Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. Coding sequences for the human acetylcholinesterase (HuAChE; EC 3.1.1.7) hydrophilic subunit were subcloned in an expression plasmid vector under the control of cytomegalovirus IE gene enhancer-promoter. The human embryonic kidney cell line 293, transiently transfected with this vector, expressed catalytically active acetylcholinesterase. 2. The recombinant gene product exhibits biochemical traits similar to native "true" acetylcholinesterase as manifested by characteristic substrate inhibition, a Km of 117 microM toward acetylthiocholine, and a high sensitivity to the specific acetylcholinesterase inhibitor BW284C51. 3. The transiently transfected 293 cells (100 mm dish) produce in 24 hr active enzyme capable of hydrolyzing 1500 nmol acetylthiocholine per min. Eighty percent of the enzymatic activity appears in the cell growth medium as soluble acetylcholinesterase; most of the cell associated activity is confined to the cytosolic fraction requiring neither detergent nor high salt for its solubilization. 4. The active secreted recombinant enzyme appears in the monomeric, dimeric, and tetrameric globular hydrophilic molecular forms. 5. In conclusion, the catalytic subunit expressed from the hydrophilic AChE cDNA species has the inherent potential to be secreted in the soluble globular form and to generate polymorphism through self-association.
Cell Mol Neurobiol 1991 Feb
PMID:Recombinant human acetylcholinesterase is secreted from transiently transfected 293 cells as a soluble globular enzyme. 184 51

1. We analyzed the mode of attachment of 16 S tailed acetylcholinesterase (AChE; EC 3.1.1.7) to rat superior cervical ganglion (SCG) neuronal membranes. Using extractions by high-salt (HS) and nonionic detergent (Triton X-100), we found two pools of 16 S AChE. 2. The detergent-extracted (DE) 16 S AChE was tightly bound to membranes through detergent-sensitive, high-salt insensitive interactions and was distinct from high-salt-soluble 16 S AChE. The detergent-extracted (DE) 16 S AChE constituted a significant proportion of about one-third of the total 16 S AChE. 3. Treatment of the neuronal membranes by a phosphatidylinositol-specific phospholipase C (PIPLC) resulted in the release of some, but not all DE 16 S AChE, indicating that a significant amount of the neuronal DE 16 S AChE, about one-third, is anchored to membranes through a phosphatidylinositol containing residue. Thus, a covalent association of a glycolipid and catalytic or structural AChE polypeptidic chains occurs not only for dimeric AChE but also for the asymmetric species of AChE. 4. The complex polymorphism of AChE is due not only to different globular or asymmetric associations of catalytic and structural subunits but also to the alternative existence of a transmembrane domain or a glycolipid membrane anchor.
Cell Mol Neurobiol 1991 Feb
PMID:Phosphatidylinositol is involved in the attachment of tailed asymmetric acetylcholinesterase to neuronal membranes. 184 54

1. We describe two simple procedures for the rapid identification of certain structural features of glycolipid anchors in acetylcholinesterases (AChEs). 2. Treatment with alkaline hydroxylamine (that cleaves ester-linked acyl chains but not ether-linked alkyl chains) converts molecules possessing a diacylglycerol, but not those with an alkylacylglycerol, into hydrophilic derivatives. AChEs in human and bovine erythrocytes possess an alkylacylglycerol (Roberts et al., J. Biol. Chem. 263:18766-18775, 1988; Biochem. Biophys. Res. Commun. 150:271-277, 1988) and are not converted to hydrophilic dimers by alkaline hydroxylamine. Amphiphilic dimers of AChE from Drosophila, from mouse erythrocytes, and from the human erythroleukaemia cell line K562 also resist the treatment with hydroxylamine and likely possess a terminal alkylacylglycerol. This indicates that the cellular pool of free glycolipids used as precursors of protein anchors is distinct from the pool of membrane phosphatidylinositols (which contain diacylglycerols). 3. Pretreatment with alkaline hydroxylamine is required to render the amphiphilic AChE from human erythrocytes susceptible to digestion by Bacillus thuringiensis phosphatidylinositol-specific phospholipase C (PI-PLC) (Toutant et al., Eur. J. Biochem. 180:503-508, 1989). We show here that this is also the case for the AChE from mouse erythrocytes, which therefore likely possesses an additional acyl chain in the anchor that prevents the action of PI-PLC. 4. In two sublines of K562 cells (48 and 243), we observed that AChE either was directly susceptible to PI-PLC (243) or required a prior deacylation by alkaline hydroxylamine (48). This suggests that glycolipid anchors in AChE of K562-48 cells, but not those in AChE of K562-243 cells, contain the additional acylation demonstrated in AChE from human erythrocytes. These observations illustrate the cell specificity (and the lack of species-specificity) of the structure of glycolipid anchors.
Cell Mol Neurobiol 1991 Feb
PMID:Rapid analysis of glycolipid anchors in amphiphilic dimers of acetylcholinesterases. 184 55

