Inhibition of Drosophila melanogaster acetylcholinesterase by high concentrations of substrate.

Jure Stojan, Laure Brochier, Carole Alies, Jacques Philippe Colletier, Didier Fournier
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引用次数: 42

Abstract

Acetylcholine hydrolysis by acetylcholinesterase is inhibited at high substrate concentrations. To determine the residues involved in this phenomenon, we have mutated most of the residues lining the active-site gorge but mutating these did not completely eliminate hydrolysis. Thus, we analyzed the effect of a nonhydrolysable substrate analogue on substrate hydrolysis and on reactivation of an analogue of the acetylenzyme. Analyses of various models led us to propose the following sequence of events: the substrate initially binds at the rim of the active-site gorge and then slides down to the bottom of the gorge where it is hydrolyzed. Another substrate molecule can bind to the peripheral site: (a) when the choline is still inside the gorge - it will thereby hinder its exit; (b) after choline has dissociated but before deacetylation occurs - binding at the peripheral site increases deacetylation rate but (c) if a substrate molecule bound to the peripheral site slides down to the bottom of the active-site before the catalytic serine is deacetylated, its new position will prevent the approach of water, thus blocking deacetylation.

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高浓度底物对黑腹果蝇乙酰胆碱酯酶的抑制作用。
在高底物浓度下,乙酰胆碱酯酶对乙酰胆碱的水解受到抑制。为了确定参与这一现象的残基,我们对活性位点峡谷的大部分残基进行了突变,但这些突变并没有完全消除水解。因此,我们分析了不可水解的底物类似物对底物水解和乙酰酶类似物再激活的影响。对各种模型的分析使我们提出了以下事件序列:底物最初在活性位点峡谷边缘结合,然后滑到峡谷底部水解。另一种底物分子可以与外周位点结合:(a)当胆碱仍在峡谷内时,它将因此阻碍其退出;(b)在胆碱解离后,但在去乙酰化发生之前,外周位点的结合增加了去乙酰化速率,但(c)如果在催化丝氨酸去乙酰化之前,与外周位点结合的底物分子滑到活性位点的底部,它的新位置将阻止水的接近,从而阻断去乙酰化。
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