Acetylcholinesterase. Two types of inhibition by an organophosphorus compound: one the formation of phosphorylated enzyme and the other analogous to inhibition by substrate

Aldridge, W. N.; Reiner, Elsa

doi:10.1042/bj1150147

Cited by 76 publications

(35 citation statements)

References 16 publications

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“…The immediate inhibition seen at zero-time can be reversed by either dilution of the enzyme-inhibitor complex or by se-questering free melarsen oxide as melarsoprol by the addition of dimercaptopropanol. This type of inhibition has been observed previously for inactivation of acetylcholinesterase by haloxon where it was proposed that the inhibitor chemically combined with two-independent sites on the enzyme without prior formation of an appreciable amount of non-covalent Michaelis-type complex [26]. In contrast, time-dependent inactivation of acetylcholinesterase by esters of methane sulphonic acid involves the initial formation of reversible noncovalent complex prior to sulphonylation of the active-site residues [25].…”

Section: Discussionsupporting

confidence: 53%

“…The fact that the first-order lines do not pass through the origin (100% activity) is unusual, but has been reported previously with certain inhibitors of acetycholinesterase [26] where it is proposed that the inhibitor can interact with two independent sites on the enzyme without the prior formation of a Michaelis-type enzyme/inhibitor complex. A characteristic of this type of inhibition is that the net time-dependent rates of inhibition, koha, are independent of the final inhibitor concentration.…”

Section: Resultsmentioning

confidence: 81%

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Mechanism of inhibition of trypanothione reductase and glutathione reductase by trivalent organic arsenicals

Cunningham

Zvelebil

Fairlamb

1994

European Journal of Biochemistry

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The dithiol trypanothione, novel to trypanosomatids and analogous to glutathione in mammalian systems, has been shown to interact with anti-trypanocidal trivalent arsenical drugs forming a stable adduct, MelT. This adduct is a competitive inhibitor of the flavoprotein trypanothione reductase, responsible for maintaining intracellular trypanothione in the reduced form. Since trypanothione reductase and the analogous glutathione reductase both contain catalytically active sulphydryl groups we have examined the ability of several arsenicals to differentially inhibit these enzymes. Melarsen oxide [p-(4,6-diamino-s-triazin-2-yl)aminophenyl~senoxide] potently inhibits both enzymes in two stages, the first being essentially complete within 1 min, the second being time dependent, exhibiting saturable pseudo-first-order kinetics with k,,,, of 14.3Xs-' and 1.06X s-'and K, of 17.2 pM and 9.6 pM for trypanothione reductase and glutathione reductase, respectively.Inhibition requires prior reduction of the enzyme by NADPH and can be reversed by excess dithiols or prevented by MelT in the case of trypanothione reductase. In both cases a time-dependent loss of the characteristic charge-transfer absorbance band at 530 nm is observed upon addition of arsenical to pre-reduced enzyme, which with excess NADPH leads to a spectrum resembling the EH, form and is accompanied by an increased ability to reduce molecular oxygen. A model for inhibition is proposed where, first, free arsenical and previously reduced enzyme immediately establish an equilibrium with an inactive monothioarsane enzyme-inhibitor complex involving the interchange cysteine distal to the FAD; second, a subsequent rearrangement about the sulphur-arsenic bond leads to the binding of the arsenical to the charge-transfer cysteine, proximal to the FAD, forming a more stable dithioarsane complex. Molecular modelling suggests that the differences in kinetic behaviour of the two enzymes can be attributed to structural features of their respective disulphide-binding sites. Incubation of reduced trypanothione reductase with excess dihydrotrypanothione and melarsen oxide prevents direct inhibition of the enzyme, suggesting that dihydrotrypanothione acts as a protectant in vivo, preventing the direct modification of trypanothione reductase by sequestering the arsenical as MelT.The trivalent arsenical drug, melarsoprol [l], is still extensively used in the treatment of human African trypanosomiasis, caused by the parasitic protozoa Trypanosoma brucei rhodesiense and 7: brucei gambiense. The disease derives its common name of sleeping sickness from the sleep disturbances and other neurological disorders that result from the second stage of the disease, when parasitic invasion of the central nervous system has occurred. If left untreated the disease is inevitably fatal. Use of melarsoprol is usually restricted to the second stage of the disease when the alterna-

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Section: Discussionsupporting

confidence: 53%

Section: Resultsmentioning

confidence: 81%

Mechanism of inhibition of trypanothione reductase and glutathione reductase by trivalent organic arsenicals

