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)

Evidence for the involvement of Ser-203, His-447, and Glu-334 in the catalytic triad of human acetylcholinesterase was provided by substitution of these amino acids by alanine residues. Of 20 amino acid positions mutated so far in human acetylcholinesterase (AChE), these three were unique in abolishing detectable enzymatic activity (less than 0.0003 of wild type), yet allowing proper production, folding, and secretion. This is the first biochemical evidence for the involvement of a glutamate in a hydrolase triad (Schrag, J.D., Li, Y., Wu, M., and Cygler, M. (1991) Nature 351, 761-764), supporting the x-ray crystal structure data of the Torpedo californica acetylcholinesterase (Sussman, J.L., Harel, M., Frolow, F., Oefner, C., Goldman, A., Toker, L. and Silman, I. (1991) Science 253, 872-879). Attempts to convert the AChE triad into a Cys-His-Glu or Ser-His-Asp configuration by site-directed mutagenesis did not yield effective AChE activity. Another type of substitution, that of Asp-74 by Gly or Asn, generated an active enzyme with increased resistance to succinylcholine and dibucaine; thus mimicking in an AChE molecule the phenotype of the atypical butyrylcholinesterase natural variant (D70G mutation). Mutations of other carboxylic residues Glu-84, Asp-95, Asp-333, and Asp-349, all conserved among cholinesterases, did not result in detectable alteration in the recombinant AChE, although polypeptide productivity of the D95N mutant was considerably lower. In contrast, complete absence of secreted human AChE polypeptide was observed when Asp-175 or Asp-404 were substituted by Asn. These two aspartates are conserved in the entire cholinesterase/thyroglobulin family and appear to play a role in generating and/or maintaining the folded state of the polypeptide. The x-ray structure of the Torpedo acetylcholinesterase supports this assumption by revealing the participation of these residues in salt bridges between neighboring secondary structure elements.
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PMID:Mutagenesis of human acetylcholinesterase. Identification of residues involved in catalytic activity and in polypeptide folding. 151 12

The structure and some functional sites of human milk bile salt activated lipase (BAL) were studied by cDNA cloning and chemical analysis of the enzyme. Eighteen cDNA clones of human BAL were identified from lactating human breast cDNA libraries in lambda gt11 and lambda gt10 with antibody and synthetic oligonucleotides as probes. The sequence of four clones was sufficient to construct a 3018-bp BAL cDNA structure. This sequence codes for an open reading frame of 742 amino acid residues. There is a putative signal sequence of 20 residues which is followed by the amino-terminal sequence of BAL, and the mature BAL contains 722 amino acid residues. The cDNA sequence also contains a 678-base 5'-untranslated sequence, a 97-base 3'-untranslated region, and a 14-base poly(A) tail. The sequence of a 1.8-kbp insert of clone G10-4A differs from that of the other cDNA in that it contains a deletion of 198 bases (1966-2163) corresponding to 66 amino acid residues. By use of BAL cDNA as probe, it was found that the major molecular species of BAL mRNA in human mammary gland HBL-100 cells had a size of 2.9 kb and two minor species had sizes of 3.8 and 5.1 kb by Northern blot analyses. The deduced BAL protein structure contains in the carboxyl-terminal region 16 repeating units of 11 amino acids each. The repeating units have the basic structure Pro-Val-Pro-Pro-Thr-Gly-Asp-Ser-Gly-Ala-Pro with only minor substitutions. The amino acid sequence of human BAL is related to that of pancreatic lysophospholipase, cholesterol esterase, cholinesterase, acetylcholinesterase, and thyroglobulin. Ten of the 14 cyanogen bromide fragments of diisopropyl fluorophosphate inhibited human milk BAL were isolated, determined for N-terminal sequences, analyzed for amino sugars, and tested for some functional properties. These chemical studies established that the active site of human milk BAL is located at serine-194, the N-glycosylation site is present at asparagine-187, the O-glycosylation region is in the 16 repeating units near the C-terminus, and the heparin binding domain is in the N-terminal region. We have also determined the location of disulfide bridges as Cys64-Cys80 and Cys246-Cys257. The cyanogen bromide cleavage and the partial sequencing of CNBr peptides also confirmed the location of methionines in the polypeptide chain as well as the deduced cDNA sequence of BAL.
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PMID:Structure of human milk bile salt activated lipase. 198 41

The complete amino acid sequence of a mammalian acetylcholinesterase from fetal bovine serum (FBS AChE) is presented. This enzyme has a high degree of sequence identity with other cholinesterases, liver carboxyesterases, esterase-6, lysophospholipase, and thyroglobulin. The locations of 191 amino acids in 10 regions of the FBS enzyme were compared with corresponding sequences of Torpedo, human, and Drosophila AChEs and human serum butyrylcholinesterase (BChE). In one region there is a marked difference in both the number of amino acids and their sequence between mammalian AChE and other AChEs and the human serum BChE. The amino acid sequence of FBS AChE showed overall homologies of 90% with human AChE, 60% with T. california AChE, 50% with human serum BChE, and 39% with Drosophila AChE in these regions.
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PMID:Complete amino acid sequence of fetal bovine serum acetylcholinesterase and its comparison in various regions with other cholinesterases. 236 60

