Bartels CF

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Full name : Bartels Cynthia F

First name : Cynthia F

Mail : Department of Genetics and Genome Sciences, Case Western Reserve University, 10900 Euclid Ave, Cleveland, OH 44106

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Country : USA

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References (46)

Title : Reaction kinetics of biotinylated organophosphorus toxicant, FP-biotin, with human acetylcholinesterase and human butyrylcholinesterase - Schopfer_2005_Chem.Res.Toxicol_18_747
Author(s) : Schopfer LM , Voelker T , Bartels CF , Thompson CM , Lockridge O
Ref : Chemical Research in Toxicology , 18 :747 , 2005
Abstract : A biotinylated organophosphate could be useful for identifying proteins that react with organophosphorus toxicants (OP). FP-biotin, 10-(fluoroethoxyphosphinyl)-N-(biotinamidopentyl)decanamide, was synthesized and found to be stable in methanol and chloroform but less stable in water. Because acetylcholinesterase (AChE, EC 3.1.1.7) and butyrylcholinesterase (BChE, EC 3.1.1.8) are known to be sensitive targets of OP, their reactivity with FP-biotin was tested. The rate constant for reaction with human AChE was 1.8 x 10(7) M(-1) min(-1), and for human BChE, it was 1.6 x 10(8) M(-1) min(-1). A phosphorus stereoisomer, constituting about 50% of the FP-biotin preparation, appeared to be the reactive species. The binding affinity was estimated to be >85 nM for AChE and >5.8 nM for BChE. It was concluded that FP-biotin is a potent OP, well-suited for searching for new biomarkers of OP exposure.
ESTHER : Schopfer_2005_Chem.Res.Toxicol_18_747
PubMedSearch : Schopfer_2005_Chem.Res.Toxicol_18_747
PubMedID: 15833035

Title : High activity of human butyrylcholinesterase at low pH in the presence of excess butyrylthiocholine - Masson_2003_Eur.J.Biochem_270_315
Author(s) : Masson P , Nachon F , Bartels CF , Froment MT , Ribes F , Matthews C , Lockridge O
Ref : European Journal of Biochemistry , 270 :315 , 2003
Abstract : Butyrylcholinesterase is a serine esterase, closely related to acetylcholinesterase. Both enzymes employ a catalytic triad mechanism for catalysis, similar to that used by serine proteases such as alpha-chymotrypsin. Enzymes of this type are generally considered to be inactive at pH values below 5, because the histidine member of the catalytic triad becomes protonated. We have found that butyrylcholinesterase retains activity at pH
ESTHER : Masson_2003_Eur.J.Biochem_270_315
PubMedSearch : Masson_2003_Eur.J.Biochem_270_315
PubMedID: 12605682

Title : Substrate activation in acetylcholinesterase induced by low pH or mutation in the pi-cation subsite - Masson_2002_Biochim.Biophys.Acta_1594_313
Author(s) : Masson P , Schopfer LM , Bartels CF , Froment MT , Ribes F , Nachon F , Lockridge O
Ref : Biochimica & Biophysica Acta , 1594 :313 , 2002
Abstract : Substrate inhibition is considered a defining property of acetylcholinesterase (AChE), whereas substrate activation is characteristic of butyrylcholinesterase (BuChE). To understand the mechanism of substrate inhibition, the pH dependence of acetylthiocholine hydrolysis by AChE was studied between pH 5 and 8. Wild-type human AChE and its mutants Y337G and Y337W, as well as wild-type Bungarus fasciatus AChE and its mutants Y333G, Y333A and Y333W were studied. The pH profile results were unexpected. Instead of substrate inhibition, wild-type AChE and all mutants showed substrate activation at low pH. At high pH, there was substrate inhibition for wild-type AChE and for the mutant with tryptophan in the pi-cation subsite, but substrate activation for mutants containing small residues, glycine or alanine. This is particularly apparent in the B. fasciatus AChE. Thus a single amino acid substitution in the pi-cation site, from the aromatic tyrosine of B. fasciatus AChE to the alanine of BuChE, caused AChE to behave like BuChE. Excess substrate binds to the peripheral anionic site (PAS) of AChE. The finding that AChE is activated by excess substrate supports the idea that binding of a second substrate molecule to the PAS induces a conformational change that reorganizes the active site.
ESTHER : Masson_2002_Biochim.Biophys.Acta_1594_313
PubMedSearch : Masson_2002_Biochim.Biophys.Acta_1594_313
PubMedID: 11904227

Title : Wild-type and A328W mutant human butyrylcholinesterase tetramers expressed in Chinese hamster ovary cells have a 16-hour half-life in the circulation and protect mice from cocaine toxicity - Duysen_2002_J.Pharmacol.Exp.Ther_302_751
Author(s) : Duysen EG , Bartels CF , Lockridge O
Ref : Journal of Pharmacology & Experimental Therapeutics , 302 :751 , 2002
Abstract : Human butyrylcholinesterase (BChE) hydrolyzes cocaine to inactive metabolites. A mutant of human BChE, A328W, hydrolyzed cocaine 15-fold faster compared with wild-type BChE. Although the catalytic properties of human BChE secreted by Chinese hamster ovary (CHO) cells are identical to those of native BChE, a major difference became evident when the recombinant BChE was injected into rats and mice. Recombinant BChE disappeared from the circulation within minutes, whereas native BChE stayed in the blood for a week. Nondenaturing gel electrophoresis showed that the recombinant BChE consisted mainly of monomers and dimers. In contrast, native BChE is a tetramer. The problem of the short residence time was solved by finding a method to assemble the recombinant BChE into tetramers. Coexpression in CHO cells of BChE and 45 residues from the N terminus of the COLQ protein yielded 70% tetrameric BChE. The resulting purified recombinant BChE tetramers had a half-life of 16 h in the circulation of rats and mice. The 16-h half-life was achieved without modifying the carbohydrate content of recombinant BChE. The protective effect of recombinant wild-type and A328W mutant BChE against cocaine toxicity was tested by measuring locomotor activity in mice. Pretreatment with wild-type BChE or A328W tetramers at a dose of 2.8 units/g i.p. reduced cocaine-induced locomotor activity by 50 and 80%. These results indicate that recombinant human BChE could be useful for treating cocaine toxicity in humans.
ESTHER : Duysen_2002_J.Pharmacol.Exp.Ther_302_751
PubMedSearch : Duysen_2002_J.Pharmacol.Exp.Ther_302_751
PubMedID: 12130740

