Detheux M, Vandercammen A, Van Schaftingen E
Laboratoire de Chimie Physiologique, International Institute of Cellular and Molecular Pathology, Brussels, Belgium.
Eur J Biochem. 1991 Sep 1;200(2):553-61. doi: 10.1111/j.1432-1033.1991.tb16218.x.
In the absence of fructose 6-phosphate, the regulatory protein of rat liver glucokinase (hexokinase IV or D) inhibited this enzyme, though with a much (15-fold) lower potency than in the presence of a saturating concentration of fructose 6-phosphate. Evidence is provided that this inhibition is not due to contaminating fructose 6-phosphate. In the presence of regulatory protein, sorbitol 6-phosphate, a potent analog of fructose 6-phosphate, exerted a hyperbolic, partial inhibition on glucokinase, the degree of which increased with the concentration of regulatory protein. Plots of the reciprocal of the difference between the rates in the absence and in the presence of sorbitol 6-phosphate versus 1/[sorbitol 6-phosphate] at various concentrations of regulatory protein were linear, and demonstrated that the apparent affinity for sorbitol 6-phosphate increased with the concentration of regulatory protein. Plots of the reciprocal of the difference between 1/v in the presence and in the absence of sorbitol 6-phosphate versus 1/[sorbitol 6-phosphate] were also linear and crossed the axis at a value independent of the concentration of regulatory protein. Fructose 1-phosphate released the inhibition exerted by the regulatory protein in a hyperbolic fashion. The concentration of this effector required for a half-maximal effect increased linearly with the concentrations of sorbitol 6-phosphate and of regulatory protein. These results are consistent with a model in which the regulatory protein exists under two conformations, one form which binds inhibitors and glucokinase, and the other which binds activators, although not glucokinase. Sorbitol 6-phosphate, 2-deoxysorbitol 6-phosphate and mannitol 1-phosphate, all analogs of the open-chain configuration of fructose 6-phosphate, inhibited glucokinase in the presence of regulatory protein at lower concentrations than fructose 6-phosphate, whereas fixed analogs of the furanose form of fructose 6-phosphate were inactive or behaved as activators. This indicated that fructose 6-phosphate in its open-chain configuration is recognized by the regulatory protein. A series of compounds exerted an activating effect. These included, in order of decreasing potency: fructose 1-phosphate, psicose 1-phosphate, ribitol 5-phosphate, analogs of fructose 1-phosphate and of ribitol 5-phosphate and, at much higher concentrations, inorganic phosphate.
在缺乏6-磷酸果糖的情况下，大鼠肝脏葡萄糖激酶（己糖激酶IV或D）的调节蛋白会抑制这种酶，尽管其效力比在饱和浓度的6-磷酸果糖存在时低得多（15倍）。有证据表明这种抑制并非由于污染的6-磷酸果糖所致。在调节蛋白存在的情况下，6-磷酸山梨醇是6-磷酸果糖的一种有效类似物，对葡萄糖激酶产生双曲线型的部分抑制作用，其抑制程度随调节蛋白浓度的增加而增加。在不同浓度的调节蛋白下，绘制6-磷酸山梨醇存在与不存在时速率差值的倒数对1/[6-磷酸山梨醇]的曲线是线性关系，表明对6-磷酸山梨醇的表观亲和力随调节蛋白浓度的增加而增加。绘制存在与不存在6-磷酸山梨醇时1/v差值的倒数对1/[6-磷酸山梨醇]的曲线也是线性的，且与横轴的交点值与调节蛋白浓度无关。1-磷酸果糖以双曲线方式解除调节蛋白施加的抑制作用。产生半最大效应所需的这种效应物浓度随6-磷酸山梨醇和调节蛋白浓度的增加而线性增加。这些结果与一种模型一致，即调节蛋白存在两种构象，一种构象结合抑制剂和葡萄糖激酶，另一种构象结合激活剂，但不结合葡萄糖激酶。6-磷酸山梨醇、2-脱氧-6-磷酸山梨醇和1-磷酸甘露醇，均为6-磷酸果糖开链构型的类似物，在调节蛋白存在时，它们抑制葡萄糖激酶的浓度低于6-磷酸果糖，而6-磷酸果糖呋喃糖形式的固定类似物则无活性或表现为激活剂。这表明调节蛋白识别开链构型的6-磷酸果糖。一系列化合物发挥激活作用。按效力递减顺序包括：1-磷酸果糖、1-磷酸阿洛酮糖、5-磷酸核糖醇、1-磷酸果糖和5-磷酸核糖醇的类似物，以及在高得多的浓度下的无机磷酸。