The nature of the putative autoantigen in Graves' ophthalmopathy (Go) remains an enigma but the sequence similarity between thyroglobulin (Tg) and acetylcholinesterase (ACHE) provides a rationale for epitopes which are common to the thyroid gland and the eye orbit. In an attempt to define the shared epitope, we have screened a lambda gt 11 human thyroid cDNA library using a polyclonal antibody to Torpedo ACHE and isolated two clones, which upon sequencing, were shown to contain Tg segments, corresponding to portions of the C terminal part of the molecule which has a high similarity with ACHE. Having demonstrated the existence of an epitope common to Tg and ACHE, the clones have been further tested and found to be positive in lysis plaque assays with 1/10 sera from patients with Hashimoto's thyroiditis (HT), 8/8 from patients with Graves' ophthalmopathy and 0/8 normal sera. We have investigated the physiological significance of this common epitope by in situ immunolocalization studies in which the polyclonal antibody to Torpedo ACHE (which was used for screening the library) and immunoglobulins (Igs) from 6 Go patients tested were shown to bind to end plate regions of human foetal muscle fibres which were concurrently shown to be rich in cholinesterase activity: Igs from 3 normal individuals and 2 patients with Hashimoto's thyroiditis did not bind. The results demonstrate and characterize an epitope which is common to Tg and ACHE and show that Go patients Igs contain antibodies which bind to muscle end plates rich in cholinesterase. The significance of these findings to the pathogenesis of Go is discussed.
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PMID:Definition, at the molecular level, of a thyroglobulin-acetylcholinesterase shared epitope: study of its pathophysiological significance in patients with Graves' ophthalmopathy. 248 81

Cholinesterases (ChEs) are highly polymorphic proteins, capable of rapidly hydrolyzing the neurotransmitter acetylcholine and involved in terminating neurotransmission in neuromuscular junctions and cholinergic synapses. In an attempt to delineate the structure and detailed properties of the human protein(s) and the gene(s) coding for the acetylcholine hydrolyzing enzymes, a human cDNA coding for ChE was isolated by use of oligodeoxynucleotide screening of cDNA libraries. For this purpose, a method for increasing the effectiveness of oligonucleotide screening by introducing deoxyinosine in sites of codon ambiguity and using tetramethyl-ammonium salt washes to remove false-positive hybrids was employed. The resulting isolated 2.4-kilobase (kb) cholinesterase cDNA sequences encode for the entire mature secretory protein, preceded by an N-terminal signal peptide. The human ChE primary sequence shows almost no homology to other serine hydrolases, with the exception of a hexapeptide at the active site. In contrast, it displays extensive homology with acetylcholinesterase form Torpedo californica and Drosophila melanogaster as well as with bovine thyroglobulin. These extensive homologies probably suggest the need of the entire coding sequence for the physiological function(s) fulfilled by the enzyme and further suggest a common, unique, ancestral gene for these cDNAs. In turn, the cDNA was used as a probe to isolate genomic DNA sequences for the 5'-region of the human ChE gene. The genomic DNA fragment encoding part of the 5'-region of ChEcDNA was detected by DNA blot hybridization, enriched 70-fold by gel electrophoresis and electroelution, cloned in lambda phage and isolated. Sequencing of the cloned DNA revealed that it did indeed include part of the 5'-region of ChEcDNA, starting at an adjacent 5'-position to the nucleotides coding for the initiator methionine, and ending with an EcoRI restriction site inherent to the ChEcDNA sequence. The isolated fragment of the human cholinesterase gene is currently employed to complete the structural characterization of this and related genes.
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PMID:Molecular biological search for human genes encoding cholinesterases. 307 58

The appearance and distribution of acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE) in 12 human thyroid cancers and three normal thyroids were examined by electron microscopic study with indirect thiocholine method. The demonstration of AChE and BuChE activities in only two of nine cases of follicular and papillary carcinoma examined and none of the three cases of medullary carcinoma shows that the cholinesterases are not specific enzymes for the thyroid tumors. In normal thyroid tissue samples examined, no activities of AChE and BuChE were detected. On ultrastructural level AChE reaction product was revealed in the perinuclear space, in the endoplasmic reticulum, and in the Golgi complex of some but not in all cells in less-differentiated regions of the tumors. In contrast to the distribution of AChE, no staining for BuChE was noted in the Golgi elements. Ultrastructural localization of AChE activity in the thyroid cancer cells corresponds exactly to the current understanding of glycoproteins synthesis and processing in normal cells. The authors postulate that the copy of AChE gene suppressed in normal thyroid epithelium cells may be expressed in some follicular thyroid carcinoma cells. Their hypothesis is logical on the basis of recent finding of a significant homology between AChE and thyroglobulin.
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PMID:Acetylcholinesterase and butyrylcholinesterase activities in human thyroid cancer cells. 333 19

