Quinn DM


Full name : Quinn Daniel M

First name : Daniel M

Mail : Department of Chemistry\; The University of Iowa\; 327 CB\; Iowa City\; IA 52242

Zip Code :

City :

Country : USA

Email : daniel-quinn@uiowa.edu

Phone : +13193351335

Fax : 319-335-1270

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References (44)

Title : Resurrection Biology: Aged Acetylcholinesterase Brought Back to Life - Quinn_2018_J.Med.Chem_61_7032
Author(s) : Quinn DM
Ref : Journal of Medicinal Chemistry , 61 :7032 , 2018
Abstract : Organophosphorus agents such as sarin and soman that phosphylate the active site serine of the enzyme acetylcholinesterase are notorious and pernicious, not only because they have been used by tyrants to effect mass murder of their own populations but also because they are sought by terrorists to inflict mass casualties on civilian populations. These threats underscore the need to develop effective antidotes against such agents. Phosphylation of acetylcholinesterase produces two adducts, an initial neutral adduct that can be reactivated with oxime nucleophiles, and a subsequent monoanionic adduct (called aged acetylcholinesterase) which has proven over two generations to be impervious to reactivation. This Viewpoint discusses a recent article in the journal that describes the first successful efforts to resurrect the activity of aged acetylcholinesterase.
ESTHER : Quinn_2018_J.Med.Chem_61_7032
PubMedSearch : Quinn_2018_J.Med.Chem_61_7032
PubMedID: 30110162

Title : Why is Aged Acetylcholinesterase So Difficult to Reactivate? - Quinn_2017_Molecules_22_
Author(s) : Quinn DM , Topczewski J , Yasapala N , Lodge A
Ref : Molecules , 22 : , 2017
Abstract : Organophosphorus agents are potent inhibitors of acetylcholinesterase. Inhibition involves successive chemical events. The first is phosphylation of the active site serine to produce a neutral adduct, which is a close structural analog of the acylation transition state. This adduct is unreactive toward spontaneous hydrolysis, but in many cases can be reactivated by nucleophilic medicinal agents, such as oximes. However, the initial phosphylation reaction may be followed by a dealkylation reaction of the incipient adduct. This reaction is called aging and produces an anionic phosphyl adduct with acetylcholinesterase that is refractory to reactivation. This review considers why the anionic aged adduct is unreactive toward nucleophiles. An alternate approach is to realkylate the aged adduct, which would render the adduct reactivatable with oxime nucleophiles. However, this approach confronts a considerable-and perhaps intractable-challenge: the aged adduct is a close analog of the deacylation transition state. Consequently, the evolutionary mechanisms that have led to transition state stabilization in acetylcholinesterase catalysis are discussed herein, as are the challenges that they present to reactivation of aged acetylcholinesterase.
ESTHER : Quinn_2017_Molecules_22_
PubMedSearch : Quinn_2017_Molecules_22_
PubMedID: 28869561

Title : Development of quantitative structure activity relationships for the binding affinity of methoxypyridinium cations for human acetylcholinesterase - Morrill_2015_J.Mol.Graph.Model_62_181
Author(s) : Morrill JA , Topczewski JJ , Lodge AM , Yasapala N , Quinn DM
Ref : J Mol Graph Model , 62 :181 , 2015
Abstract : Among the most toxic substances known are the organophosphorus (OP) compounds used as pesticides and chemical warfare agents. Owing to their high toxicity there is a number of efforts underway to develop effective therapies for OP agent exposure. To date all therapies in use treat inhibited acetylcholinesterase (AChE), but are ineffective for the treatment of inhibited AChE, which has undergone a subsequent hydrolysis process, referred to as aging. Toward developing a therapy for treating victims of OP intoxication in the aged state we have developed Quantitative Structure-Activity Relationships (QSARs) based on the AM1 semiempirical quantum mechanical method using the program, CODESSA (COmprehensive Descriptors for Structural and Statistical Analysis). Using this methodology we obtained a multiple correlation QSAR equation which gave R2=0.9359 for a random training set of 38 ligands and R2=0.9236 for prediction on a random test set of 9 ligands.
ESTHER : Morrill_2015_J.Mol.Graph.Model_62_181
PubMedSearch : Morrill_2015_J.Mol.Graph.Model_62_181
PubMedID: 26454505

Title : Reversible inhibition of human acetylcholinesterase by methoxypyridinium species - Topczewski_2013_Bioorg.Med.Chem.Lett_23_5786
Author(s) : Topczewski JJ , Lodge AM , Yasapala SN , Payne MK , Keshavarzi PM , Quinn DM
Ref : Bioorganic & Medicinal Chemistry Lett , 23 :5786 , 2013
Abstract : The irreversible inhibition of acetylcholinesterase (AChE) by organophosphorous chemical warfare agents necessitates that antidotes be administered for effective treatment. Currently no antidote is known that resurrects the phosphyl-AChE complex once aging has occurred. This report characterizes the affinities of over 30 new AChE inhibitors which could act as resurrecting agents for the aged AChE-OP adduct.
ESTHER : Topczewski_2013_Bioorg.Med.Chem.Lett_23_5786
PubMedSearch : Topczewski_2013_Bioorg.Med.Chem.Lett_23_5786
PubMedID: 24076173

Title : Kinetic assessment of N-methyl-2-methoxypyridinium species as phosphonate anion methylating agents - Topczewski_2013_Org.Lett_15_1084
Author(s) : Topczewski JJ , Quinn DM
Ref : Org Lett , 15 :1084 , 2013
Abstract : Organophosphate nerve agents and pesticides are potent inhibitors of acetylcholinesterase (AChE). Although oxime nucleophiles can reactivate the AChE-phosphyl adduct, the adduct undergoes a reaction called aging. No compounds have been described that reactivate the aged-AChE adduct. A family of 2-methoxypyridinium species which reverse aging in a model system is presented. A kinetic study of this system, which includes an SAR analysis, demonstrates that the reaction is highly tunable based on the ring substituents.
ESTHER : Topczewski_2013_Org.Lett_15_1084
PubMedSearch : Topczewski_2013_Org.Lett_15_1084
PubMedID: 23410111

Title : Accumulation of tetrahedral intermediates in cholinesterase catalysis: a secondary isotope effect study - Tormos_2010_J.Am.Chem.Soc_132_17751
Author(s) : Tormos JR , Wiley KL , Wang Y , Fournier D , Masson P , Nachon F , Quinn DM
Ref : Journal of the American Chemical Society , 132 :17751 , 2010
Abstract : In a previous communication, kinetic beta-deuterium secondary isotope effects were reported that support a mechanism for substrate-activated turnover of acetylthiocholine by human butyrylcholinesterase (BuChE) wherein the accumulating reactant state is a tetrahedral intermediate ( Tormos , J. R. ; et al. J. Am. Chem. Soc. 2005 , 127 , 14538 - 14539 ). In this contribution additional isotope effect experiments are described with acetyl-labeled acetylthiocholines (CL(3)COSCH(2)CH(2)N(+)Me(3); L = H or D) that also support accumulation of the tetrahedral intermediate in Drosophila melanogaster acetylcholinesterase (DmAChE) catalysis. In contrast to the aforementioned BuChE-catalyzed reaction, for this reaction the dependence of initial rates on substrate concentration is marked by pronounced substrate inhibition at high substrate concentrations. Moreover, kinetic beta-deuterium secondary isotope effects for turnover of acetylthiocholine depended on substrate concentration, and gave the following: (D3)k(cat)/K(m) = 0.95 +/- 0.03, (D3)k(cat) = 1.12 +/- 0.02 and (D3)betak(cat) = 0.97 +/- 0.04. The inverse isotope effect on k(cat)/K(m) is consistent with conversion of the sp(2)-hybridized substrate carbonyl in the E + A reactant state into a quasi-tetrahedral transition state in the acylation stage of catalysis, whereas the markedly normal isotope effect on k(cat) is consistent with hybridization change from sp(3) toward sp(2) as the reactant state for deacylation is converted into the subsequent transition state. Transition states for Drosophila melanogaster AChE-catalyzed hydrolysis of acetylthiocholine were further characterized by measuring solvent isotope effects and determining proton inventories. These experiments indicated that the transition state for rate-determining decomposition of the tetrahedral intermediate is stabilized by multiple protonic interactions. Finally, a simple model is proposed for the contribution that tetrahedral intermediate stabilization provides to the catalytic power of acetylcholinesterase.
ESTHER : Tormos_2010_J.Am.Chem.Soc_132_17751
PubMedSearch : Tormos_2010_J.Am.Chem.Soc_132_17751
PubMedID: 21105647

