Inhibition of <i>Drosophila melanogaster</i> acetylcholinesterase by high concentrations of substrate

Stojan, Jure; Brochier, L.; Alies, Carole; Colletier, J.P.; Fournier, Didier

doi:10.1111/j.1432-1033.2004.04048.x

Cited by 42 publications

(38 citation statements)

References 52 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In contrast to BuChE, and consistent with persistent observations for AChE,6 this enzyme shows substrate inhibition at high substrate concentrations. As will be discussed below, isotope effects again indicate that the deacylation stage of catalysis is rate limited by decomposition of an accumulating tetrahedral intermediate.…”

Section: Introductionsupporting

confidence: 87%

Accumulation of Tetrahedral Intermediates in Cholinesterase Catalysis: A Secondary Isotope Effect Study

et al. 2010

Self Cite

View full text Add to dashboard Cite

In a previous communication, kinetic β-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. (2005) JACS 127, 14538–14539). In this paper additional isotope effect experiments are described with acetyl-labeled acetylthiocholines (CL3COSCH2CH2N+Me3; 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 β -deuterium secondary isotope effects for turnover of acetylthiocholine depended on substrate concentration, and gave the following: D3kcat/Km = 0.95 ± 0.03, D3kcat = 1.12 ± 0.02 and D3 β kcat = 0.97 ± 0.04. The inverse isotope effect on kcat/Km is consistent with conversion of the sp2 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 kcat is consistent with hybridization change from sp3 toward sp2 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.

show abstract

Section: Introductionsupporting

confidence: 87%

Accumulation of Tetrahedral Intermediates in Cholinesterase Catalysis: A Secondary Isotope Effect Study

et al. 2010

Self Cite

View full text Add to dashboard Cite

show abstract

“…AChE is subjected to substrate inhibition at high ACTI concentration (Stojan, Brochier, Alies, Colletier, & Fournier, 2004). This can be explained by the enzyme structure as a complex ellipsoid shape protein with a gouge 20 Å deep and 5 Å wide buried deep into the protein shape that contains the substrate recognition and catalytic sites (Silman & Sussman, 2008;Dvir et al, 2010).…”

Section: Assay Of Acetylcholine Esterase (Ache) Activitymentioning

confidence: 99%

Screening Flavonoids for Inhibition of Acetylcholinesterase Identified Baicalein as the Most Potent Inhibitor

Balkis¹,

Tran²,

Lee³

et al. 2015

JAS

View full text Add to dashboard Cite

Screening phenolic and polyphenolic compounds for inhibitory activity against electric eels acetylcholinesterase (AChE) identified baicalein, a major flavone derived from the roots of Scutellaria baicalensis, as the most potent inhibitor with IC50 (concentration required for 50% inhibition) of 0.61 µM. None of the hydroxybenzoic and hydroxycinnamic acids screened showed inhibitory activity measured at 100 µM. Structure-activity relationships based on IC50 values of the active flavonoids showed that inhibitory activity (a) required the unsaturated 2-phenyl-chroman structure, (b) has strong requirement for the A-ring A5-OH, A6-OH and A7-OH groups (b) does not depend on B-ring hydroxyl groups, and (d) was reduced by bulky sugar substitution of the saturated C-ring C3-OH. Enzyme kinetic analysis showed that baicalein is a mixed inhibitor of AChE with K 1 (equilibrium constant of dissociation of the inhibitor bound enzyme complex) and K 2 (equilibrium constant of dissociation of the inhibitor bound enzyme-substrate complex) of 0.91 and 1.98 µM, respectively.

show abstract

“…Thus k 11 > k 5 and k 10 > k 4 . All this information can be easily drawn by ENZO as the comprehensive reaction scheme shown in Figure 8 [15] and subsequently the experimental data can be evaluated by fitting the parameters of the corresponding differential equations.…”

Section: Resultsmentioning

confidence: 99%

ENZO: A Web Tool for Derivation and Evaluation of Kinetic Models of Enzyme Catalyzed Reactions

et al. 2011

Self Cite

View full text Add to dashboard Cite

We describe a web tool ENZO (Enzyme Kinetics), a graphical interface for building kinetic models of enzyme catalyzed reactions. ENZO automatically generates the corresponding differential equations from a stipulated enzyme reaction scheme. These differential equations are processed by a numerical solver and a regression algorithm which fits the coefficients of differential equations to experimentally observed time course curves. ENZO allows rapid evaluation of rival reaction schemes and can be used for routine tests in enzyme kinetics. It is freely available as a web tool, at http://enzo.cmm.ki.si.

show abstract

Inhibition of Drosophila melanogaster acetylcholinesterase by high concentrations of substrate

Cited by 42 publications

References 52 publications

Accumulation of Tetrahedral Intermediates in Cholinesterase Catalysis: A Secondary Isotope Effect Study

Accumulation of Tetrahedral Intermediates in Cholinesterase Catalysis: A Secondary Isotope Effect Study

Screening Flavonoids for Inhibition of Acetylcholinesterase Identified Baicalein as the Most Potent Inhibitor

ENZO: A Web Tool for Derivation and Evaluation of Kinetic Models of Enzyme Catalyzed Reactions

Contact Info

Product

Resources

About