1. The acetylcholinesterase (AChE) gene from the important malaria vector Anopheles stephensi has been isolated by homology to the Drosophila acetylcholinesterase gene. 2. The complete sequence and intron-exon organization has been determined. The encoded protein has 69% identity to Drosophila AChE and 38 and 36% identity to Torpedo AChE and human butyrylcholinesterase, respectively.
Cell Mol Neurobiol 1991 Feb
PMID:The acetylcholinesterase gene of Anopheles stephensi. 190 15

Previous investigations have suggested that immune-sensitization increases airway smooth muscle responsiveness to cholinomimetic stimulation by reducing the rate of degradation of acetylcholine. To examine the hypothesis that increased cholinomimetic responsiveness of tracheal smooth muscle (TSM) caused by immune-sensitization results from inhibition of acetylcholinesterase (AChase) activity, we developed a method for direct measurement of AChase activity in homogenates of TSM obtained from mongrel dogs actively sensitized in vivo to ragweed pollen extract (n = 7) and sham-sensitized littermate controls (n = 7). For both sensitized and control specimens, saturation of AChase was obtained at approximately 3.12 mM substrate (acetylthiocholine); however, maximal enzyme activity in homogenates of ragweed-sensitized tissues was significantly less (0.862 +/- 0.088 absorbance units/min/mg protein [AU/min/mg]) compared to control homogenates (1.590 +/- 0.129 AU/min/mg; P less than 0.001). Kinetic analysis (Eadie-Hofstee plot) indicated similar Michaelis constants (Km) for AChase from ragweed-sensitized (0.360 +/- 0.063) and control (0.336 +/- 0.062) homogenates (P = NS). The concentration of physostigmine eliciting half-maximal inhibition (Ki) of AChase activity also was similar for tissues from sensitized (-7.92 +/- 0.032 log M) and control animals (-7.86 +/- 0.012 log M; P = NS). Pretreatment with selected mediators of anaphylaxis (10(-4) M histamine, 10(-6) M serotonin, 10(-5) M prostaglandin E2, 10(-6) M prostaglandin F2 alpha, and 10(-7) M leukotriene D4) did not affect AChase activity. Our data demonstrate reduced AChase activity in homogenates of canine TSM after active immune-sensitization in vivo. This corresponds to functional augmentation of cholinomimetic contraction in actively sensitized tissues.
Am J Respir Cell Mol Biol 1991 Jul
PMID:Reduced activity of acetylcholinesterase in canine tracheal smooth muscle homogenates after active immune-sensitization. 190 87

Propidium has been demonstrated in previous studies to be a selective ligand for the peripheral anionic site on acetylcholinesterase (EC 3.1.1.7). Its association with this site can be advantageously monitored by direct fluorescent titration. We have measured the ability of acetylcholine, acetylthiocholine, haloxon [di-(2-chloroethyl)3-chloro-4-methylcoumarin-7-ylphosphate] , and a coumarin derivative (3-chloro-7-hydroxy-4-methylcoumarin) to dissociate propidium from the peripheral anionic site of Torpedo californica acetylcholinesterase. Measurements were made by back-titration of propidium after complete inhibition of the active center with diisopropylfluorophosphate. Both acetylcholine and acetylthiocholine show substrate inhibition at high substrate concentrations. The concentrations required for occupation of the peripheral site, as ascertained by competition with propidium, correlated well with the concentration dependence for the kinetics of substrate inhibition. These observations are consistent with substrate inhibition being due to binding of acetylcholine or acetylthiocholine at a peripheral anionic site. Displacement of propidium by haloxon and coumarin indicated that these inhibitors also bind to the peripheral anionic site. The dissociation constants ascertained from peripheral site occupation are in agreement with the constants obtained from inhibition kinetics. Evidence is presented that competition with propidium obtained by direct fluorescence titrations, when combined with inhibition kinetics, provides a more reliable means for ascertaining site selectivity of various inhibitors than does a kinetic analysis alone.
Mol Pharmacol 1991 Jan
PMID:Role of the peripheral anionic site on acetylcholinesterase: inhibition by substrates and coumarin derivatives. 198 54

1. Comparison of partial amino acid sequences of G2-acetylcholinesterase (AChE) from bovine erythrocytes and G4-AChE from bovine caudate nucleus revealed no differences in primary structure between the two enzymes. The first 33 residues of the N-terminal sequences were identical. 2. In addition, the amino acid sequences of four peptides generated by tryptic and cyanogen bromide cleavage were identical for bovine erythrocyte and brain AChE, suggesting one identical major coding exon for the adult bovine AChE forms. Comparison of these sequences with that of fetal bovine serum AChE (Doctor et al., 1988), showed differences in residues 16, 181, 212, and 216. 3. Deglycosylation studies of the two adult enzyme forms revealed that the core protein of erythrocyte AChE has an approximately 4 kDa lower molecular mass than brain AChE. This most probably reflects differences in the C-terminal sequences of the two enzymes.
Cell Mol Neurobiol 1991 Feb
PMID:Comparative studies on the primary structure of acetylcholinesterases from bovine caudate nucleus and bovine erythrocytes. 201 55