Title : A Secondary Isotope Effect Study of Equine Serum Butyrylcholinesterase- Catalyzed Hydrolysis of Acetylthiocholine - Wiley_2010_Chem.Biol.Interact_187_124
Author(s) : Wiley KL , Tormos JR , Quinn DM
Ref : Chemico-Biological Interactions , 187 :124 , 2010
Abstract : beta-Secondary deuterium isotope effects have been measured for equine serum butyrylcholinesterase-catalyzed hydrolysis of acetyl-L(3)-thiocholine (L=H or (2)H). The dependencies of initial rates on isotopic substrate concentrations show close adherence to Michaelis-Menten kinetics, and yield the following isotope effects: (D3)k(cat)/K(m)=0.98+/-0.02 and (D3)k(cat)=1.10+/-0.02. The modestly inverse isotope effect on k(cat)/K(m) is consistent with partial rate limitation by a step that converts the sp(2)-hybridized ester carbonyl of the E+A reactant state into a quasi-tetrahedral transition state in the acylation stage of catalysis. On the other hand, the markedly normal isotope effect on k(cat) indicates that the Michaelis complex that accumulates at substrate saturation of the active site during catalytic turnover is a tetrahedral intermediate, whose decomposition is the rate-limiting step. These results compliment a previous report [J.R. Tormos et al., J. Am. Chem. Soc. 127 (2005) 14538-14539] that showed that substrate-activated hydrolysis of acetylthiocholine (ATCh), catalyzed by recombinant human butyrylcholinesterase, is also rate limited by decomposition of an accumulating tetrahedral intermediate.
ESTHER : Wiley_2010_Chem.Biol.Interact_187_124
PubMedSearch : Wiley_2010_Chem.Biol.Interact_187_124
PubMedID: 20493178

Title : The reactant state for substrate-activated turnover of acetylthiocholine by butyrylcholinesterase is a tetrahedral intermediate - Tormos_2005_J.Am.Chem.Soc_127_14538
Author(s) : Tormos JR , Wiley KL , Seravalli J , Nachon F , Masson P , Nicolet Y , Quinn DM
Ref : Journal of the American Chemical Society , 127 :14538 , 2005
Abstract : Secondary beta-deuterium kinetic isotope effects have been measured as a function of substrate concentration for recombinant human butyrylcholinesterase-catalyzed hydrolysis of acetyl-L3-thiocholine (L = 1H or 2H). The isotope effect on V/K is inverse, D3V/K = 0.93 +/- 0.03, which is consistent with conversion of the sp2 hybridized carbonyl carbon of the scissile ester bond of the E + A reactant state to a quasi-tetrahedral structure in the acylation transition state. In contrast, the isotope effect on Vmax under conditions of substrate activation is markedly normal, D3(betaVmax) = 1.29 +/- 0.06, an observation that is consistent with accumulation of a tetrahedral intermediate as the reactant state for catalytic turnover. Generally, tetrahedral intermediates for nonenzymatic ester hydrolyses are high-energy steady-state intermediates. Apparently, butyrylcholinesterase displays an unusual ability to stabilize such intermediates. Hence, the catalytic power of cholinesterases can largely be understood in terms of their ability to stabilize tetrahedral intermediates in the multistep reaction mechanism.
ESTHER : Tormos_2005_J.Am.Chem.Soc_127_14538
PubMedSearch : Tormos_2005_J.Am.Chem.Soc_127_14538
PubMedID: 16231883

Title : Poster (15) Molecular recognition in AChE catalysis - the view from near and far. -
Author(s) : Moussavi-Harami F , Sikorski RS , Quinn DM , Feaster SR , Doctor BP
Ref : In: Cholinesterases in the Second Millennium: Biomolecular and Pathological Aspects , (Inestrosa NC, Campos EO) P. Universidad Catolica de Chile-FONDAP Biomedicina :328 , 2004

Title : Importance of arginines 63 and 423 in modulating the bile salt-dependent and bile salt-independent hydrolytic activities of rat carboxyl ester lipase - Liang_2000_J.Biol.Chem_275_24040
Author(s) : Liang Y , Medhekar R , Brockman HL , Quinn DM , Hui DY
Ref : Journal of Biological Chemistry , 275 :24040 , 2000
Abstract : Previous studies using chemical modification approach have shown the importance of arginine residues in bile salt activation of carboxyl ester lipase (CEL) activity. However, the x-ray crystal structure of CEL failed to show the involvement of arginine residues in CEL-bile salt interaction. The current study used a site-specific mutagenesis approach to determine the role of arginine residues 63 and 423 in bile salt-dependent and bile salt-independent hydrolytic activities of rat CEL. Mutations of Arg(63) to Ala(63) (R63A) and Arg(423) to Gly(423) (R423G) resulted in enzymes with increased bile salt-independent hydrolytic activity against lysophosphatidylcholine, having 6.5- and 2-fold higher k(cat) values, respectively, in comparison to wild type CEL. In contrast, the R63A and R423A mutant enzymes displayed 5- and 11-fold decreases in k(cat), in comparison with wild type CEL, for bile salt-dependent cholesteryl ester hydrolysis. Although taurocholate induced similar changes in circular dichroism spectra for wild type, R63A, and R423G proteins, this bile salt was less efficient in protecting the mutant enzymes against thermal inactivation in comparison with control CEL. Lipid binding studies revealed less R63A and R423G mutant CEL were bound to 1,2-diolein monolayer at saturation compared with wild type CEL. These results, along with computer modeling of the CEL protein, indicated that Arg(63) and Arg(423) are not involved directly with monomeric bile salt binding. However, these residues participate in micellar bile salt modulation of CEL enzymatic activity through intramolecular hydrogen bonding with the C-terminal domain. These residues are also important, probably through similar intramolecular hydrogen bond formation, in stabilizing the enzyme in solution and at the lipid-water interface.
ESTHER : Liang_2000_J.Biol.Chem_275_24040
PubMedSearch : Liang_2000_J.Biol.Chem_275_24040
PubMedID: 10811659

