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:3.1.1.7 (acetylcholinesterase)
28,390 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The aim of the work was to elucidate the role of water in the reaction between acetylcholinesterase (acetylcholine hydrolase, EC 3.1.1.7) and methanesulfonyl fluoride, accelerated by accelerators. The reaction between the enzyme and methanesulfonyl fluoride in the presence of individual monovalent cations of the Hofmeister series was investigated. The results obtained were analyzed in comparison with the effect of methanesulfonylation of the specific accelerators tetramethylammonium and tetraethylammonium under various experimental conditions. The monovalent cations of the Hofmeister series accelerate the reaction. Their effect--as well as that of specific accelerators--significantly correlates with the effect of these agents on the structure of water. These findings, together with others, led to the following model of the role of hydration water in acylation of acetylcholinesterase. The accelerator, which may also be the cationic head of the natural substrate, binds to the anionic site of the enzyme and reduces the hydration of the nucleophilic serine -OH in the esteratic site, thus enhancing the nucleophilicity of -OH. This results in an improvement of the binding between the acylating agent and the esteratic site of acetylcholinesterase.
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PMID:The role of hydration in an enzyme reaction. 356 86

Proteolytic fragmentation of [3H]diisopropylfluorophosphate-labelled catalytic subunits of different molecular forms of acetylcholinesterase demonstrates that all forms extracted from the electric organ from Torpedo marmorata are true acetylcholinesterases. This is supported by immunochemical results showing that the radiolabelled polypeptides are readily recognized by specific anti-acetylcholinesterase antibodies. Although distinct structural differences exist, all forms contain a similar peptide carrying the serine hydroxyl of the esteratic subsite. Dimeric, detergent-soluble acetylcholinesterase is present in the low-salt-soluble extract (Mr of the catalytic subunit 66,000) together with a monomeric form (apparent Mr 76,000). This monomeric polypeptide is hydrophilic, enzymatically inactive, and might represent a precursor of the asymmetric forms of acetylcholinesterase.
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PMID:Inactive monomeric acetylcholinesterase in the low-salt-soluble extract of the electric organ from Torpedo marmorata. 359 80

The trichothecene T-2 toxin was rapidly hydrolyzed by rat liver microsomal fraction into HT-2 toxin which was the main metabolite. The metabolism was completely blocked by paraoxon, a serine esterase inhibitor, but not affected by EDTA or 4-hydroxy mercury benzoate, inhibitors of arylesterase and esterases containing SH-group in active site, respectively. Among the serine esterases carboxylesterase (EC 3.1.1.1), but not cholinesterase (EC 3.1.1.8) hydrolysed T-2 toxin to HT-2 toxin. Carboxylesterase activity from liver microsomes was separated into at least five different isoenzymes by isoelectric focusing, and only the isoenzyme of pI 5.4 was able to hydrolyse T-2 toxin to HT-2 toxin. The toxicity of T-2 toxin in mice was enhanced by pre-treatment with tri-o-cresyl phosphate (TOCP), a specific carboxylesterase inhibitor. This confirms the importance of carboxylesterase in detoxification of trichothecenes.
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PMID:Metabolism of T-2 toxin by rat liver carboxylesterase. 370 11

Active-site tryptic peptides were isolated from three genetic types of human serum cholinesterase. The active-site peptide was identified by labeling the active-site serine with [3H]diisopropylfluorophosphate. Peptides were purified by high-performance liquid chromatography. Amino acid composition and sequence analysis showed that the peptide from the usual genotype contained 29 residues with the sequence Ser-Val-Thr-Leu-Phe-Gly-Glu-Ser-Ala-Gly-Ala-Ala-Ser-Val-Ser-Leu-His-Leu- Leu-Ser-Pro-Gly-Ser-His-Ser-Leu-Phe-Thr-Arg. The active-site serine was the eighth residue from the N-terminal. The peptide containing the active-site serine from the atypical genotype contained 22 residues with the sequence Ser-Val-Thr-Leu-Phe-Gly-Glu-Ser-Ala-Gly-Ala-Ala-Ser-Val-Ser-Leu-His-Leu- Leu-Ser-Pro-Gly. The peptide from the atypical-silent genotype contained eight residues with the sequence Gly-Glu-Ser-Ala-Gly-Ala-Ala-Ser. Thus, the sequences of the atypical and atypical-silent active-site peptides were identical to the corresponding portions of the usual peptide.
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PMID:Amino acid sequence of the active site of human serum cholinesterase from usual, atypical, and atypical-silent genotypes. 374 70