Title : The active site of human paraoxonase (PON1) - Josse_2001_J.Appl.Toxicol_21 Suppl 1_S7
Author(s) : Josse D , Lockridge O , Xie W , Bartels CF , Schopfer LM , Masson P
Ref : J Appl Toxicol , 21 Suppl 1 :S7 , 2001
Abstract : Ideally we would like to treat people exposed to nerve agents with an enzyme that rapidly destroys nerve agents. The enzymes considered for such a role include human butyrylcholinesterase (BChE), acetylcholinesterase (AChE), carboxylesterase and paraoxonase (PON1). Success has been achieved in endowing BChE with the ability to hydrolyze organophosphates. The G117H mutant of BCHE hydrolyzes sarin and VX, whereas the double mutant G117H/E197Q hydrolyzes soman (Millard et al. Biochemistry 1995; 34: 15925-15933; 1998; 37: 237-247). However, the rates of organophosphate hydrolysis are slow and a faster organophosphate hydrolase is being sought. Native PON1 hydrolyzes paraoxon with a catalytic efficiency, of 2.4 x 10(6) M(-1) x min(-1), and our goal is to improve the organophosphate hydrolase activity of PON1. To achieve this we need to identify the amino acids in the active site of PON1. Using site-directed mutagenesis and expression in human 293T cells, we have identified the following eight amino acids as being essential to PON1 activity: W280, H114, H133, H154, H242, H284, E52 and D53. Fluorescence of PON1 complexed to terbium ion shows that at least one tryptophan is close to the calcium binding site.
ESTHER : Josse_2001_J.Appl.Toxicol_21 Suppl 1_S7
PubMedSearch : Josse_2001_J.Appl.Toxicol_21 Suppl 1_S7
PubMedID: 11920913

Title : The butyrylcholinesterase K-variant shows similar cellular protein turnover and quaternary interaction to the wild-type enzyme - Altamirano_2000_J.Neurochem_74_869
Author(s) : Altamirano CV , Bartels CF , Lockridge O
Ref : Journal of Neurochemistry , 74 :869 , 2000
Abstract : A recent study has linked the butyrylcholinesterase (BChE) K-variant and the apolipoprotein epsilon4 isoform to late-onset Alzheimer's disease. These findings have been controversial and have led us to examine the differences between wild-type and K-variant BChE in enzyme activity, protein stability, and quaternary structure. J-variant BChE (E497V/A539T) was also studied because it is associated with the K-variant mutation. The K-variant mutation (A539T) is located in the C-terminal tetramerization domain. Wild-type, K-variant, and J-variant BChE were expressed in Chinese hamster ovary cells and purified. The purified enzymes had similar binding affinity (Km) values and catalytic rates for butyrylthiocholine and benzoylcholine. In pulse-chase studies the K-variant, J-variant, and wildtype BChE were degraded rapidly within the cell, with a half-time of approximately 1.5 h. Less than 5% of the intracellular BChE was exported. The C-terminal peptide containing the K-variant mutation interacted with itself as strongly as did the wild-type peptide in the yeast two-hybrid system. Both K-variant and wild-type BChE assembled into tetramers in the presence of poly-L-proline or the proline-rich attachment domain of the collagen tail. The native K-variant BChE in serum showed the same proportion of tetramers as the native serum wild-type BChE. We conclude that the K-variant BChE is similar to wild-type BChE in enzyme activity, protein turnover, and tetramer formation.
ESTHER : Altamirano_2000_J.Neurochem_74_869
PubMedSearch : Altamirano_2000_J.Neurochem_74_869
PubMedID: 10646540

Title : Determination of the DNA sequences of acetylcholinesterase and butyrylcholinesterase from cat and demonstration of the existence of both in cat plasma - Bartels_2000_Biochem.Pharmacol_60_479
Author(s) : Bartels CF , Xie W , Miller-Lindholm AK , Schopfer LM , Lockridge O
Ref : Biochemical Pharmacology , 60 :479 , 2000
Abstract : Cat serum contains 0.5 mg/L of butyrylcholinesterase (BChE, EC 3.1.1. 8) and 0.3 mg/L of acetylcholinesterase (AChE, EC 3.1.1.7); this can be compared with 5 mg/mL and < 0.01 mg/L, respectively, in human serum. Cat BChE differed from human BChE in the steady-state turnover of butyrylthiocholine, having a 3-fold higher k(cat) and 2-fold higher K(m) and K(ss) values. Sequencing of the cat BCHE cDNA revealed 70 amino acid differences between cat and human BChE, three of which could account for these kinetic differences. These amino acids, which were located in the region of the active site, were Phe398Ile, Pro285Leu, and Ala277Leu (where the first amino acid was found in human and the second in cat). Sequencing genomic DNA for cat and human ACHE demonstrated that there were 33 amino acid differences between the cat and human AChE enzymes, but that there were no differences in the active site region. In addition, a polymorphism in intron 3 of the human ACHE gene was detected, as well as a silent polymorphism at Y116 of the cat ACHE gene.
ESTHER : Bartels_2000_Biochem.Pharmacol_60_479
PubMedSearch : Bartels_2000_Biochem.Pharmacol_60_479
PubMedID: 10874122
Gene_locus related to this paper: felca-ACHE , felca-BCHE , tiger-BCHE

Title : An improved cocaine hydrolase: the A328Y mutant of human butyrylcholinesterase is 4-fold more efficient - Xie_1999_Mol.Pharmacol_55_83
Author(s) : Xie W , Altamirano CV , Bartels CF , Speirs RJ , Cashman JR , Lockridge O
Ref : Molecular Pharmacology , 55 :83 , 1999
Abstract : Butyrylcholinesterase (BChE) has a major role in cocaine detoxication. The rate at which human BChE hydrolyzes cocaine is slow, with a kcat of 3.9 min(-1) and Km of 14 microM. Our goal was to improve cocaine hydrolase activity by mutating residues near the active site. The mutant A328Y had a kcat of 10.2 min(-1) and Km of 9 microM for a 4-fold improvement in catalytic efficiency (kcat/Km). Since benzoylcholine (kcat 15,000 min(-1)) and cocaine form the same acyl-enzyme intermediate but are hydrolyzed at 4000-fold different rates, it was concluded that a step leading to formation of the acyl-enzyme intermediate was rate-limiting. BChE purified from plasma of cat, horse, and chicken was tested for cocaine hydrolase activity. Compared with human BChE, horse BChE had a 2-fold higher kcat but a lower binding affinity, cat BChE was similar to human, and chicken BChE had only 10% of the catalytic efficiency. Naturally occurring genetic variants of human BChE were tested for cocaine hydrolase activity. The J and K variants (E497V and A539T) had k(cat) and Km values similar to wild-type, but because these variants are reduced to 66 and 33% of normal levels in human blood, respectively, people with these variants may be at risk for cocaine toxicity. The atypical variant (D70G) had a 10-fold lower binding affinity for cocaine, suggesting that persons with the atypical variant of BChE may experience severe or fatal cocaine intoxication when administered a dose of cocaine that is not harmful to others.
ESTHER : Xie_1999_Mol.Pharmacol_55_83
PubMedSearch : Xie_1999_Mol.Pharmacol_55_83
PubMedID: 9882701

Title : ACHE Knockout Mouse\; Cat AChE and Cat BChE Sequences\; Tetramers of BChE -
Author(s) : Lockridge O , Xie W , Chatonnet A , Taylor P , Bartels CF
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :41 , 1998
PubMedID:

Title : Prolonged paralysis after a test dose of mivacurium in a patient with atypical serum cholinesterase -
Author(s) : Vanlinthout LE , Bartels CF , Lockridge O , Callens K , Booij LH
Ref : Anesthesia & Analgesia , 87 :1199 , 1998
PubMedID: 9806709

Title : Acetylcholinesterase and Butyrylcholinesterase of Cat -
Author(s) : Bartels CF , Xie W , Miller-Lindholm AK , Lockridge O
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :147 , 1998
PubMedID:

Title : A Comparison of Blood Cholinesterase Activities, Pyridostigmine Inhibition of Red Cell Acetylcholinesterase, and Butyrylcholinesterase Phenotypes in Gulf War Veterans and Normal Controls -
Author(s) : Gentry MK , Powell S , Bitsko N , Bartels CF , Bartko J , Chung R , Lockridge O , Ribas J , Roy M , Malone J , Doctor BP
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :300 , 1998
PubMedID:

Title : Butyrylcholinesterase-catalysed hydrolysis of aspirin, a negatively charged ester, and aspirin-related neutral esters - Masson_1998_Biochim.Biophys.Acta_1387_41
Author(s) : Masson P , Froment MT , Fortier PL , Visicchio JE , Bartels CF , Lockridge O
Ref : Biochimica & Biophysica Acta , 1387 :41 , 1998
Abstract : Although aspirin (acetylsalicylic acid) is negatively charged, it is hydrolysed by butyrylcholinesterase (BCHE). Catalytic parameters were determined in 100 mM Tris buffer, pH 7.4, in the presence and absence of metal cations. The presence of Ca2+ or Mg2+ (<100 mM) in buffer did not change the Km, but accelerated the rate of hydrolysis of aspirin by wild-type or D70G mutant BCHE by 5-fold. Turnover numbers were of the order of 5000-12000 min-1 for the wild-type enzyme and the D70G and D70K enzymes in 100 mM Tris, pH 7.4, containing 50 mM CaCl2 at 25 degreesC; Km values were 6 mM for wild-type, 16 mM for D70G and 38 mM for D70K. People with 'atypical' BCHE have the D70G mutation. The apparent inhibition seen at high aspirin concentration was not due to inhibition by excess substrate but to spontaneous hydrolysis of aspirin, causing inhibition by salicylate. The wild-type and D70G enzymes were competitively inhibited by salicylic acid; the D70K enzyme showed a complex parabolic inhibition, suggesting multiple binding. The effect of salicylate was substrate-dependent, the D70K mutant being activated by salicylate with butyrylthiocholine as substrate. Km value for wild-type enzyme was lower than for D70 mutants, suggesting that residue 70 located at the rim of the active site gorge was not the major site for the initial encounter aspirin-BCHE complex. On the other hand, the virtual absence of affinity of the W82A mutant for aspirin indicated that W82 was the major residue involved in formation of the Michaelis complex. Molecular modelling of aspirin binding to BCHE indicated perpendicular interactions between the aromatic rings of W82 and aspirin. Kinetic study of BCHE-catalysed hydrolysis of different acetyl esters showed that the rate limiting step was acetylation. The bimolecular rate constants for hydrolysis of aspirin by wild-type, D70G and D70K enzymes were found to be close to 1x106 M-1 min-1. These results support the contention that the electrostatic steering due to the negative electrostatic field of the enzyme plays a role in substrate binding, but plays no role in the catalytic steps, i.e. in the enzyme acetylation.
ESTHER : Masson_1998_Biochim.Biophys.Acta_1387_41
PubMedSearch : Masson_1998_Biochim.Biophys.Acta_1387_41
PubMedID: 9748494

Title : Importance of aspartate-70 in organophosphate inhibition, oxime re-activation and aging of human butyrylcholinesterase - Masson_1997_Biochem.J_325_53
Author(s) : Masson P , Froment MT , Bartels CF , Lockridge O
Ref : Biochemical Journal , 325 :53 , 1997
Abstract : Asp-70 is the defining amino acid in the peripheral anionic site of human butyrylcholinesterase (BCHE), whereas acetylcholinesterase has several additional amino acids, the most important one being Trp-277 (Trp-279 in Torpedo AChE). We studied mutants D70G, D70K and A277W to evaluate the role of Asp-70 and Trp-277 in reactions with organophosphates. We found that Asp-70 was important for binding positively charged echothiophate, but not neutral paraoxon and iso-OMPA. Asp-70 was also important for binding of positively charged pralidoxime (2-PAM) and for activation of re-activation by excess 2-PAM. Excess 2-PAM had an effect similar to substrate activation, suggesting the binding of 2 mol of 2-PAM to wild-type but not to the D70G mutant. A surprising result was that Asp-70 was important for irreversible aging, the D70G mutant having a 3- and 8-fold lower rate of aging for paraoxon-inhibited and di-isopropyl fluorophosphate-inhibited BCHE. Mutants of Asp-70 had the same rate constants for phosphorylation and re-activation by 2-PAM as wild-type. The A277W mutant behaved like wild-type in all assays. Our results predict that people with the atypical (D70G) variant of BCHE will be more sensitive to the toxic effects of echothiophate, but will be equally sensitive to paraoxon and di-isopropyl fluorophosphate. People with the D70G mutation will be resistant to re-activation of their inhibited BCHE by 2-PAM, but this will be offset by the lower rate of irreversible aging of inhibited BCHE, allowing some regeneration by spontaneous hydrolysis.
ESTHER : Masson_1997_Biochem.J_325_53
PubMedSearch : Masson_1997_Biochem.J_325_53
PubMedID: 9224629

Title : Role of aspartate 70 and tryptophan 82 in binding of succinyldithiocholine to human butyrylcholinesterase - Masson_1997_Biochemistry_36_2266
Author(s) : Masson P , Legrand P , Bartels CF , Froment MT , Schopfer LM , Lockridge O
Ref : Biochemistry , 36 :2266 , 1997
Abstract : The atypical variant of human butyrylcholinesterase has Gly in place of Asp 70. Patients with this D70G mutation respond abnormally to the muscle relaxant succinyldicholine, experiencing hours of apnea rather than the intended 3 min. Asp 70 is at the rim of the active site gorge 12 A from the active site Ser 198. An unanswered question in the literature is why the atypical variant has a 10-fold increase in Km for compounds with a single positive charge but a 100-fold increase in Km for compounds with two positive charges. We mutated residues Asp 70, Trp 82, Trp 231, Glu 197, and Tyr 332 and expressed mutant enzymes in mammalian cells. Steady-state kinetic parameters for hydrolysis of butyrylthiocholine, benzoylcholine, succinyldithiocholine, and o-nitrophenyl butyrate were determined. The wild type and the D70G mutant had identical k(cat) values for all substrates. Molecular modeling and molecular dynamics suggested that succinyldicholine could bind in two consecutive orientations in the active site gorge; formation of one complex caused a conformational change in the omega loop involving Asp 70 and Trp 82. We propose the formation of three enzyme-substrate intermediates preceding the acyl-enzyme intermediate; kinetic data support this contention. Substrates with a single positive charge interact with Asp 70 just once, whereas substrates with two positive charges, for example succinyldithiocholine, interact with Asp 70 in two complexes, thus explaining the 10- and 100-fold increases in Km in the D70G mutant.
ESTHER : Masson_1997_Biochemistry_36_2266
PubMedSearch : Masson_1997_Biochemistry_36_2266
PubMedID: 9047329