Title : Theoretical and experimental investigations of electrostatic effects on acetylcholinesterase catalysis and inhibition - Malany_1999_Chem.Biol.Interact_119-120_99
Author(s) : Malany S , Baker NA , Verweyst M , Medhekar R , Quinn DM , Velan B , Kronman C , Shafferman A
Ref : Chemico-Biological Interactions , 119-120 :99 , 1999
Abstract : The role of electrostatics in the function of acetylcholinesterase (AChE) has been investigated by both theoretical and experimental approaches. Second-order rate constants (kE = k(cat)/Km) for acetylthiocholine (ATCh) turnover have been measured as a function of ionic strength of the reaction medium for wild-type and mutant AChEs. Also, binding and dissociation rate constants have been measured as a function of ionic strength for the respective charged and neutral transition state analog inhibitors m-(N,N,N-trimethylammonio)trifluoroacetophenone (TMTFA) and m-(t-butyl)trifluoroacetophenone (TBTFA). Linear free-energy correlations between catalytic rate constants and inhibition constants indicate that kE for ATCh turnover is rate limited by terminal binding events. Comparison of binding rate constants for TMTFA and TBTFA attests to the sizable electrostatic discrimination of AChE. Free energy profiles for cationic ligand release from the active sites of wild-type and mutant AChEs have been calculated via a model that utilizes the structure of T. californica AChE, a spherical ligand, and energy terms that account for electrostatic and van der Waals interactions and chemical potential. These calculations indicate that EA and EI complexes are not bound with respect to electrostatic interactions, which obviates the need for a 'back door' for cationic ligand release. Moreover, the computed energy barriers for ligand release give linear free-energy correlations with log(kE) for substrate turnover, which supports the general correctness of the computational model.
ESTHER : Malany_1999_Chem.Biol.Interact_119-120_99
PubMedSearch : Malany_1999_Chem.Biol.Interact_119-120_99
PubMedID: 10421443

Title : Change in the mode of inhibition of acetylcholinesterase by (4- nitrophenyl)sulfonoxyl derivatives of conformationally constrained choline analogues - Savle_1998_Chem.Res.Toxicol_11_19
Author(s) : Savle PS , Medhekar RA , Kelley EL , May JG , Watkins SF , Fronczek FR , Quinn DM , Gandour RD
Ref : Chemical Research in Toxicology , 11 :19 , 1998
Abstract : A chiral, five-step synthesis of 2-(hydroxymethyl)-2,4-dimethylmorpholine (12) from (R)- and (S)-2-methylglycidols gives an overall yield of 63%. Morpholines (R)- and (S)-12 are converted into 2-(azidomethyl)-2,4-dimethylmorpholine (15) via 2,4-dimethyl-2-[[(4-nitrophenyl)sulfonoxy]methyl]morpholine (14). The tertiary morpholines 12, 14, and 15 are quaternarized to afford 2-(hydroxymethyl)-2,4,4-trimethylmorpholinum iodide (2), 2,4,4-trimethyl-2-[[(4-nitrophenyl)sulfonoxy]methyl]morpholinium iodide (3), and 2-(azidomethyl)-2,4,4-trimethylmorpholinium iodide (4), respectively, which all inhibit acetylcholinesterase (AChE). These morpholinium inhibitors are compared with conformationally constrained aryl hemicholinium AChE inhibitors. Enantiomers of 2 and 4 are reversible competitive inhibitors of AChE, with values of Ki = 360 +/- 30 microM for (S)-2, 650 +/- 90 microM for (R)-2, 450 +/- 70 microM for (S)-4, and 560 +/- 30 microM for (R)-4, respectively. Enantiomers of 3 are noncompetitive inhibitors of AChE with values of Ki = 19.0 +/- 0.9 microM for (S)-3 and 50 +/- 2 microM for (R)-3, respectively. AChE shows a 2-fold chiral discrimination in the case of inhibition by 2 and 3. Inhibition also changes from competitive to noncompetitive when (3-hydroxyphenyl)-N,N,N-trimethylammonium iodide (18) [Ki = 0.21 +/- 0.06 microM; Lee, B. H., Stelly, T. C., Colucci, W. J., Garcia, J. G., Gandour, R. D., and Quinn, D. M. (1992) Chem. Res. Toxicol. 5, 411-418] is converted into [3-[(4-nitrophenyl)sulfonoxy]phenyl]-N,N,N-trimethylammonium iodide (5), Ki = 6.0 +/- 0.5 microM. These results indicate that the 4-nitrobenzenesulfonyl group controls the mode of inhibition.
ESTHER : Savle_1998_Chem.Res.Toxicol_11_19
PubMedSearch : Savle_1998_Chem.Res.Toxicol_11_19
PubMedID: 9477222

Title : Structure of the Acylation Transition State for Acetylcholinesterase Catalysis -
Author(s) : Malany S , Sikorski RS , Seravalli J , Medhekar R , Quinn DM
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. :197 , 1998

Title : Correlation of Isotope and Viscosity Effects -
Author(s) : Malany S , Taylor P , Quinn DM , Sawai M , Shafferman A , Velan B , Kronman C
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. :232 , 1998

Title : Low-Barrier Hydrogen Bond in the Catalytic Triad of Serine Enzymes -
Author(s) : Medhekar R , Baker NA , Sando K , Kearney W , Quinn DM
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. :233 , 1998

Title : Electrostatic influence on the kinetics of ligand binding to acetylcholinesterase. Distinctions between active center ligands and fasciculin - Radic_1997_J.Biol.Chem_272_23265
Author(s) : Radic Z , Kirchhoff PD , Quinn DM , McCammon JA , Taylor P
Ref : Journal of Biological Chemistry , 272 :23265 , 1997
Abstract : To explore the role that surface and active center charges play in electrostatic attraction of ligands to the active center gorge of acetylcholinesterase (AChE), and the influence of charge on the reactive orientation of the ligand, we have studied the kinetics of association of cationic and neutral ligands with the active center and peripheral site of AChE. Electrostatic influences were reduced by sequential mutations of six surface anionic residues outside of the active center gorge (Glu-84, Glu-91, Asp-280, Asp-283, Glu-292, and Asp-372) and three residues within the active center gorge (Asp-74 at the rim and Glu-202 and Glu-450 at the base). The peripheral site ligand, fasciculin 2 (FAS2), a peptide of 6.5 kDa with a net charge of +4, shows a marked enhancement of rate of association with reduction in ionic strength, and this ionic strength dependence can be markedly reduced by progressive neutralization of surface and active center gorge anionic residues. By contrast, neutralization of surface residues only has a modest influence on the rate of cationic m-trimethylammoniotrifluoroacetophenone (TFK+) association with the active serine, whereas neutralization of residues in the active center gorge has a marked influence on the rate but with little change in the ionic strength dependence. Brownian dynamics calculations for approach of a small cationic ligand to the entrance of the gorge show the influence of individual charges to be in quantitative accord with that found for the surface residues. Anionic residues in the gorge may help to orient the ligand for reaction or to trap the ligand. Bound FAS2 on AChE not only reduces the rate of TFK+ reaction with the active center but inverts the ionic strength dependence for the cationic TFK+ association with AChE. Hence it appears that TFK+ must traverse an electrostatic barrier at the gorge entry imparted by the bound FAS2 with its net charge of +4.
ESTHER : Radic_1997_J.Biol.Chem_272_23265
PubMedSearch : Radic_1997_J.Biol.Chem_272_23265
PubMedID: 9287336

Title : Molecular modeling of the structures of human and rat pancreatic cholesterol esterases - Feaster_1997_Protein.Sci_6_73
Author(s) : Feaster SR , Quinn DM , Barnett BL
Ref : Protein Science , 6 :73 , 1997
Abstract : Structural models have been generated for rat and human cholesterol esterases by molecular modeling. For rat cholesterol esterase, three separate models were generated according to the following procedure: (1) the cholesterol esterase sequence was aligned with those of three template enzymes: Torpedo californica acetylcholinesterase, Geotrichum candidum lipase and Candida rugosa lipase; (2) the X-ray structure coordinates of the three template enzymes were used to construct cholesterol esterase models by amino acid replacements of matched sequence positions and by making sequence insertions and deletions as required; (3) bad contracts in each of the cholesterol esterase models were relaxed by molecular dynamics and mechanics; (4) the three cholesterol esterase models were merged into one by arithmetic averaging of atomic coordinates; (5) Ramachandran analysis indicated that the model generated from the AChE template possessed the best set of phi/psi angles. Therefore, this model was subjected to molecular dynamics, with harmonic constraints imposed on the C(alpha) coordinates to drive them toward the coordinates of the averaged model. (6) Subsequent relaxation by molecular mechanics produced the final rat cholesterol esterase model. A model for human cholesterol esterase was produced by repeating steps 1-3 above, albeit with the rat cholesterol esterase model as the template. Hydrophobic and electrostatic analyses of the rat and human cholesterol esterase models suggest the structural origins of molecular recognition of hydrophobic substrates and interfaces, of charged interfaces, and of bile salt activators.
ESTHER : Feaster_1997_Protein.Sci_6_73
PubMedSearch : Feaster_1997_Protein.Sci_6_73
PubMedID: 9007978