1. We have analyzed the behavior of two types of asymmetric molecular forms (A forms) of acetylcholinesterase (AChE) during development of chick hindlimb muscle, in vivo and in cell culture, and upon irreversible inactivation of peroneal muscle AChE with diisopropylfluorophosphate (DFP) in vivo. 2. In agreement with previous developmental studies on chick muscle, globular forms of AChE (G forms) are predominant in chick hindlimb at early embryonic ages, being gradually replaced by A forms as hatching (and, therefore, onset of locomotion) approaches. Of the two A-form types, AI appears and accumulates significantly earlier than AII, so that A/G and II/I ratios higher than 1 are attained only at about hatching time. 3. Cultures prepared from 11-day chick embryo hindlimb myoblasts express both types of A forms, with a combined activity of 27% of total AChE after 12 days in culture. AI forms appear again earlier and are much more abundant than type II asymmetric species through the life span of cultures. 4. All AChE activity in the peroneal muscle is irreversibly inactivated by injection of DFP in vivo. The recovery of A forms follows the same sequence described for normal development, with a delayed and slower recovery of AII forms as compared with AI. 5. Several hypotheses involving tail polypeptides or tissue target molecules, or posttranslational interconversion, are proposed to help explain the earlier appearance and accumulation of AI forms in chick muscle.
Cell Mol Neurobiol 1991 Feb
PMID:Two types of asymmetric acetylcholinesterase in chick hindlimb muscle: developmental profiles, in vivo and in cell culture, and recovery after inactivation. 201 57

1. In various brain regions, there is a puzzling disparity between large amounts of acetylcholinesterase and low levels of acetylcholine. One such area is the substantia nigra. Furthermore, within the substantia nigra, a soluble form of acetylcholinesterase is released from the dendrites of dopamine-containing nigrostriatal neurons, independent of cholinergic transmission. These two issues have prompted the hypothesis that acetylcholinesterase released in the substantia nigra has an unexpected noncholinergic function. 2. Electrophysiological studies demonstrate that this dendritic release is a function, not of the excitability of the cell from which the acetylcholinesterase is released, but of the inputs to it. In order to explore this phenomenon at the behavioral level, a novel system has been developed for detecting release of acetylcholinesterase "on-line." It can be seen that release of this protein within the substantia nigra can reflect, but is not causal to, movement. 3. Once released, the possible actions of acetylcholinesterase can be studied at both the cellular and the behavioral level. Independent of its catalytic site, acetylcholinesterase has a "modulatory" action on nigrostriatal neurons. The functional consequences of this modulation would be to enhance the sensitivity of the cells to synaptic inputs. 4. Many basic questions remain regarding the release and action of acetylcholinesterase within the substantia nigra and, indeed, within other areas of the brain. Nonetheless, tentative conclusions can be formulated that begin, in a new way, to provide a link between cellular mechanisms and the control of movement.
Cell Mol Neurobiol 1991 Feb
PMID:A noncholinergic action of acetylcholinesterase (AChE) in the brain: from neuronal secretion to the generation of movement. 201 59

1. Long before onset of synaptogenesis in the chicken neural tube, the closely related enzymes butyrylcholinesterase (BChE) and acetylcholinesterase (AChE) are expressed in a mutually exclusive manner. Accordingly, neuroblasts on the ventricular side of the neural tube transiently express BChE before they abruptly accumulate AChE while approaching the outer brain surface. 2. By exploiting AChE as a sensitive and early histochemical differentiation marker, we have demonstrated complex polycentric waves of differentiation spreading upon the cranial part of the chicken neural tube but a smooth rostrocaudal wave along the spinal cord. Shortly after expression of AChE, these cells extend long projecting neurites. In particular, segmented spinal motor axons originate from AChE-positive motoneurones; they navigate through a BChE-active zone within the rostral half of the sclerotomes before contacting BChE/AChE-positive myotome cells. At synaptogenetic stages, cholinesterases additionally are detectable in neurofibrillar laminae foreshadowing the establishment of cholinergic synapses. 3. In order to elucidate the functional significance of cholinesterases at early stages, we have investigated specific cholinesterase molecules and their mechanisms of action in vivo and in vitro. A developmental shift from the low molecular weight forms to the tetramers of both enzymes has been determined. In vitro, the addition of a selective BChE inhibitor leads to a reduction of AChE gene expression. Thus, in vivo and in vitro data suggest roles of cholinesterases in the regulation of cell proliferation and neurite growth. 4. Future research has to show whether neurogenetic functioning of cholinesterases can help to understand their reported alterations in neural tube defects, mental retardations, dementias and in some tumours.
Cell Mol Neurobiol 1991 Feb
PMID:Cholinesterases during development of the avian nervous system. 201 60


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