Cunningham

Zvelebil

Fairlamb

1994

European Journal of Biochemistry

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“…A solution for this problem would be to have ACHe be inhibited by its substrate ACH so that when ACH is at high concentrations the degradation proceeds relatively slowly and then accelerates as the concentration drops. Substrate inhibition of ACHe by ACH was noticed as long ago as 1969 [31], but its functional importance has been emphasized only recently [28,32]. For this scenario to work, ACH should be released very rapidly into the cleft so that the concentration of ACH rises quickly into the inhibitory range.…”

Section: C Reed Et Almentioning

confidence: 99%

The biological significance of substrate inhibition: A mechanism with diverse functions

2010

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Many enzymes are inhibited by their own substrates, leading to velocity curves that rise to a maximum and then descend as the substrate concentration increases. Substrate inhibition is often regarded as a biochemical oddity and experimental annoyance. We show, using several case studies, that substrate inhibition often has important biological functions. In each case we discuss, the biological significance is different. Substrate inhibition of tyrosine hydroxylase results in a steady synthesis of dopamine despite large fluctuations in tyrosine due to meals. Substrate inhibition of acetylcholinesterase enhances the neural signal and allows rapid signal termination. Substrate inhibition of phosphofructokinase ensures that resources are not devoted to manufacturing ATP when it is plentiful. In folate metabolism, substrate inhibition maintains reactions rates in the face of substantial folate deprivation. Substrate inhibition of DNA methyltransferase serves to faithfully copy DNA methylation patterns when cells divide while preventing de novo methylation of methyl-free promoter regions. Keywords:.b iological function; enzyme kinetics; substrate inhibitionThe kinetics of an enzymatic reaction are typically studied by varying the concentration of substrate and plotting the rate of product formation as a function of substrate concentration. In the conventional case this yields a typical hyperbolic Michaelis-Menten curve, and a linear reciprocal LineweaverBurk plot, from which the kinetic constants of the enzyme can be calculated. A surprisingly large number of enzymes do not behave in this conventional way. Instead, their velocity curves rise to a maximum and then decline as the substrate concentration goes up. This phenomenon is referred to as substrate inhibition, and it is estimated that it occurs in some 20% of enzymes [1]. A partial list of enzymes that show substrate inhibition appears in Box 1.Substrate inhibition is often interpreted as an abnormality that comes from using artificially high substrate concentration in a laboratory setting. In a review article on the mechanisms of substrate inhibition in 1994, Kuehl [2] commented that ''although recognized early on as an almost universal phenomenon, it has nevertheless met an almost universal disinterest. Probably the main reason for this neglect is that the majority of enzymologists and many authorities in the field regard substrate inhibition as being almost always a nonphysiological phenomenon.'' There are several reasons for suspecting that substrate inhibition is not a pathological phenomenon, but a biologically relevant regulatory mechanism. First, in many cases normal substrate concentrations are to the right of the velocity maximum, which indicates that these enzymes typically operate under substrate inhibition. Second, many enzymes have specialized sites where a second substrate molecule can bind and act as an allosteric inhibitor. For those enzymes, substrate inhibition is clearly a specially evolved property. Third, evidence is accumulating that substr...

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“…Kinetics of cationic substrate hydrolysis catalyzed by AChE deviate from Michaelis-Menten kinetics (21,22). Cationic sub-strates, including ACh, inhibit their own catalysis at concentrations exceeding the K m (Ն1 mM, i.e.…”

Section: Residue Trpmentioning

confidence: 99%

Substrate and Product Trafficking through the Active Center Gorge of Acetylcholinesterase Analyzed by Crystallography and Equilibrium Binding

Bourne

Radić

Sulzenbacher

et al. 2006

Journal of Biological Chemistry

123

139

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The principal role of acetylcholinesterase (AChE) 3 at cholinergic synapses is to terminate neurotransmission by fast hydrolysis of the substrate, acetylcholine (ACh) (1, 2). The AChE active center, containing the catalytic triad, Glu 334 -His 447 -Ser 203 in mammals (3), is located centrosymmetric to the subunit and at the base of a deep and narrow gorge (4, 5).AChE-catalyzed hydrolysis of ACh and other carboxyl esters proceeds via formation of an initial noncovalent enzyme-substrate complex.

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Acetylcholinesterase. Two types of inhibition by an organophosphorus compound: one the formation of phosphorylated enzyme and the other analogous to inhibition by substrate

Cited by 76 publications

References 16 publications

Mechanism of inhibition of trypanothione reductase and glutathione reductase by trivalent organic arsenicals

Mechanism of inhibition of trypanothione reductase and glutathione reductase by trivalent organic arsenicals

The biological significance of substrate inhibition: A mechanism with diverse functions

Substrate and Product Trafficking through the Active Center Gorge of Acetylcholinesterase Analyzed by Crystallography and Equilibrium Binding

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