Title : Mutant acetylcholinesterases as potential detoxification agents for organophosphate poisoning - Saxena_1997_Biochem.Pharmacol_54_269
Author(s) : Saxena A , Maxwell DM , Quinn DM , Radic Z , Taylor P , Doctor BP
Ref : Biochemical Pharmacology , 54 :269 , 1997
Abstract : It has been demonstrated that cholinesterases (ChEs) are an effective mode of pretreatment to prevent organophosphate (OP) toxicity in mice and rhesus monkeys. The efficacy of ChE as a bioscavenger of OP can be enhanced by combining enzyme pretreatment with oxime reactivation, since the scavenging capacity extends beyond a stoichiometric ratio of ChE to OP. Aging has proven to be a major barrier to achieving oxime reactivation of acetylcholinesterase (AChE) inhibited by the more potent OPs. To further increase the stoichiometry of OP to ChE required, we have sought AChE mutants that are more easily reactivated than wild-type enzyme. Substitution of glutamine for glutamate (E199) located at the amino-terminal to the active-site serine (S200) in Torpedo AChE generated an enzyme largely resistant to aging. Here we report the effect of the corresponding mutation on the rate of inhibition, reactivation by 1-(2-hydroxyiminomethyl-1-pyridinium)-1(4-carboxyaminopyridinium)- dimethyl ether hydrochloride (HI-6), and aging of mouse AChE inhibited by C(+)P(-)- and C(-)P(-)-epimers of soman. The E202 to Q mutation decreased the affinity of soman for AChE, slowed the reactivation of soman-inhibited AChE by HI-6, and decreased the aging of mutant AChE. These effects were more pronounced with C(-)P(-)-soman than with C(+)P(-)-soman. In vitro detoxification of soman and sarin by wild-type and E202Q AChE in the presence of 2 mM HI-6 showed that, E202Q AChE was 2-3 times more effective in detoxifying soman and sarin than wild-type AChE. These studies show that these recombinant DNA-derived AChEs are a great improvement over wild-type AChE as bioscavengers. They can be used to develop effective methods for the safe disposal of stored OP nerve agents and potential candidates for pre- or post-exposure treatment for OP toxicity.
ESTHER : Saxena_1997_Biochem.Pharmacol_54_269
PubMedSearch : Saxena_1997_Biochem.Pharmacol_54_269
PubMedID: 9271331

Title : Mechanism-based inhibitors of mammalian cholesterol esterase -
Author(s) : Feaster SR , Quinn DM
Ref : Methods Enzymol , 286 :231 , 1997
PubMedID: 9309653

Title : The X-ray structure of a transition state analog complex reveals the molecular origins of the catalytic power and substrate specificity of acetylcholinesterase. -
Author(s) : Harel M , Quinn DM , Nair HK , Silman I , Sussman JL
Ref : Journal of the American Chemical Society , 118 :2340 , 1996
Gene_locus related to this paper: torca-ACHE

Title : Molecular recognition by cholesterol esterase of active site ligands: structure-reactivity effects for inhibition by aryl carbamates and subsequent carbamylenzyme turnover - Feaster_1996_Biochemistry_35_16723
Author(s) : Feaster SR , Lee K , Baker N , Hui DY , Quinn DM
Ref : Biochemistry , 35 :16723 , 1996
Abstract : Interactions of mammalian pancreatic cholesterol esterases from pig and rat with a family of aryl carbamates CnH2n+1NHCOOAr [n = 4-9; Ar = phenyl, p-X-phenyl (X = acetamido, bromo, fluoro, nitro, trifluoromethyl), 2-naphthyl, 2-tetrahydronaphthyl, estronyl] have been investigated, with an aim of delineating the ligand structural features which lead to effective molecular recognition by the active site of the enzyme. These carbamates inhibit the catalytic activity of CEase by rapid carbamylation of the active site, a process that shows saturation kinetics. Subsequent slow decarbamylation usually leads to full restoration of activity, and therefore aryl carbamates are transient inhibitors, or pseudo-substrates, of CEase. Structural variation of carbamate inhibitors allowed molecular recognition in the fatty acid binding and steroid binding loci of the extended active site to be probed, and the electronic nature of the carbamylation transition state to be characterized. Optimal inhibitory activity is observed when the length of the carbamyl function is n = 6 and n = 7 for porcine and rat cholesterol esterases, respectively, equivalent to eight- and nine-carbon fatty acyl chains. In contrast, inhibitory activity increases progressively as the partial molecular volume of the aromatic fragment increases. Hammett plots for p-substituted phenyl-N-hexyl carbamates indicate that the rate-determining step for carbamate inhibition is phenolate anion expulsion. Effects of the bile salt activator taurocholate on the kinetically resolved phases of the pseudo-substrate turnover of aryl carbamates were also studied. Taurocholate increases the affinity of the carbamate for the active site of cholesterol esterase in the reversible, noncovalent complex that precedes carbamylation and increases the rate constants of the serial carbamylation and decarbamylation steps. Structural variation of the N-alkyl chain and of the aryl fused-ring system provides an accounting of bile salt modulation of the fatty acid and steroid binding sites, respectively. In that pseudo-substrate turnover of aryl carbamates proceeds by a three-step mechanism that is analogous to that for rapid turnover of lipid ester substrates, these investigations illuminate details of ligand recognition by the extended active site of cholesterol esterase that are prominent determinants of the substrate specificity and catalytic power of the enzyme.
ESTHER : Feaster_1996_Biochemistry_35_16723
PubMedSearch : Feaster_1996_Biochemistry_35_16723
PubMedID: 8988009

Title : Aspartate 74 as a primary determinant in acetylcholinesterase governing specificity to cationic organophosphonates - Hosea_1996_Biochemistry_35_10995
Author(s) : Hosea NA , Radic Z , Tsigelny I , Berman HA , Quinn DM , Taylor P
Ref : Biochemistry , 35 :10995 , 1996
Abstract : Through site-specific mutagenesis, we examined the determinants on acetylcholinesterase which govern the specificity and reactivity of three classes of substrates: enantiomeric alkyl phosphonates, trifluoromethyl acetophenones, and carboxyl esters. By employing cationic and uncharged pairs of enantiomeric alkyl methylphosphonyl thioates of known absolute stereochemistry, we find that an aspartate residue near the gorge entrance (D74) is responsible for the enhanced reactivity of the cationic organophosphonates. Removal of the charge with the mutation D74N causes a near equal reduction in the reaction rate constants for the Rp and Sp enantiomers and exerts a greater influence on the cationic organophosphonates than on the charged trimethylammonio trifluoromethyl acetophenone and acetylthiocholine. This pattern of reactivity suggests that the orientation of the leaving group for both enantiomers is directed toward the gorge exit and in apposition to Asp 74. Replacement of tryptophan 86 with alanine in the choline subsite also diminishes the reaction rates for cationic organophosphonates, although to a lesser extent than with the D74N mutation, while not affecting the reactions with the uncharged compounds. Hence, reaction with cationic OPs depends to a lesser degree on Trp 86 than on Asp 74. Docking of Sp and Rp cycloheptyl methylphosphonyl thiocholines and thioethylates in AChE as models of the reversible complex and transition state using molecular dynamics affords structural insight into the spatial arrangement of the substituents surrounding phosphorus prior to and during reaction. The leaving group of the Rp and Sp enantiomers, regardless of charge, is directed to the gorge exit and toward Asp 74, an orientation unique to tetrahedral ligands.
ESTHER : Hosea_1996_Biochemistry_35_10995
PubMedSearch : Hosea_1996_Biochemistry_35_10995
PubMedID: 8718893