Title : Aging of di-isopropyl-phosphorylated human butyrylcholinesterase - Masson_1997_Biochem.J_327_601
Author(s) : Masson P , Fortier PL , Albaret C , Froment MT , Bartels CF , Lockridge O
Ref : Biochemical Journal , 327 ( Pt 2) :601 , 1997
Abstract : Organophosphate-inhibited cholinesterases can be reactivated by nucleophilic compounds. Sometimes phosphylated (phosphorylated or phosphonylated) cholinesterases become progressively refractory to reactivation; this can result from different reactions. The most frequent process, termed 'aging', involves the dealkylation of an alkoxy group on the phosphyl moiety through a carbocation mechanism. In attempting to determine the amino acid residues involved in the aging of butyrylcholinesterase (BuChE), the human BuChE gene was mutated at several positions corresponding to residues located at the rim of the active site gorge and in the vicinity of the active site. Mutant enzymes were expressed in Chinese hamster ovary cells. Wild-type BuChE and mutants were inhibited by di-isopropylfluorophosphate at pH 8.0 and 25 degrees C. Di-isopropyl-phosphorylated enzymes were incubated with the nucleophilic oxime 2-pyridine aldoxime methiodide and their reactivatability was determined. Reactivatability was expressed by the first-order rate constant of aging and/or the half-life of aging (t12). The t12 was found to be of the order of 60 min for wild-type BuChE. Mutations on Glu-197 increased t12 60-fold. Mutation W82A increased t12 13-fold. Mutation D70G increased t12 8-fold. Mutations in the vicinity of the active site serine residue had either moderate or no effect on aging; t12 was doubled for F329C and F329A, increased only 4-fold for the double mutant A328G+F329S, and no change was observed for the A328G mutant, indicating that the isopropoxy chain to be dealkylated does not directly interact with Ala-328 and Phe-329. These results were interpreted by molecular modelling of di-isopropylphosphorylated wild-type and mutant enzymes. Molecular dynamics simulations indicated that the isopropyl chain that is lost interacted with Trp-82, suggesting that Trp-82 has a role in stabilizing the carbonium ion that is released in the dealkylation step. This study emphasized the important role of the Glu-197 carboxylate in stabilizing the developing carbocation, and the allosteric control of the dealkylation reaction by Asp-70. Indeed, although Asp-70 does not interact with the phosphoryl moiety, mutation D70G affects the rate of aging. This indirect control was interpreted in terms of change in the conformational state of Trp-82 owing to internal motions of the Omega loop (Cys-65-Cys-92) in the mutant enzyme.
ESTHER : Masson_1997_Biochem.J_327_601
PubMedSearch : Masson_1997_Biochem.J_327_601
PubMedID: 9359435

Title : Asp7O in the peripheral anionic site of human butyrylcholinesterase - Masson_1996_Eur.J.Biochem_235_36
Author(s) : Masson P , Froment MT , Bartels CF , Lockridge O
Ref : European Journal of Biochemistry , 235 :36 , 1996
Abstract : The goal of this work was to determine what amino acids at the mouth of the active-site gorge are important for the function of human butyrylcholinesterase. Mutants D70G, Q119Y, G283D, A277W, A277H and A277W/G283D were expressed in human embryonal kidney cells and the secreted enzymes were assayed by steady-state kinetics. The result was that only one amino acid, D70 was found to be important for function. When D70 was mutated to G, the same mutation as in the naturally occurring atypical butyrylcholinesterase, the affinity for positively charged substrates and positively charged inhibitors decreased 5-30-fold. The D70G mutant had another striking abnormality in that it was virtually devoid of the phenomenon of substrate activation by excess butyrylthiocholine. Thus, though kcat was the same for wild-type and D70G mutant, being 24000 min(-1) at low butyrylthiocholine concentrations (0.01-0.1 mM), it failed to increase for the D70G mutant at 40 mM butyrylthiocholine, whereas it increased threefold for wild type. The D70G mutant was more sensitive to changes in salt concentration, its catalytic rate decreasing more than that of the wild type. The D70G mutant appeared to have a greater surface negative charge than wild type suggesting that the D70G mutant had a conformation different from that of the wild type. That D70 affects the function of butyrylcholinesterase, together with its location at the mouth of the active-site gorge, supports the hypothesis that D70 is a component of the peripheral anionic site of butyrylcholinesterase. Mutants containing aromatic amino acids at the mouth of the gorge had increased binding affinity for propidium and fasciculin, but unaltered function, suggesting that aromatic amino acids are not important to the function of the peripheral anionic site of butyrylcholinesterase.
ESTHER : Masson_1996_Eur.J.Biochem_235_36
PubMedSearch : Masson_1996_Eur.J.Biochem_235_36
PubMedID: 8631355

Title : Characterization of 12 silent alleles of the human butyrylcholinesterase (BCHE) gene - Primo-Parmo_1996_Am.J.Hum.Genet_58_52
Author(s) : Primo-Parmo SL , Bartels CF , Wiersema B , van der Spek AF , Innis JW , La Du BN
Ref : American Journal of Human Genetics , 58 :52 , 1996
Abstract : The silent phenotype of human butyrylcholinesterase (BChE), present in most human populations in frequencies of approximately 1/100,000, is characterized by the complete absence of BChE activity or by activity <10% of the average levels of the usual phenotype. Heterogeneity in this phenotype has been well established at the phenotypic level, but only a few silent BCHE alleles have been characterized at the DNA level. Twelve silent alleles of the human butyrylcholinesterase gene (BCHE) have been identified in 17 apparently unrelated patients who were selected by their increased sensitivity to the muscle relaxant succinylcholine. All of these alleles are characterized by single nucleotide substitutions or deletions leading to distinct changes in the structure of the BChE enzyme molecule. Nine of the nucleotide substitutions result in the replacement of single amino acid residues. Three of these variants, BCHE*33C, BCHE*198G, and BCHE*201T, produce normal amounts of immunoreactive but enzymatically inactive BChE protein in the plasma. The other six amino acid substitutions, encoded by BCHE*37S, BCHE*125F, BCHE*170E, BCHE*471R, and BCHE*518L, seem to cause reduced expression of BChE protein, and their role in determining the silent phenotype was confirmed by expression in cell culture. The other four silent alleles, BCHE*271STOP, BCHE*500STOP, BCHE*FS6, and BCHE*I2E3-8G, encode BChES truncated at their C-terminus because of premature stop codons caused by nucleotide substitutions, a frame shift, or altered splicing. The large number of different silent BCHE alleles found within a relatively small number of patients shows that the heterogeneity of the silent BChE phenotype is high. The characterization of silent BChE variants will be useful in the study of the structure/function relationship for this and other closely related enzymes.
ESTHER : Primo-Parmo_1996_Am.J.Hum.Genet_58_52
PubMedSearch : Primo-Parmo_1996_Am.J.Hum.Genet_58_52
PubMedID: 8554068

Title : Acetylcholinesterase Gene Sequence and Copy Number are Normal in Alzheimer's Disease Patients Treated with the Organophosphate Metrifonate -
Author(s) : Bartels CF , Moriearty PL , Becker RE , Mountjoy CP , Lockridge O
Ref : In Enzyme of the Cholinesterase Family - Proceedings of Fifth International Meeting on Cholinesterases , (Quinn, D.M., Balasubramanian, A.S., Doctor, B.P., Taylor, P., Eds) Plenum Publishing Corp. :479 , 1995
PubMedID:

Title : Peripheral Anionic Site of Wild-Type and Mutant Human Butyrylcholinesterase -
Author(s) : Masson P , Froment MT , Bartels CF , Lockridge O
Ref : In Enzyme of the Cholinesterase Family - Proceedings of Fifth International Meeting on Cholinesterases , (Quinn, D.M., Balasubramanian, A.S., Doctor, B.P., Taylor, P., Eds) Plenum Publishing Corp. :230 , 1995
PubMedID:

Title : Tissue distribution of human acetylcholinesterase and butyrylcholinesterase messenger RNA - Jbilo_1994_Toxicon_32_1445
Author(s) : Jbilo O , Bartels CF , Chatonnet A , Toutant JP , Lockridge O
Ref : Toxicon , 32 :1445 , 1994
Abstract : Cholinesterase inhibitors occur naturally in the calabar bean (eserine), green potatoes (solanine), insect-resistant crab apples, the coca plant (cocaine) and snake venom (fasciculin). There are also synthetic cholinesterase inhibitors, for example man-made insecticides. These inhibitors inactivate acetylcholinesterase and butyrylcholinesterase as well as other targets. From a study of the tissue distribution of acetylcholinesterase and butyrylcholinesterase mRNA by Northern blot analysis, we have found the highest levels of butyrylcholinesterase mRNA in the liver and lungs, tissues known as the principal detoxication sites of the human body. These results indicate that butyrylcholinesterase may be a first line of defense against poisons that are eaten or inhaled.
ESTHER : Jbilo_1994_Toxicon_32_1445
PubMedSearch : Jbilo_1994_Toxicon_32_1445
PubMedID: 7886701

Title : Endogenous butyrylcholinesterase in SV40 transformed cell lines: COS-1, COS-7, MRC-5 SV40, and WI-38 VA13 - Kris_1994_In.Vitro.Cell.Dev.Biol.Animal_10_680
Author(s) : Kris M , Jbilo O , Bartels CF , Masson P , Rhode S , Lockridge O
Ref : In Vitro Cellular & Developmental Biology Animal , 10 :680 , 1994
Abstract : Comparison of proteins expressed by SV40 transformed cell lines and untransformed cell lines is of interest because SV40 transformed cells are immortal, whereas untransformed cells senesce after about 50 doublings. In MRC-5 SV40 cells, only seven proteins have previously been reported to shift from undetectable to detectable after transformation by SV40 virus. We report that butyrylcholinesterase is an 8th protein in this category. Butyrylcholinesterase activity in transformed MRC-5 SV40 cells increased at least 150-fold over its undetectable level in MRC-5 parental cells. Other SV40 transformed cell lines, including COS-1, COS-7, and WI-38 VA13, also expressed endogenous butyrylcholinesterase, whereas the parental, untransformed cell lines, CV-1 and WI-38, had no detectable butyrylcholinesterase activity or mRNA. Infection of CV-1 cells by SV40 virus did not result in expression of butyrylcholinesterase, showing that the butyrylcholinesterase promoter was not activated by the large T antigen of SV40. We conclude that butyrylcholinesterase expression resulted from events related to cell immortalization and did not result from activation by the large T antigen.
ESTHER : Kris_1994_In.Vitro.Cell.Dev.Biol.Animal_10_680
PubMedSearch : Kris_1994_In.Vitro.Cell.Dev.Biol.Animal_10_680
PubMedID: 7842168

Title : Mutation His322Asn in human acetylcholinesterase does not alter electrophoretic and catalytic properties of the erythrocyte enzyme - Masson_1994_Blood_83_3003
Author(s) : Masson P , Froment MT , Sorenson RC , Bartels CF , Lockridge O
Ref : Blood , 83 :3003 , 1994
Abstract : The YT blood group antigen is located on human red blood cell (RBC) acetylcholinesterase. Wild-type acetylcholinesterase, YT1, has histidine at codon 322, whereas the genetic variant of acetylcholinesterase, YT2, has asparagine. This mutation is located within exon 2 of the ACHE gene, an exon that is present in all alternatively spliced forms of acetylcholinesterase. Therefore, acetylcholinesterase in brain and muscle has the same mutation as RBC acetylcholinesterase. We compared the electrophoretic and kinetic properties of RBC acetylcholinesterases having His 322 or Asn 322. We found no differences in the isoelectric point, mobility on non-denaturing gel electrophoresis, affinity for acetylthiocholine, activity per milligram of RBC ghost protein, substrate inhibition constants, binding to the peripheral site ligand (propidium), and binding to active site ligands (tetrahydroaminoacridine and edrophonium). Thus, although the point mutation elicits antibody production in nonmatching blood transfusion recipients, it has no effect on the enzymatic properties of acetylcholinesterase.
ESTHER : Masson_1994_Blood_83_3003
PubMedSearch : Masson_1994_Blood_83_3003
PubMedID: 8180397

Title : Mutation at codon 322 in the human acetylcholinesterase (ACHE) gene accounts for YT blood group polymorphism - Bartels_1993_Am.J.Hum.Genet_52_928
Author(s) : Bartels CF , Zelinski T , Lockridge O
Ref : American Journal of Human Genetics , 52 :928 , 1993
Abstract : Acetylcholinesterase is present in innervated tissues, where its function is to terminate nerve impulse transmission. It is also found in the red blood cell membrane, where its function is unknown. We report the first genetic variant of human acetylcholinesterase and support the identity of acetylcholinesterase as the YT blood group antigen. DNA sequencing shows that the wild-type sequence of acetylcholinesterase with His322 (CAC) is the YT1 blood group antigen and that the rare variant of acetylcholinesterase with Asn322 (AAC) is the YT2 blood group antigen. Two additional point mutations in the acetylcholinesterase gene do not affect the amino acid sequence of the mature enzyme.
ESTHER : Bartels_1993_Am.J.Hum.Genet_52_928
PubMedSearch : Bartels_1993_Am.J.Hum.Genet_52_928
PubMedID: 8488842

Title : DNA mutations associated with the human butyrylcholinesterase J-variant - Bartels_1992_Am.J.Hum.Genet_50_1104
Author(s) : Bartels CF , James K , La Du BN
Ref : American Journal of Human Genetics , 50 :1104 , 1992
Abstract : The J-variant of human serum butyrylcholinesterase (BChE) causes both an approximately two-thirds reduction of circulating enzyme molecules and a corresponding decrease in the level of BChE activity present in serum. Since the level of serum BChE activity and the duration of succinylcholine apnea are inversely correlated, this marked decrease in activity makes individuals with the J-variant more susceptible than usual subjects to prolonged apnea from succinylcholine. We reinvestigated the same family in which Garry et al. identified the J-variant phenotype. The atypical, fluoride, and K-variant mutations were also identified in members of the 47-person pedigree. DNA amplification by PCR, followed by direct sequencing of the amplified DNA, led to the finding that the J-variant phenotype of human serum BChE was associated with two DNA point mutations in the coding region. One of these was the mutation previously identified with the K-variant phenotype (GCA----ACA; Ala539----Thr). The other was an adenine-to-thymine transversion at nucleotide 1490, which changed amino acid 497 from glutamic acid to valine (GAA----GTA; Glu497----Val). This latter point mutation was named the J-variant mutation (formal name BCHE*497V). The J-variant mutation has not been identified without the K-variant mutation. The J-variant mutation created an RsaI-enzyme RFLP. Two additional point mutations, located in the noncoding regions of the gene, were also found to be linked with the J-variant and K-variant point mutations on the same allele. These noncoding polymorphic mutations had previously been found linked to the atypical and K-variant point mutations. A summary table shows dibucaine, fluoride, and Hoffmann-La Roche compound Ro 2-0683 inhibition numbers for 119 samples whose DNA has been sequenced. Eighteen BChE genotypes are represented.
ESTHER : Bartels_1992_Am.J.Hum.Genet_50_1104
PubMedSearch : Bartels_1992_Am.J.Hum.Genet_50_1104
PubMedID: 1349196