Cholinesterases are serine esterases that rapidly hydrolyze the neurotransmitter acetylcholine. In humans, cholinesterases exhibit extensive polymorphism in terms of their substrate specificity, sensitivity to selective inhibitors, hydrophobicity, and cellular as well as subcellular localization. It is not yet known whether the various cholinesterase forms originate from different genes or are products of posttranscriptional and posttranslational processing. The extent to which these enzyme forms are homologous in their amino acid sequence is also not known. However, a consensus organophosphate-binding hexapeptide sequence Phe-Gly-Glu-Ser-Ala-Gly was found both in "true" acetylcholinesterase from the electric organ of Torpedo [McPhee-Quigley et al: J Biol Chem 260:12185-12189, 1985] and in "pseudocholinesterase" (butyrylcholinesterase) from human serum [Lockridge: "Cholinesterases--Fundamental and Applied Aspects." New York: de Gruyter pp 5-12, 1984], suggesting that this region in the protein is conserved in all cholinesterases. Based on this common sequence, we prepared synthetic oligodeoxynucleotides and used them as labeled probes to screen a cDNA library from fetal human brain mRNA, cloned in lambda gt10 phages. A cDNA clone of 770 nucleotides in length was isolated. It contains an open reading frame terminating with the sequence Ser-Val-Thr-Leu-Phe-Gly-Glu-Ser-Ala-Gly-Ala-Ala, which includes the consensus hexapeptide used for designing the DNA probe. Furthermore, the sequence of this 12-amino acid peptide is identical to the sequence reported for the organophosphate binding site of human serum pseudocholinesterase [Lockridge: "Cholinesterases--Fundamental and Applied Aspects." New York: de Gruyter, pp 5-12, 1984]. These findings confirm that the isolated clone is indeed part of a human cholinesterase cDNA.
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PMID:Use of synthetic oligodeoxynucleotide probes for the isolation of a human cholinesterase cDNA clone. 375 63

Two distinct classes of acetylcholinesterase exist in near equal amounts in the electric organ of Torpedo californica. A globular 5.6 S form is a dimer which possesses a hydrophobic region. The second form is present as elongated species that sediment at 17 and 13 S and contain structural subunits disulfide-linked to the catalytic subunits. Removal of the structural subunits by mild proteolysis yields a tetramer of catalytic subunits which sediments at 11 S. To compare the primary structures of the catalytic subunits of the 5.6 S and 11 S forms of acetylcholinesterase, amino acid sequences from the active sites and from the amino-terminal regions have been elucidated. Active site serines were labeled with [3H]isopropyl fluorophosphate. After digestion with trypsin, the resultant peptides were resolved by elution from a size-exclusion column followed by reverse-phase high performance liquid chromatography. Each active site tryptic peptide contained 24 residues and identical sequences were found in this peptide for the 5.6 S and 11 S forms of the enzyme. The sequence flanking the active site serine revealed extensive homology with the published sequence of human serum cholinesterase as well as a lesser degree of homology with other known serine proteases and esterases. The sequences of the amino-terminal region also appear to be identical for both enzyme forms although we note variation in the ratio of Glu and Gln at position 5. The amino-terminal sequence exhibits only partial homology with the published sequence of human serum cholinesterase.
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PMID:Primary structures of the catalytic subunits from two molecular forms of acetylcholinesterase. A comparison of NH2-terminal and active center sequences. 390 71

Steady-state and time-correlated fluorescence polarizations have been examined for selected complexes and covalent conjugates of the 11S and (17 + 13)S forms of Torpedo acetylcholinesterase. The 11S form exists as a tetramer of apparently identical subunits, whereas the (17 + 13)S forms contain two or three sets of tetramers disulfide-linked to an elongated collagen-like tail unit. Pyrenebutyl methylphosphonofluoridate and (dansylsulfonamido)pentyl methylphosphonofluoridate were conjugated at the active center serine whereas propidium was employed as a fluorescent ligand for the spatially removed peripheral anionic site. Steady-state polarization of the pyrenebutyl conjugates indicates rotational correlation times of approximately 400 ns for the 11S species and greater than 1100 ns for the (17 + 13)S species. Hence, the tail unit severely restricts rotational motion of the catalytic subunits. Time-correlated fluorescence polarization analysis of the 11S species indicates multiple rotational correlation times. Anisotropy decay of the propidium complex (tau = 6 ns) occurs in exponential manner with a rotational correlation time of approximately 150 ns, while covalent adducts at the active center exhibit rotational correlation times greater than or equal to 300 ns. Anisotropy decay of the (dansylsulfonamido)pentyl conjugate (tau = 16 ns) appears exponential with a correlation time of approximately 320 ns, whereas decay of the pyrenebutyl conjugate (tau = 100 ns) is described by two correlation times, phi S = 18 ns and phi L = 320 ns, of small (15%) and large (85%) amplitudes, respectively. Two limiting models have been considered to explain the results.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Flexibility of the molecular forms of acetylcholinesterase measured with steady-state and time-correlated fluorescence polarization spectroscopy. 408 69