Title : Structures of Complexes of Acetylcholinesterase with Covalently and Non-Covalently Bound Inhibitors -
Author(s) : Sussman JL , Harel M , Raves ML , Quinn DM , Nair HK , Silman I
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. :59 , 1995

Title : Amino acid residues controlling reactivation of organophosphonyl conjugates of acetylcholinesterase by mono- and bisquaternary oximes - Ashani_1995_J.Biol.Chem_270_6370
Author(s) : Ashani Y , Radic Z , Tsigelny I , Vellom DC , Pickering NA , Quinn DM , Doctor BP , Taylor P
Ref : Journal of Biological Chemistry , 270 :6370 , 1995
Abstract : Single and multiple site mutants of recombinant mouse acetylcholinesterase (rMoAChE) were inhibited with racemic 7-(methylethoxyphosphinyloxy)-1-methylquinolinium iodide (MEPQ) and the resulting mixture of two enantiomers, CH3PR,S(O)(OC2H5)-AChE(EMPR,S-AChE), were subjected to reactivation with 2-(hydroxyiminomethyl)-1-methylpyridinium methanesulfonate (P2S) and 1-(2'-hydroxyiminomethyl-1'-pyridinium)-3-(4"-carbamoyl-1"- pyridinium)-2-oxapropane dichloride (HI-6). Kinetic analysis of the reactivation profiles revealed biphasic behavior with an approximate 1:1 ratio of two presumed reactivatable enantiomeric components. Equilibrium dissociation and kinetic rate constants for reactivation of site-specific mutant enzymes were compared with those obtained for wild-type rMoAChE, tissue-derived Torpedo AChE and human plasma butyrylcholinesterase. Substitution of key amino acid residues at the entrance to the active-site gorge (Trp-286, Tyr-124, Tyr-72, and Asp-74) had a greater influence on the reactivation kinetics of the bisquaternary reactivator HI-6 compared with the monoquaternary reactivator P2S. Replacement of Phe-295 by Leu enhanced reactivation by HI-6 but not by P2S. Of residues forming the choline-binding subsite, the E202Q mutation had a dominant influence where reactivation by both oximes was decreased 16- to 33-fold. Residues Trp-86 and Tyr-337 in this subsite showed little involvement. These kinetic findings, together with energy minimization of the oxime complex with the phosphonylated enzyme, provide a model for differences in the reactivation potencies of P2S and HI-6. The two kinetic components of oxime reactivation of MEPQ-inhibited AChEs arise from the chirality of O-ethyl methylphosphonyl moieties conjugated with Ser-203 and may be attributable to the relative stability of the phosphonyl oxygen of the two enantiomers in the oxyanion hole.
ESTHER : Ashani_1995_J.Biol.Chem_270_6370
PubMedSearch : Ashani_1995_J.Biol.Chem_270_6370
PubMedID: 7890775

Title : Amino Acid Residues in Acetylcholinesterase which Influence Fasciculin Inhibition -
Author(s) : Radic Z , Quinn DM , Vellom DC , Camp S , Taylor P
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. :183 , 1995

Title : Amino Acid Residues that Control Mono- and Bisquaternary Oxime-Induced Reactivation of O-Ethyl Methylphosphonylated Cholinesterases -
Author(s) : Ashani Y , Radic Z , Tsigelny I , Vellom DC , Pickering NA , Quinn DM , Doctor BP , Taylor P
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. :133 , 1995

Title : Allosteric control of acetylcholinesterase catalysis by fasciculin - Radic_1995_J.Biol.Chem_270_20391
Author(s) : Radic Z , Quinn DM , Vellom DC , Camp S , Taylor P
Ref : Journal of Biological Chemistry , 270 :20391 , 1995
Abstract : The interaction of fasciculin 2 was examined with wild-type and several mutant forms of acetylcholinesterase (AChE) where Trp86, which lies at the base of the active center gorge, is replaced by Tyr, Phe, and Ala. The fasciculin family of peptides from snake venom bind to a peripheral site near the rim of the gorge, but at a position which still allows substrates and other inhibitors to enter the gorge. The interaction of a series of charged and uncharged carboxyl esters, alkyl phosphoryl esters, and substituted trifluoroacetophenones were analyzed with the wild-type and mutant AChEs in the presence and absence of fasciculin. We show that Trp86 is important for the alignment of carboxyl ester substrates in the AChE active center. The most marked influence of Trp86 substitution in inhibiting catalysis is seen for carboxyl esters that show rapid turnover. The extent of inhibition achieved with bound fasciculin is also greatest for efficiently catalyzed, charged substrates. When Ala is substituted for Trp86, fasciculin becomes an allosteric activator instead of an inhibitor for certain substrates. Analysis of the kinetics of acylation by organophosphates and conjugation by trifluoroacetophenones, along with deconstruction of the kinetic constants for carboxyl esters, suggests that AChE inhibition by fasciculin arises from reductions of both the commitment to catalysis and diffusional entry of substrate into the gorge. The former is reflected in the ratio of the rate constant for substrate acylation to that for dissociation of the initial complex. The action of fasciculin appears to be mediated allosterically from its binding site at the rim of the gorge to affect the orientation of the side chain of Trp86 which lies at the gorge base.
ESTHER : Radic_1995_J.Biol.Chem_270_20391
PubMedSearch : Radic_1995_J.Biol.Chem_270_20391
PubMedID: 7657613

Title : London Dispersion Interactions in Molecular Recognition by Acetylcholinesterase -
Author(s) : Quinn DM , Nair HK , Seravalli J , Lee K , Arbuckle T , Radic Z , Vellom DC , Pickering NA , Taylor P
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. :228 , 1995

Title : The Function of Electrostatics in Acetylcholinesterase Catalysis -
Author(s) : Quinn DM , Seravalli J , Nair HK , Medhekar R , Husseini B , Radic Z , Vellom DC , Pickering NA , Taylor P
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. :203 , 1995

Title : Identification of amino acid residues involved in the binding of Huperzine A to cholinesterases - Saxena_1994_Protein.Sci_3_1770
Author(s) : Saxena A , Qian N , Kovach IM , Kozikowski AP , Pang YP , Vellom DC , Radic Z , Quinn DM , Taylor P , Doctor BP
Ref : Protein Science , 3 :1770 , 1994
Abstract : Huperzine A, a potential agent for therapy in Alzheimer's disease and for prophylaxis of organophosphate toxicity, has recently been characterized as a reversible inhibitor of cholinesterases. To examine the specificity of this novel compound in more detail, we have examined the interaction of the 2 stereoisomers of Huperzine A with cholinesterases and site-specific mutants that detail the involvement of specific amino acid residues. Inhibition of fetal bovine serum acetylcholinesterase by (-)-Huperzine A was 35-fold more potent than (+)-Huperzine A, with KI values of 6.2 nM and 210 nM, respectively. In addition, (-)-Huperzine A was 88-fold more potent in inhibiting Torpedo acetylcholinesterase than (+)-Huperzine A, with KI values of 0.25 microM and 22 microM, respectively. Far larger KI values that did not differ between the 2 stereoisomers were observed with horse and human serum butyrylcholinesterases. Mammalian acetylcholinesterase, Torpedo acetylcholinesterase, and mammalian butyrylcholinesterase can be distinguished by the amino acid Tyr, Phe, or Ala in the 330 position, respectively. Studies with mouse acetylcholinesterase mutants, Tyr 337 (330) Phe and Tyr 337 (330) Ala yielded a difference in reactivity that closely mimicked the native enzymes. In contrast, mutation of the conserved Glu 199 residue to Gln in Torpedo acetylcholinesterase produced only a 3-fold increase in KI value for the binding of Huperzine A.
ESTHER : Saxena_1994_Protein.Sci_3_1770
PubMedSearch : Saxena_1994_Protein.Sci_3_1770
PubMedID: 7849595