Title : DNA mutation associated with the human butyrylcholinesterase K-variant and its linkage to the atypical variant mutation and other polymorphic sites - Bartels_1992_Am.J.Hum.Genet_50_1086
Author(s) : Bartels CF , Jensen FS , Lockridge O , van der Spek AF , Rubinstein HM , Lubrano T , La Du BN
Ref : American Journal of Human Genetics , 50 :1086 , 1992
Abstract : Genomic DNA from two families exhibiting the K-variant phenotype of serum butyrylcholinesterase was amplified by PCR and sequenced to determine the molecular basis of this variant. The K-variant phenotype was found to be associated with a DNA transition from guanine to adenine at nucleotide 1615, which caused an amino acid change from alanine 539 to threonine (GCA----ACA; Ala539----Thr). There was a 30% reduction of serum butyrylcholinesterase activity associated with this mutation. Amplification and sequencing of DNA from a random sample of 47 unrelated people gave a frequency of .128 for the K-variant allele. Thus, 1 person in 63 should be homozygous for the K-variant, making the K-variant the most common butyrylcholinesterase variant. The K-variant mutation was also found to be present in 17 (89%) of 19 butyrylcholinesterase genes containing the point mutation which causes the atypical phenotype of butyrylcholinesterase (GAT----GGT; Asp70----Gly). The presence of the K-variant in the same molecule as the atypical variant does not contribute to the qualitative change in the atypical enzyme, but it most likely accounts for the approximately one-third reduction in Vmax of butyrylcholinesterase activity in atypical serum. Two additional point mutations located in noncoding regions of the gene were also observed to be in linkage disequilibrium with the K-variant mutation. As many as four different point mutations have been identified within a single butyrylcholinesterase gene. Inhibition tests of the enzyme in plasma are usually used to distinguish the K-variant from the usual enzyme when the former is present with the heterozygous atypical variant (AK phenotype vs. UA phenotype). Inhibition tests were performed on plasma enzyme from the four possible genotypic combinations of the heterozygous atypical mutation with or without the K-variant mutation on either allele; we found that the AK phenotype was caused by three genotypes (A/K, AK/K, and U/A) and that the UA phenotype was caused by two genotypes (U/A and U/AK).
ESTHER : Bartels_1992_Am.J.Hum.Genet_50_1086
PubMedSearch : Bartels_1992_Am.J.Hum.Genet_50_1086
PubMedID: 1570838

Title : Structural basis of the butyrylcholinesterase H-variant segregating in two Danish families - Jensen_1992_Pharmacogenet_2_234
Author(s) : Jensen FS , Bartels CF , La Du BN
Ref : Pharmacogenetics , 2 :234 , 1992
Abstract : The rare H-variant of human butyrylcholinesterase is a quantitative variant that reduces serum butyrylcholinesterase activity by about 90%. Individuals who are heterozygous for both the H-variant and the atypical variant are abnormally sensitive to the muscle relaxant succinylcholine. By using standard phenotypic serum assays, the Danish Cholinesterase Research Unit identified four individuals from two unrelated pedigrees who were heterozygous for both the H-variant (H) and the atypical (A) variant. DNA of these A/H individuals was extracted from white blood cells. Using the polymerase chain reaction and subsequent DNA sequencing, a point mutation was found at nucleotide 424 which changed amino acid 142 from valine to methionine. The previously identified atypical mutation, Asp 70 to Gly, was also seen, which segregated apart from the H-variant mutation in family studies. These two mutations were found in all four A/H individuals.
ESTHER : Jensen_1992_Pharmacogenet_2_234
PubMedSearch : Jensen_1992_Pharmacogenet_2_234
PubMedID: 1306123

Title : Identification of two different point mutations associated with the fluoride-resistant phenotype for human butyrylcholinesterase - Nogueira_1992_Am.J.Hum.Genet_51_821
Author(s) : Nogueira CP , Bartels CF , McGuire MC , Adkins S , Lubrano T , Rubinstein HM , Lightstone H , van der Spek AF , Lockridge O , La Du BN
Ref : American Journal of Human Genetics , 51 :821 , 1992
Abstract : The fluoride variant of human butyrylcholinesterase owes its name to the observation that it is resistant to inhibition by 0.050 mM sodium fluoride in the in vitro assay. Individuals who are heterozygous for the fluoride and atypical alleles experience about 30 min of apnea, rather than the usual 3-5 min, after receiving succinyldicholine. Earlier we reported that the atypical variant has a nucleotide substitution which changes Asp 70 to Gly. In the present work we have identified two different point mutations associated with the fluoride-resistant phenotype. Fluoride-1 has a nucleotide substitution which changes Thr 243 to Met (ACG to ATG). Fluoride-2 has a substitution which changes Gly 390 to Val (GGT to GTT). These results were obtained by DNA sequence analysis of the butyrylcholinesterase gene after amplification by PCR. The subjects for these analyses were 4 patients and 21 family members.
ESTHER : Nogueira_1992_Am.J.Hum.Genet_51_821
PubMedSearch : Nogueira_1992_Am.J.Hum.Genet_51_821
PubMedID: 1415224

Title : Part 1: Genetic Variant of Human Acetylcholinesterase Part 2: SV-40 Transformed Cell Lines, for Example COS-1, but Not Parental Untransformed Cell Lines, Express Butyrylcholinesterase (BChE) -
Author(s) : Lockridge O , Bartels CF
Ref : In Multidisciplinary approaches to cholinesterase functions - Proceedings of Fourth International Meeting on Cholinesterases , (Shafferman, A. and Velan, B., Eds) Plenum Press, New York :53 , 1992
PubMedID:

Title : Heterogeneity of the Silent Phenotype of Human Butyrylcholinesterase - Identification of Eight New Mutations -
Author(s) : Primo-Parmo SL , Bartels CF
Ref : In Multidisciplinary approaches to cholinesterase functions - Proceedings of Fourth International Meeting on Cholinesterases , (Shafferman, A. and Velan, B., Eds) Plenum Press, New York :61 , 1992
PubMedID:

Title : Poster: Expression of the fluoride variant of human butyrylcholinesterase in chinese hamster ovary cells -
Author(s) : Adkins S , Vaughan TA , Bartels CF , La Du BN , Lockridge O
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :196 , 1991
PubMedID:

Title : Poster: Nomenclature for human butyrylcholinesterase genetic variants identified by DNA sequencing -
Author(s) : Lockridge O , Bartels CF , Nogueira CP , Arpagaus M , Adkins S , La Du BN
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :193 , 1991
PubMedID:

Title : Poster: Practical consequences of having more than one mutation within the same butyrylcholinesterase gene -
Author(s) : La Du BN , Bartels CF , Lockridge O
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :192 , 1991
PubMedID:

Title : Poster: DNA Coding for the K polymorphism in linkage disequilibrium with atypical human butyrylcholinesterase complicates phenotyping -
Author(s) : Bartels CF , Lockridge O , La Du BN , van der Spek AF , Rubinstein HM , Lubrano T
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :191 , 1991
PubMedID:

Title : Poster: Identification of two different mutations associated with human butyrylcholinesterase fluoride resistance in serum -
Author(s) : Bartels CF , Nogueira CP , McGuire MC , Adkins S , Lockridge O , La Du BN , Rubinstein HM , Lubrano T , van der Spek AF , Lightstone H
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :190 , 1991
PubMedID:

Title : Poster: A DNA point mutation associated with the H-variant of human butyrylcholinesterase -
Author(s) : Jensen FS , Bartels CF , La Du BN
Ref : In: Cholinesterases: Structure, Function, Mechanism, Genetics, and Cell Biology , (Massoulie J, Barnard EA, Chatonnet A, Bacou F, Doctor BP, Quinn DM) American Chemical Society, Washington, DC :189 , 1991
PubMedID:

Title : Proposed nomenclature for human butyrylcholinesterase genetic variants identified by DNA sequencing - La Du_1991_Cell.Mol.Neurobiol_11_79
Author(s) : La Du BN , Bartels CF , Nogueira CP , Arpagaus M , Lockridge O
Ref : Cellular Molecular Neurobiology , 11 :79 , 1991
Abstract : 1. New information identifying nucleotide alterations of human butyrylcholinesterase allows the use of more specific nomenclature for the variants commonly known as atypical, fluoride, silent, and K variant. 2. In addition to suggesting a system of trivial names and abbreviations, we provide a list of formal names that follow the guidelines of the Committee for Human Gene Nomenclature. 3. It is suggested that formal names be included in publications whenever possible.
ESTHER : La Du_1991_Cell.Mol.Neurobiol_11_79
PubMedSearch : La Du_1991_Cell.Mol.Neurobiol_11_79
PubMedID: 2013061

Title : Use of the polymerase chain reaction for homology probing of butyrylcholinesterase from several vertebrates - Arpagaus_1991_J.Biol.Chem_266_6966
Author(s) : Arpagaus M , Chatonnet A , Masson P , Newton M , Vaughan TA , Bartels CF , Nogueira CP , La Du BN , Lockridge O
Ref : Journal of Biological Chemistry , 266 :6966 , 1991
Abstract : Genomic blots from man, monkey, cow, sheep, pig, rabbit, dog, rat, mouse, guinea pig, and chicken DNA were hybridized with probes derived from the four exons of the human butyrylcholinesterase gene (BCHE) (Arpagaus, M., Kott, M., Vatsis, K. P., Bartels, C. F., La Du, B. N., and Lockridge, O. (1990) Biochemistry 29, 124-131). Results showed that the BCHE gene was present in a single copy in the genome of all these vertebrates. The polymerase chain reaction was used to amplify genomic DNA from these animals with oligonucleotides derived from the human BCHE coding sequence. The amplified segment contained 423 bp of BCHE sequence including the active site serine of the enzyme (amino acid 198) and a component of the anionic site, aspartate 70. Amplification was successful for monkey, pig, cow, dog, sheep, and rabbit DNA, but unsuccessful for rat, guinea pig, mouse, and chicken DNA. Amplified segments were cloned in M13 and sequenced. The mouse sequence was obtained by sequencing a genomic clone. The highest identity of the human amino acid sequence was found with monkey (100%) and the lowest with mouse (91.5%). The sequence around the active site serine 198, Phe-Gly-Glu-Ser-Ala-Gly-Ala, was conserved in all eight animals as was the anionic site component, aspartate 70. A phylogenetic tree of mammalian butyrylcholinesterases was constructed using the partial BCHE sequences.
ESTHER : Arpagaus_1991_J.Biol.Chem_266_6966
PubMedSearch : Arpagaus_1991_J.Biol.Chem_266_6966
PubMedID: 2016308
Gene_locus related to this paper: bovin-BCHE , canfa-BCHE , macmu-BCHE , pig-BCHE , sheep-BCHE

Title : Structure of the gene for human butyrylcholinesterase. Evidence for a single copy - Arpagaus_1990_Biochemistry_29_124
Author(s) : Arpagaus M , Kott M , Vatsis KP , Bartels CF , La Du BN , Lockridge O
Ref : Biochemistry , 29 :124 , 1990
Abstract : We have isolated five genomic clones for human butyrylcholinesterase (BChE), using cDNA probes encoding the catalytic subunit of the hydrophilic tetramer [McTiernan et al. (1987) Proc. Natl. Acad. Sci. U.S.A. 84, 6682-6686]. The BChE gene is at least 73 kb long and contains four exons. Exon 1 contains untranslated sequences and two potential translation initiation sites at codons -69 and -47. Exon 2 (1525 bp) contains 83% of the coding sequence for the mature protein, including the N-terminal and the active-site serine, and a third possible translation initiation site (likely functional), at codon -28. Exon 3 is 167 nucleotides long. Exon 4 (604 bp) codes for the C-terminus of the protein and the 3' untranslated region where two polyadenylation signals were identified. Intron 1 is 6.5 kb long, and the minimal sizes of introns 2 and 3 are estimated to be 32 kb each. Southern blot analysis of total human genomic DNA is in complete agreement with the gene structure established by restriction endonuclease mapping of the genomic clones: this strongly suggests that the BChE gene is present in a single copy.
ESTHER : Arpagaus_1990_Biochemistry_29_124
PubMedSearch : Arpagaus_1990_Biochemistry_29_124
PubMedID: 2322535

Title : Identification of a frameshift mutation responsible for the silent phenotype of human serum cholinesterase, Gly 117 (GGT----GGAG) - Nogueira_1990_Am.J.Hum.Genet_46_934
Author(s) : Nogueira CP , McGuire MC , Graeser C , Bartels CF , Arpagaus M , van der Spek AF , Lightstone H , Lockridge O , La Du BN
Ref : American Journal of Human Genetics , 46 :934 , 1990
Abstract : A frameshift mutation that causes a silent phenotype for human serum cholinesterase was identified in the DNA of seven individuals of two unrelated families. The mutation, identified using the polymerase chain reaction, causes a shift in the reading frame from Gly 117, where GGT (Gly)----GGAG (Gly+ 1 base) to a new stop codon created at position 129. This alteration is upstream of the active site (Ser 198), and, if any protein were made, it would represent only 22% of the mature enzyme found in normal serum. Results of analysis of the enzymatic activities in serum agreed with the genotypes inferred from the nucleotide sequence. Rocket immunoelectrophoresis using alpha-naphthyl acetate to detect enzymatic activity showed an absence of cross-reactive material, as expected. One additional individual with a silent phenotype did not show the same frameshift mutation. This was not unexpected, since there must be considerable molecular heterogeneity involved in causes for the silent cholinesterase phenotype. This is the first report of a molecular mechanism underlying the silent phenotype for serum cholinesterase. The analytical approach used was similar to the one we recently employed to identify the mutation that causes the atypical cholinesterase variant.
ESTHER : Nogueira_1990_Am.J.Hum.Genet_46_934
PubMedSearch : Nogueira_1990_Am.J.Hum.Genet_46_934
PubMedID: 2339692

