Torres J C, Guixé V, Babul J
Departamento de Biología, Facultad de Ciencias, Universidad de Chile, Santiago.
Arch Biochem Biophys. 1995 Aug 20;321(2):517-25. doi: 10.1006/abbi.1995.1425.
A method for assessing rates of the futile cycle is presented, and it is illustrated in vitro. Glycolytic- and gluconeogenic-type futile cycles are simulated for the reactions catalyzed by phosphofructokinase (EC 2.7.1.11) and fructose-1,6-bisphosphatase (EC 3.1.3.11) in assays systems in which the cycle rates in either direction can be varied and determined. While either system is sustaining a net flux of carbons in a given direction, the flux of radioactively labeled carbons in the opposite direction is determined. Different cycle rates are obtained by varying phosphofructokinase activity while keeping fructose-1,6-bisphosphatase activity constant in the "gluconeogenic" simulation and varying fructose-1,6-bisphosphatase while keeping phosphofructokinase activity constant in the "glycolytic" simulation. A direct, linear relationship was found between the cycle rate and the radioactive labeling of fructose 1,6-bisphosphate from [U-14C]glucose 6-phosphate during net gluconeogenic carbon flux. Also, a direct, linear relationship was found between cycle rate and radioactive labeling of fructose-6-P from [U-14C]fructose-1,6-bisP during net glycolytic carbon flux. The applicability, advantages, and problems of the method are discussed.
本文介绍了一种评估无效循环速率的方法，并在体外进行了说明。在测定系统中，模拟了由磷酸果糖激酶（EC 2.7.1.11）和果糖-1,6-二磷酸酶（EC 3.1.3.11）催化的糖酵解型和糖异生型无效循环，在该系统中，循环速率在任一方向上均可变化并测定。当任一系统在给定方向上维持碳的净通量时，测定相反方向上放射性标记碳的通量。在“糖异生”模拟中，通过改变磷酸果糖激酶活性同时保持果糖-1,6-二磷酸酶活性恒定，以及在“糖酵解”模拟中，通过改变果糖-1,6-二磷酸酶同时保持磷酸果糖激酶活性恒定，可获得不同的循环速率。发现在净糖异生碳通量期间，循环速率与来自[U-14C]葡萄糖6-磷酸的果糖1,6-二磷酸的放射性标记之间存在直接的线性关系。此外，发现在净糖酵解碳通量期间，循环速率与来自[U-14C]果糖-1,6-二磷酸的果糖-6-磷酸的放射性标记之间存在直接的线性关系。讨论了该方法的适用性、优点和问题。