Title : Molecular recognition in acetylcholinesterase catalysis: free-energy correlations for substrate turnover and inhibition by trifluoro ketone transition-state analogs - Nair_1994_Biochemistry_33_8566
Author(s) : Nair HK , Seravalli J , Arbuckle T , Quinn DM
Ref : Biochemistry , 33 :8566 , 1994
Abstract : Ten meta-substituted aryl trifluoromethyl ketones (m-XC6H4COCF3; X = H, CH3, CF3, C2H5, isopropyl, t-butyl, NH2, NMe2, N+Me3, NO2) have been evaluated as inhibitors of acetylcholinesterases from Electrophorus electricus and Torpedo californica. Trifluoro ketones that have small meta substituents (X = H, CH3, CF3, C2H5, NH2, NO2) are rapid reversible inhibitors, whereas the remaining compounds in this study show time-dependent inhibition. Dissociation constants (Ki values) for these compounds span a range of approximately 10(7)-fold, with trifluoroacetophenone (X = H) being the least potent and m-(N,N,N-trimethylammonio)trifluoroacetophenone (X = Me3N+) being the most potent inhibitor. For the latter compound Ki values are 1.5 and 15 fM for inhibitions of the respective acetylcholinesterases (Nair, H. K., Lee, K., & Quinn, D. M. (1993) J. Am. Chem. Soc. 115, 9939-9941). Linear correlations of log(kcat/Km) for substrate turnover versus pKi of inhibitors have slopes of approximately 0.6, which suggest that aryl trifluoro ketones bind to AChE in a manner that structurally resembles transition states in the acylation stage of catalysis. Substituent variation in the inhibitors allows one to gauge the importance for AChE function of molecular recognition in the quaternary ammonium binding locus of the active site. This locus is frequently termed the "anionic site" and consists of E199, W84, and perhaps Y130 and F330. Correlations of pKi versus hydrophobicity constant are linear for alkyl and trifluoromethyl substituents but fail for nitrogen-containing substituents. However, three-dimensional correlations of pKi versus sigma m and molar refractivity of substituents indicate that dispersion interactions in the anionic locus contribute approximately 10(5)-fold (delta delta G = 7 kcal mol-1) to the above-mentioned 10(7)-fold range of inhibitor potencies. The remaining approximately 100-fold arises from the inductive electronic effects of substituents on the stability of the tetrahedral adduct that forms between the ketone carbonyl of inhibitors and S200 in the esteratic locus of the active site. Values of k(on), the second-order rate constant for binding of time-dependent inhibitors, monitor a diffusion-controlled process. Moreover, k(on) for the quaternary ammonio inhibitor is 20-70-fold higher than for inhibitors that have uncharged meta substituents, which likely reflects the effect of the electrical field of AChE on ligand and substrate binding.
ESTHER : Nair_1994_Biochemistry_33_8566
PubMedSearch : Nair_1994_Biochemistry_33_8566
PubMedID: 8031791

Title : Cryptic Catalysis and Cholinesterase Function -
Author(s) : Quinn DM , Selwood T
Ref : In Multidisciplinary approaches to cholinesterase functions - Proceedings of Fourth International Meeting on Cholinesterases , (Shafferman, A. and Velan, B., Eds) Plenum Press, New York :141 , 1992

Title : Inhibition of acetylcholinesterase by hemicholiniums, conformationally constrained choline analogues. Evaluation of aryl and alkyl substituents. Comparisons with choline and (3- hydroxyphenyl)trimethylammonium - Lee_1992_Chem.Res.Toxicol_5_411
Author(s) : Lee BH , Stelly TC , Colucci WJ , Garcia JG , Gandour RD , Quinn DM
Ref : Chemical Research in Toxicology , 5 :411 , 1992
Abstract : 2-Substituted-2-hydroxy-4,4-dimethylmorpholiniums (hemicholiniums) inhibit acetylcholinesterase (EC hydrolysis of acetylthiocholine (ATCh). The 4-substituted arenes [NH2, NHC(O)CH3, Cl, CN, and NO2] have values of inhibition constants (Ki) that range from 220 to 3690 microM, which correlate with Hammett sigma, rho approximately 0.8. The alkyl compounds, hydrogen, methyl, tert-butyl, and trifluoromethyl, have values of Ki of 550, 560, 1200, and 1200 microM, respectively. These values compare favorably with Ki = 960 microM for choline. The conformation of AChE-bound choline must be gauche to support our suggestion that hemicholiniums are conformationally constrained analogues of choline. (3-Hydroxyphenyl)trimethylammonium (5) inhibits most strongly, Ki = 0.21 microM, of the compounds examined in this study. The solvent isotope effect (H2OKi/D2OKi = 0.83 +/- 0.04) suggests that inhibition by 5 involves hydrogen bonding. The binding by AChE of the hemicholiniums of various sizes and the strong binding of 5 support an earlier proposal [Schowen, K. B., Smissman, E. E., and Stephen, W. F., Jr. (1975) J. Med. Chem. 18, 292-300] that the active site of AChE has ample space for rotation about the C-C bond in choline. Compound 5, which has one more carbon between the hydroxy and trimethylammonium than does choline, inhibits much more potently than either choline or the hemicholiniums. Compound 5 provides a correct spacer to span the trimethylammonium recognition site and the esteratic site of AChE. This aromatic spacer interacts favorably with the hydrophobic active site, and the phenolic hydroxyl probably hydrogen bonds to the histidine in the esteratic site. Choline in any conformation and the hemicholiniums are too short to make a strong hydrogen bond.
ESTHER : Lee_1992_Chem.Res.Toxicol_5_411
PubMedSearch : Lee_1992_Chem.Res.Toxicol_5_411
PubMedID: 1504265

Title : Cholesterol esterase catalyzed hydrolysis of mixed micellar thiophosphatidylcholines: a possible charge-relay mechanism - Sutton_1991_Biochemistry_30_5888
Author(s) : Sutton LD , Froelich S , Hendrickson HS , Quinn DM
Ref : Biochemistry , 30 :5888 , 1991
Abstract : Mechanistic features of cholesterol esterase catalyzed hydrolysis of two thiophospholipids, rac-1-(hexanoylthio)-2-hexanoyl-3-glycerophosphorylcholine (6TPC) and rac-1-(decanoylthio)-2-decano-yl-3-glycerophosphorylcholine (10TPC), have been characterized. The hydrolysis of 10TPC that is contained in mixed micelles with Triton X-100 occurs strictly at the micellar interface, since the reaction rate is independent of the micelle concentration but depends hyperbolically on the mole fraction of the substrate in the micelles. This latter observation allows one to calculate the interfacial kinetic parameters V*max and K*m. The hydrolyses of 10TPC and p-nitrophenyl butyrate are similarly inhibited by the transition state analogue inhibitor phenyl-n-butylborinic acid, and therefore, physiological and nonphysiological substrates are processed at the same active site. The similarity of k*cat values for the acyl-similar substrates 10TPC and p-nitrophenyl decanoate indicates that the phospholipase A1 activity of cholesterol esterase is partially rate limited by turnover of a decanoyl-enzyme intermediate. Solvent isotope effects on V*max and V*max/K*m (which monitors acylation only) are approximately 2-3 and are consistent with transition states that are stabilized by general acid-base proton transfers. Proton inventories of V*max/K*m indicate that simultaneous proton transfers stabilize the acylation transition state, which requires a multifunctional acid-base machinery (perhaps a charge-relay system) in the cholesterol esterase active site. Similar results are obtained for the 6TPC reaction, both in the presence and absence of Triton X-100 micelles.
ESTHER : Sutton_1991_Biochemistry_30_5888
PubMedSearch : Sutton_1991_Biochemistry_30_5888
PubMedID: 2043629