Title : Phenotypic and molecular biological analysis of human butyrylcholinesterase variants - La Du_1990_Clin.Biochem_23_423
Author(s) : La Du BN , Bartels CF , Nogueira CP , Hajra A , Lightstone H , van der Spek AF , Lockridge O
Ref : Clinical Biochemistry , 23 :423 , 1990
Abstract : Our laboratory has recently shown that several variant forms of human butyrylcholinesterase, associated with unusual sensitivity to succinylcholine, are caused by specific mutations within the structural DNA coding for this enzyme. Atypical (dibucaine-resistant) butyrylcholinesterase is caused by a point mutation at nucleotide position 209(GAT-- greater than GGT), which changes aspartate 70 to glycine. One fluoride-resistant variant family has a point mutation at nucleotide 728(ACG-- greater than ATG), which changes threonine 243 to methionine. Another type of fluoride-resistant variant has a point mutation at nucleotide 1169(GGT-- greater than GTT), which changes glycine 390 to valine. One type of silent phenotype is due to a frame-shift mutation at nucleotide position 351(GGT-- greater than GGAG). A polymorphic site at nucleotide position 1615 (GCA/ACA), coding for Ala/Thr, accounts for the quantitative K-variant, which causes an approximate one-third reduction of activity, if Thr occupies that position at codon 539. Examples are given to illustrate the advantages of using a combination of the new DNA analytical techniques, including: the use of allele-specific probes, with the standard serum cholinesterase phenotyping methods. More accurate typing of patients with certain variants is now possible; pedigree analysis will be aided by the improved methodology.
ESTHER : La Du_1990_Clin.Biochem_23_423
PubMedSearch : La Du_1990_Clin.Biochem_23_423
PubMedID: 2253336

Title : Two polymorphisms in the non-coding regions of the BCHE gene -
Author(s) : Bartels CF , van der Spek AF , La Du BN
Ref : Nucleic Acids Research , 18 :6171 , 1990
PubMedID: 1978284

Title : Identification of a frameshift mutation (gly 117, GGT-to-GGAG) responsible for a silent phenotype of human serum cholinesterase. (Abstract) -
Author(s) : Nogueira CP , McGuire MC , Bartels CF , van der Spek AF , Lightstone H , Lockridge O , La Du BN
Ref : American Journal of Human Genetics , 45 (suppl.) :A210 , 1989
PubMedID:

Title : Identification of the structural mutation responsible for the dibucaine-resistant (atypical) variant form of human serum cholinesterase - McGuire_1989_Proc.Natl.Acad.Sci.U.S.A_86_953
Author(s) : McGuire MC , Nogueira CP , Bartels CF , Lightstone H , Hajra A , van der Spek AF , Lockridge O , La Du BN
Ref : Proc Natl Acad Sci U S A , 86 :953 , 1989
Abstract : A point mutation in the gene for human serum cholinesterase was identified that changes Asp-70 to Gly in the atypical form of serum cholinesterase. The mutation in nucleotide 209, which changes codon 70 from GAT to GGT, was found by sequencing a genomic clone and sequencing selected regions of DNA amplified by the polymerase chain reaction. The entire coding sequences for usual and atypical cholinesterases were compared, and no other consistent base differences were found. A polymorphic site near the C terminus of the coded region was detected, but neither allele at this locus segregated consistently with the atypical trait. The nucleotide-209 mutation was detected in all five atypical cholinesterase families examined. There was complete concordance between this mutation and serum cholinesterase phenotypes for all 14 heterozygous and 6 homozygous atypical subjects tested. The mutation causes the loss of a Sau3A1 restriction site; the resulting DNA fragment length polymorphism was verified by electrophoresis of 32P-labeled DNA restriction fragments from usual and atypical subjects. Dot-blot hybridization analysis with a 19-mer allele-specific probe to the DNA amplified by the polymerase chain reaction distinguished between the usual and atypical genotypes. We conclude that the Asp-70----Gly mutation (acidic to neutral amino acid substitution) accounts for reduced affinity of atypical cholinesterase for choline esters and that Asp-70 must be an important component of the anionic site. Heterogeneity in atypical alleles may exist, but the Asp-70 point mutation may represent an appreciable portion of the atypical gene pool.
ESTHER : McGuire_1989_Proc.Natl.Acad.Sci.U.S.A_86_953
PubMedSearch : McGuire_1989_Proc.Natl.Acad.Sci.U.S.A_86_953
PubMedID: 2915989

Title : Brain cDNA clone for human cholinesterase - McTiernan_1987_Proc.Natl.Acad.Sci.U.S.A_84_6682
Author(s) : McTiernan C , Adkins S , Chatonnet A , Vaughan TA , Bartels CF , Kott M , Rosenberry TL , La Du BN , Lockridge O
Ref : Proceedings of the National Academy of Sciences of the United States of America , 84 :6682 , 1987
Abstract : A cDNA library from human basal ganglia was screened with oligonucleotide probes corresponding to portions of the amino acid sequence of human serum cholinesterase (EC 3.1.1.8). Five overlapping clones, representing 2.4 kilobases, were isolated. The sequenced cDNA contained 207 base pairs of coding sequence 5' to the amino terminus of the mature protein in which there were four ATG translation start sites in the same reading frame as the protein. Only the ATG coding for Met-(-28) lay within a favorable consensus sequence for functional initiators. There were 1722 base pairs of coding sequence corresponding to the protein found circulating in human serum. The amino acid sequence deduced from the cDNA exactly matched the 574 amino acid sequence of human serum cholinesterase, as previously determined by Edman degradation. Therefore, our clones represented cholinesterase (EC 3.1.1.8) rather than acetylcholinesterase (EC 3.1.1.7). It was concluded that the amino acid sequences of cholinesterase from two different tissues, human brain and human serum, were identical. Hybridization of genomic DNA blots suggested that a single gene, or very few genes, coded for cholinesterase.
ESTHER : McTiernan_1987_Proc.Natl.Acad.Sci.U.S.A_84_6682
PubMedSearch : McTiernan_1987_Proc.Natl.Acad.Sci.U.S.A_84_6682
PubMedID: 3477799
Gene_locus related to this paper: human-BCHE

Title : Complete amino acid sequence of human serum cholinesterase - Lockridge_1987_J.Biol.Chem_262_549
Author(s) : Lockridge O , Bartels CF , Vaughan TA , Wong CK , Norton SE , Johnson LL
Ref : Journal of Biological Chemistry , 262 :549 , 1987
Abstract : 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.
ESTHER : Lockridge_1987_J.Biol.Chem_262_549
PubMedSearch : Lockridge_1987_J.Biol.Chem_262_549
PubMedID: 3542989
Gene_locus related to this paper: human-BCHE