its ability to inhibit acetylcholinesterase activity. Though carbofuran has a long history of use, there is little information available with respect to its metabolic fate and disposition in mammals. The present study was designed to investigate the comparative in vitro metabolism of carbofuran from human, rat, and mouse liver microsomes (HLM, RLM, MLM, respectively), and characterize the specific enzymes involved in such metabolism, with particular reference to human metabolism. Carbofuran is metabolized by cytochrome P450 (CYP) leading to the production of one major ring oxidation metabolite, 3-hydroxycarbofuran, and two minor metabolites. The affinity of carbofuran for CYP enzymes involved in the oxidation to 3-hydroxycarbofuran is significantly less in HLM (Km=1.950 mM) than in RLM (Km=0.210 mM), or MLM (Km=0.550 mM). Intrinsic clearance rate calculations indicate that HLM are 14-fold less efficient in the metabolism of carbofuran to 3-hydroxycarbofuran than RLM or MLM. A screen of 15 major human CYP isoforms for metabolic ability with respect to carbofuran metabolism demonstrated that CYP3A4 is the major isoform responsible for carbofuran oxidation in humans. CYP1A2 and 2C19 are much less active while other human CYP isoforms have minimal or no activity toward carbofuran. In contrast with the human isoforms, members of the CYP2C family in rats are likely to have a primary role in carbofuran metabolism. Normalization of HLM data with the average levels of each CYP in native HLM, indicates that carbofuran metabolism is primarily mediated by CYP3A4 (percent total normalized rate (% TNR)=77.5), although CYP1A2 and 2C19 play ancillary roles (% TNR=9.0 and 6.0, respectively). This is substantiated by the fact that ketoconazole, a specific inhibitor of CYP3A4, is an excellent inhibitor of 3-hydroxycarbofuran formation in HLM (IC50: 0.31 microM). Chlorpyrifos, an irreversible non-competitive inhibitor of CYP3A4, inhibits the formation of 3-hydroxycarbofuran in HLM (IC50: 39 microM). The use of phenotyped HLM demonstrated that individuals with high levels of CYP3A4 have the greatest potential to metabolize carbofuran to its major metabolite. The variation in carbofuran metabolism among 17 single-donor HLM samples is over 5-fold and the best correlation between CYP isoform activity and carbofuran metabolism was observed with CYP3A4 (r2=0.96). The interaction of carbofuran and the endogenous CYP3A4 substrates, testosterone and estradiol, were also investigated. Testosterone metabolism was activated by carbofuran in HLM and CYP3A4, however, less activation was observed for carbofuran metabolism by testosterone in HLM and CYP3A4. No interactions between carbofuran and estradiol metabolism were observed.
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PMID:In vitro metabolism of carbofuran by human, mouse, and rat cytochrome P450 and interactions with chlorpyrifos, testosterone, and estradiol. 1556 Aug 89