Title : The Chemical Mechanism of Acetylcholinesterase Reactions: Biological Catalysis at the Speed Limit -
Author(s) : Quinn DM , Pryor AN , Selwood T , Lee BH , Acheson SA , Barlow PN
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 :252 , 1991

Title : Anatomy of acetylcholinesterase catalysis: reaction dynamics analogy for human erythrocyte and electric eel enzymes - Acheson_1990_Biochim.Biophys.Acta_1040_199
Author(s) : Acheson SA , Quinn DM
Ref : Biochimica & Biophysica Acta , 1040 :199 , 1990
Abstract : The anatomy of catalysis (i.e., reaction dynamics, thermodynamics and transition state structures) is compared herein for acetylcholinesterases from human erythrocytes and Electrophorus electricus. The two enzymes have similar relative activities for the substrate o-nitrochloroacetanilide and o-nitrophenyl acetate. In addition, with each substrate K values and solvent deuterium kinetic isotope effects for kES and kE are similar for the two enzymes. Solvent isotope effects in mixed isotopic buffers indicate that the acylation stages of o-nitrochloroacetanilide turnover by the two enzymes are rate-limited by virtual transition states that are weighted averages of contributions from transition states of serial chemical and physical steps. Similar experiments show that the transition states for Vmax of o-nitrophenyl acetate turnover by the two enzymes are stabilized by simple general acid-base (i.e., one-proton) catalysis. These comparisons demonstrate that acetylcholinesterases from diverse sources display functional analogy in that reaction dynamics and transition state structures are closely similar.
ESTHER : Acheson_1990_Biochim.Biophys.Acta_1040_199
PubMedSearch : Acheson_1990_Biochim.Biophys.Acta_1040_199
PubMedID: 2400771

Title : Haloketone transition state analog inhibitors of cholesterol esterase - Sohl_1988_Biochem.Biophys.Res.Commun_151_554
Author(s) : Sohl J , Sutton LD , Burton DJ , Quinn DM
Ref : Biochemical & Biophysical Research Communications , 151 :554 , 1988
Abstract : The cholesterol esterase-catalyzed hydrolysis of p-nitro-phenyl butyrate is reversibly inhibited by four phenyl haloalkyl ketones. Inhibitor potency is greatest for halogenated acetophenones and parallels the extent of hydration of the various ketones in buffered D2O. These results are consistent with an inhibition mechanism wherein haloketones reversibly form hemiketal adducts at the active site that structurally mimic tetrahedral intermediates of the cholesterol esterase catalytic cycle.
ESTHER : Sohl_1988_Biochem.Biophys.Res.Commun_151_554
PubMedSearch : Sohl_1988_Biochem.Biophys.Res.Commun_151_554
PubMedID: 3348795

Title : p-Nitrophenyl and cholesteryl-N-alkyl carbamates as inhibitors of cholesterol esterase - Hosie_1987_J.Biol.Chem_262_260
Author(s) : Hosie L , Sutton LD , Quinn DM
Ref : Journal of Biological Chemistry , 262 :260 , 1987
Abstract : p-Nitrophenyl and cholesteryl-N-alkyl carbamates are good inhibitors of porcine pancreatic cholesterol esterase-catalyzed hydrolysis of p-nitrophenyl butyrate. p-Nitrophenyl-N-butyl and N-octyl carbamates (compounds 1 and 2, respectively) are potent active site-directed irreversible inhibitors of this enzyme. The inhibition of cholesterol esterase by compound 1 or 2 shows saturation kinetics with increasing inhibitor concentration. The activity of cholesterol esterase in the presence of compound 1 or 2 can be protected by the competitive inhibitor, phenylboronic acid. First-order decreases in cholesterol esterase activity effected by compound 1 or 2 are also observed in the presence of taurocholate/phosphatidylcholine micelles. Dilution of the inhibited enzyme results in a gradual return of activity, the rate of which is increased in the presence of the nucleophile hydroxylamine. Hence, inhibition of cholesterol esterase-catalyzed hydrolysis of p-nitrophenyl butyrate by compound 1 or 2 in the aqueous or micellar phase occurs via a carbamyl-cholesterol esterase mechanism. The turnover of the butyl carbamylenzyme is increased in the presence of micelles, which indicates that the micelles have a direct effect on the catalytic activity of the enzyme. However, this effect is dependent on the structure of the substrate as the turnover of the octyl carbamylenzyme is unaffected in the presence of micelles. A comparison of the second-order rate constants for the inhibition of cholesterol esterase by compound 1 or 2 indicates that the octyl derivative is the more potent inhibitor. Cholesteryl-N-alkyl carbamates do not carbamylate cholesterol esterase but instead act as reversible inhibitors. This is due to the stability of cholesteryl carbamates relative to p-nitrophenyl carbamates.
ESTHER : Hosie_1987_J.Biol.Chem_262_260
PubMedSearch : Hosie_1987_J.Biol.Chem_262_260
PubMedID: 3793726
Gene_locus related to this paper: pig-i3ltk9

Title : Acetylcholinesterase: enzyme structure, reaction dynamics and virtual transition states -
Author(s) : Quinn DM
Ref : Chem Rev , 87 :955 , 1987

Title : Phenyl-n-butylborinic acid is a potent transition state analog inhibitor of lipolytic enzymes - Sutton_1986_Biochem.Biophys.Res.Commun_134_386
Author(s) : Sutton LD , Stout JS , Hosie L , Spencer PS , Quinn DM
Ref : Biochemical & Biophysical Research Communications , 134 :386 , 1986
Abstract : The cholesterol esterase and lipoprotein lipase catalyzed hydrolyses of the water-soluble substrate p-nitrophenyl butyrate are competitively inhibited by butaneboronic acid and phenylboronic acid. Phenyl-n-butylborinic acid has been synthesized and characterized as an ultrapotent transition state analog inhibitor: Ki = 2.9 +/- 0.6 nM and 1.7 +/- 0.3 microM for the cholesterol esterase and lipoprotein lipase reactions, respectively. These results are interpreted in terms of transition state structure and stabilization.
ESTHER : Sutton_1986_Biochem.Biophys.Res.Commun_134_386
PubMedSearch : Sutton_1986_Biochem.Biophys.Res.Commun_134_386
PubMedID: 3947331