The 60-kDa esterase was isolated from liver microsomes of 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced rabbits and its complete amino acid sequence determined. Automated sequence analysis of intact protein, as well as characterization of the peptides obtained from enzymatic and chemical cleavages, led to the elucidation of the primary structure. The protein is a single polypeptide consisting of 539 residues and molecular weight 59,478. The active site serine is 195, and another diisopropylphospho binding site is at histidyl 441. Carbohydrate chains are attached at aspariginyl residues 61 and 363. Although 2,3,7,8-tetrachlorodibenzo-p-dioxin treatment induces this esterase severalfold, the amino acid sequence of the induced enzyme is identical to that of the enzyme isolated from liver microsomes of untreated rabbits. The sequence of the microsomal esterase is 30% identical with the sequences of human serum cholinesterase and the acetylcholinesterase from Torpedo californica. There is also a close homology between the 60-kDa esterase and the COOH-terminal domain of bovine thyroglobulin.
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PMID:Complete covalent structure of 60-kDa esterase isolated from 2,3,7,8-tetrachlorodibenzo-p-dioxin-induced rabbit liver microsomes. 334 53

The complete amino acid sequence of human serum cholinesterase (choline esterase II (unspecific), EC 3.1.1.8) was determined by Edman degradation of purified peptides. The protein contains 574 amino acids per subunit and nine carbohydrate chains attached to 9 asparagines. The four subunits of cholinesterase appear to be identical. The active site serine is the 198th residue from the amino terminus. The sequence of human serum cholinesterase is 53.8% identical with the sequence of acetylcholinesterase from Torpedo californica and 28% identical with the carboxyl-terminal portion of bovine thyroglobulin.
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PMID:Complete amino acid sequence of human serum cholinesterase. 354 89

Cholinesterases (acetylcholinesterase and butyrylcholinesterase) exhibit additional catalytic activities apart from their well-known action in hydrolyzing choline esters. An amine-sensitive aryl acylamidase activity is exhibited by both acetyl- and butyrylcholinesterases. A metallocarboxypeptidase-like activity is found associated with both acetyl- and butyrylcholinesterases. The peptidase activity exhibited by butyrylcholinesterase was located in a 50-kDa COOH-terminal fragment. Acetylcholinesterase is implicated in noncholinergic functions in the substantia nigra. A relationship between tumorigenesis, cell differentiation, and cholinesterases has been speculated. The sequence similarities between different esterases, lipases, thyroglobulin, cell adhesion proteins, and cholinesterases would make it appear that cholinesterases are capable of exhibiting more than one biological activity and their functions are wider than what is hitherto known.
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PMID:Noncholinergic functions of cholinesterases. 822 8

The disulfide bonds and N-glycosylation sites in a glycoprotein from the Rathke's gland secretion of the Kemp's ridley turtle (Lepidochelys kempi) have been characterized with respect to peptide sequences and glycan structures. The glycoprotein constitutes about 70% of the total protein in the secretion, and based on partial sequence information, it shows more than 20% identity with both the catalytic (esterases) and the noncatalytic (thyroglobulin) members of the esterase/lipase family of proteins. For the determination of the disulfide locations, the glycoprotein was digested with chymotrypsin, and the three HPLC peptide peaks yielding fluorescent products after treatment with tributylphosphine (Bu3P) and 4-(aminosulfonyl)-7-fluoro-2,1,3-benzoxadiazole (ABD-F) were collected. The three fractions were treated with the same reagents in separate experiments, the resulting pairs of ABD-Cys-containing peptides were separated by HPLC, and the sequence of each individual peptide was determined. The peptide identity established that three disulfide bonds existed in the glycoprotein: Cys 65-Cys 91, Cys 254-Cys 265, and Cys 130-Cys 404; the first two of these are conserved in all the members of the esterase family. For the study of the glycosylation sites, the glycoprotein was reduced with Bu3P and the SH groups covalently blocked with ABD-F, and the resulting product was digested with chymotrypsin. The glycopeptides were isolated by affinity chromatography, separated by reverse-phase HPLC, and subjected to sequence analysis and fast atom bombardment mass spectrometry before and after separation of the glycans and the peptides through the action of glycoamidase. Three separate glycosylation sites were identified, each containing multiple glycans. The sugar analyses of the hydrolysates of the glycoprotein indicated that only GlcNAc and Man were present as building blocks, and the mass spectrometric data showed that Man3GlcNAc2-, GlcNAc2-4Man3GlcNAc2-, and possibly GlcNAc2Man2GlcNAc2- were the major glycan structures, distributed differently at the three sites. The three glycosylation sites match three of the nine sites glycosylated in human serum choline esterase, and one of them, Asn 106, is also found as one of two glycosylation sites in the homologous segment of thyroglobulin.
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PMID:Characterization of the disulfide bonds and the N-glycosylation sites in the glycoprotein from Rathke's gland secretions of Kemp's ridley sea turtle (Lepidochelys kempi). 878 16


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