The transfer of detergent solubilized and purified gamma-glutamyl transpeptidase (gamma-GTase), of hog kidney cortex, from proteoliposomes into human erythrocyte ghost membranes has been studied. The transfer of gamma-glutamyl transpeptidase was observed upon incubation of gamma-GTase incorporated dipalmitoylphosphatidylcholine vesicles with erythrocyte ghost membranes at 37 degrees C for 12 h. The extent of transfer was dependent upon the fluidity of donor proteoliposomes, being more when dipalmitoylphosphatidylcholine proteoliposomes were used compared to dimyristoylphosphatidylcholine, and intermediate values were observed when binary mixtures of DMPC and DPPC were used. Moreover, the transfer of gamma-GTase was facilitated when rigid basic phospholipid proteoliposomes were used as donor. The transfer of gamma-GTase has been observed to be associated with the removal of intrinsic membrane proteins and lipids from erythrocytes, mainly acetylcholinesterase, sphingomyelin, and cholesterol. An enhancement in the fluorescence due to resonance energy transfer was observed when ghost membranes containing fluorescent donor probe were incubated with proteoliposomes containing fluorescent acceptor probe, indicating that fusion but not adsorption of vesicles occurs during the transfer process. However, the inability of entrapped [14C]-sucrose delivery from proteoliposomes into ghost membrane vesicle suggest that fusion per se is not primarily involved in the transfer process. It appears that the transfer of gamma-glutamyl transpeptidase occurs by a collisional transfer process resulting in intermembrane protein transfer. The gamma-glutamyl transpeptidase implanted ghost membranes exhibited the uptake of L-glutamate which was inhibited by serine-borate, an inhibitor of transpeptidase activity. In addition, the uptake of L-glutamate was inhibited by the dipeptide gamma-glutamyl-L-glutamate, thus supporting the proposed role of gamma-glutamyl transpeptidase in the uptake of amino acids in biological membranes.
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PMID:Proteoliposome interaction with human erythrocyte membranes. Functional implantation of gamma-glutamyl transpeptidase. 612 71

In this study, the effect of sixteen different enzymes on serum C1 and its subcomponents was investigated. The sixteen enzymes could be divided into three groups. First, enzymes which activate native C1: trypsin (optimal concentration 2.4 x 10(-4) mM); alpha-chymotrypsin (2.3 x 10(3) mM); thrombin (1.0 x 10(-5) mM); plasmin (1.9 x 10(-5) mM); elastase (5.8 x 10(-5) mM); pronase (3.0 x 10(-6) mM). All these enzymes are serine esterase and activate native serum C1 bound to EAC4 at the given concentration within 10 min at 30 degrees C. Furthermore, native C1 inhibited by a pentosanpolysulfoester, Sp54, is unable to undergo the internal activation but can be externally activated by the serine esterases. Second, enzymes which do not activate native C1 but result in a dose and time-dependent loss of C1 activity: collagenase; pepsin; carboxypeptidase B. Third, enzymes which have no effect on C1 and C1: Lysozyme; neuraminidase; beta-galactosidase; L-amino acid oxidase; arginase; streptokinase, and acetylcholinesterase.
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PMID:Activation of the first component of complement, C1: comparison of the effect of sixteen different enzymes on serum C1. 619 90

The interaction of serine protease (esterases) with 6-chloro-2-pyrones was investigated. Time-dependent inactivation of chymotrypsin, alpha-lytic protease, pig liver elastase, and cholinesterase was found with 3- and 5-benzyl-6-chloro-2-pyrone, as well as 3- and 5-methyl-6-chloro-2-pyrone. No inactivation was observed with the unsubstituted 6-chloro-2-pyrone. The substituted pyrones did not inactivate papain or carboxypeptidase A, as well as a number of other nonproteolytic enzymes. The substituted chloropyrones, therefore, show considerable selectivity toward serine proteases. Analogues in which the 6-chloro substituent is replaced by H or OH do not inactivate. The presence of the halogen is, therefore, essential for inactivation. Chymotrypsin catalyzes the hydrolysis of 3-benzyl-6-chloro-2-pyrone. At pH 7.5, (E)-4-benzyl-2-pentenedioic acid is the major product, and 2-benzyl-2-pentenedioic anhydride is a minor product. The ration of hydrolysis product found to the number of enzyme molecules inactivated varies from 14 to 40. The enzyme inactivated with the 3-benzyl compound does not show a spectrum characteristic of the pyrone ring. This suggests that inactivation by 3-benzyl-6-chloro-2-pyrone occurs in a mechanism-based fashion after enzymatic lactone hydrolysis. When the enzyme is inactivated with the 5-benzyl compound, absorbance due to the pyrone ring is observed. We suggest that inactivation occurs through an active site directed mechanism involving a 1,6-conjugate addition of an active site nucleophile to the pyrone ring.
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PMID:Novel inactivators of serine proteases based on 6-chloro-2-pyrone. 641 Nov 20


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