Title : Interfacial reaction dynamics and acyl-enzyme mechanism for lipoprotein lipase-catalyzed hydrolysis of lipid p-nitrophenyl esters - Burdette_1986_J.Biol.Chem_261_12016
Author(s) : Burdette RA , Quinn DM
Ref : Journal of Biological Chemistry , 261 :12016 , 1986
Abstract : The fatty acyl (lipid) p-nitrophenyl esters p-nitrophenyl caprylate, p-nitrophenyl laurate and p-nitrophenyl palmitate that are incorporated at a few mol % into mixed micelles with Triton X-100 are substrates for bovine milk lipoprotein lipase. When the concentration of components of the mixed micelles is approximately equal to or greater than the critical micelle concentration, time courses for lipoprotein lipase-catalyzed hydrolysis of the esters are described by the integrated form of the Michaelis-Menten equation. Least square fitting to the integrated equation therefore allows calculation of the interfacial kinetic parameters Km and Vmax from single runs. The computational methodology used to determine the interfacial kinetic parameters is described in this paper and is used to determine the intrinsic substrate fatty acyl specificity of lipoprotein lipase catalysis, which is reflected in the magnitude of kcat/Km and kcat. The results for interfacial lipoprotein lipase catalysis, along with previously determined kinetic parameters for the water-soluble esters p-nitrophenyl acetate and p-nitrophenyl butyrate, indicate that lipoprotein lipase has highest specificity for the substrates that have fatty acyl chains of intermediate length (i.e. p-nitrophenyl butyrate and p-nitrophenyl caprylate). The fatty acid products do not cause product inhibition during lipoprotein lipase-catalyzed hydrolysis of lipid p-nitrophenyl esters that are contained in Triton X-100 micelles. The effects of the nucleophiles hydroxylamine, hydrazine, and ethylenediamine on Km and Vmax for lipoprotein lipase catalyzed hydrolysis of p-nitrophenyl laurate are consistent with trapping of a lauryl-lipoprotein lipase intermediate. This mechanism is confirmed by analysis of the product lauryl hydroxamate when hydroxylamine is the nucleophile. Hence, lipoprotein lipase-catalyzed hydrolysis of lipid p-nitrophenyl esters that are contained in Triton X-100 micelles occurs via an interfacial acyl-lipoprotein lipase mechanism that is rate-limited by hydrolysis of the acyl-enzyme intermediate.
ESTHER : Burdette_1986_J.Biol.Chem_261_12016
PubMedSearch : Burdette_1986_J.Biol.Chem_261_12016
PubMedID: 3745178

Title : Acylenzyme mechanism and solvent isotope effects for cholesterol esterase-catalyzed hydrolysis of p-nitrophenyl butyrate - Stout_1985_Biochim.Biophys.Acta_837_6
Author(s) : Stout JS , Sutton LD , Quinn DM
Ref : Biochimica & Biophysica Acta , 837 :6 , 1985
Abstract : The mechanism of cholesterol esterase- (carboxylic ester hydrolase, EC catalyzed hydrolysis of the water-soluble ester p-nitrophenyl butyrate has been characterized for commercially available preparations from bovine and porcine pancreas and for a purified preparation from porcine pancreas. Kinetic evidence for an acylenzyme mechanism is provided by experiments wherein the butyryl enzyme is trapped by MeOH, EtOH or n-BuOH. For the last alcohol the transacylation product n-butyl n-butyrate was characterized by GC-mass spectrometry. Solvent isotope effects have been measured for Vmax/Km, which is the rate constant for acylation, and for Vmax, which monitors rate-determining deacylation. Isotope effects of 1.5-3 on these rate constants indicate that both steps of the acylenzyme mechanism for cholesterol esterase catalysis involve transition states that are stabilized by general acid-base proton bridges.
ESTHER : Stout_1985_Biochim.Biophys.Acta_837_6
PubMedSearch : Stout_1985_Biochim.Biophys.Acta_837_6
PubMedID: 4052438

Title : Solvent isotope effects for lipoprotein lipase catalyzed hydrolysis of water-soluble p-nitrophenyl esters - Quinn_1985_Biochemistry_24_3144
Author(s) : Quinn DM
Ref : Biochemistry , 24 :3144 , 1985
Abstract : Solvent deuterium isotope effects on the rates of lipoprotein lipase (LpL) catalyzed hydrolysis of the water-soluble esters p-nitrophenyl acetate (PNPA) and p-nitrophenyl butyrate (PNPB) have been measured and fall in the range 1.5-2.2. The isotope effects are independent of substrate concentration, LpL stability, and reaction temperature and hence are effects on chemical catalysis and not due to a medium effect of D2O on LpL stability and/or conformation. pL (L = H or D) vs. rate profiles for the Vmax/Km of LpL-catalyzed hydrolysis of PNPB increase sigmoidally with increasing pL. Least-squares analysis of the profiles gives pKaH2O = 7.10 +/- 0.01, pKaD2O = 7.795 +/- 0.007, and a solvent isotope effect on limiting velocity at high pL of 1.97 +/- 0.03. Because the pL-rate profiles are for the Vmax/Km of hydrolysis of a water-soluble substrate, the measured pKa's are intrinsic acid-base ionization constants for a catalytically involved LpL active-site amino acid side chain. Benzeneboronic acid, a potent inhibitor of LpL-catalyzed hydrolysis of triacylglycerols [Vainio, P., Virtanen, J. A., & Kinnunen, P. K. J. (1982) Biochim. Biophys. Acta 711, 386-390], inhibits LpL-catalyzed hydrolysis of PNPB, with Ki = 6.9 microM at pH 7.36, 25 degrees C. This result and the solvent isotope effects for LpL-catalyzed hydrolysis of water-soluble esters are interpreted in terms of a proton transfer mechanism that is similar in many respects to that of the serine proteases.
ESTHER : Quinn_1985_Biochemistry_24_3144
PubMedSearch : Quinn_1985_Biochemistry_24_3144
PubMedID: 4027237
Gene_locus related to this paper: human-LPL

Title : Lipoprotein lipase catalyzed hydrolysis of water-soluble p-nitrophenyl esters. Inhibition by apolipoprotein C-II - Quinn_1982_Biochemistry_21_6872
Author(s) : Quinn DM , Shirai K , Jackson RL , Harmony JA
Ref : Biochemistry , 21 :6872 , 1982
Abstract : Bovine milk lipoprotein lipase (LpL) catalyzes the hydrolysis of the water-soluble esters p-nitrophenyl acetate (PNPA) and p-nitrophenyl butyrate (PNPB). The same protein and same active site are involved in hydrolysis of water-soluble p-nitrophenyl esters and emulsified trioleoylglycerol since (a) trioleoylglycerol hydrolysis and PNPB hydrolysis activities coelute from the heparin-Sepharose affinity column used to purify LpL and (b) LpL-catalyzed hydrolyses of trioleoylglycerol and PNPB are inhibited to equal extents by phenylmethanesulfonyl fluoride. The effect of apolipoprotein C-II (apoC-II) on the LpL-catalyzed hydrolysis of PNPA and PNPB has been determined. ApoC-II inhibits hydrolysis of both esters, with a maximum extent of inhibition of 70-90%. Inhibition of the LpL-catalyzed hydrolysis of PNPB is specific for apoC-II, since apolipoproteins A-I, C-I, and C-III-2 have little effect on this reaction, and is partial noncompetitive in form. KI values for apoC-II inhibition of the LpL-catalyzed hydrolysis of PNPA and PNPB are in the range 0.26-0.83 microM. The effect of apoC-II on the temperature dependences of LpL-catalyzed hydrolysis of both esters and on NaCl inhibition of LpL-catalyzed PNPB hydrolysis is consistent with a change in rate-determining step with LpL and apoC-II interact. These results indicate not only that there is an interaction between apoC-II and LpL in aqueous solution in the absence of a lipid interface but also that this interaction conformationally modulates the active site of the enzyme.
ESTHER : Quinn_1982_Biochemistry_21_6872
PubMedSearch : Quinn_1982_Biochemistry_21_6872
PubMedID: 7159570
Gene_locus related to this paper: bovin-lipli