来自叶片的丙酮酸磷酸双激酶的性质及作用机制

Sugar-cane leaf pyruvate,P(i) dikinase was prepared free of enzymes that would interfere with studies on the stoicheiometry and mechanism of the reaction it catalyses. The reaction was unequivocally shown to involve the conversion of equimolar amounts of pyruvate, ATP and P(i) into phosphoenolpyruvate, AMP and PP(i). 2. The purified enzyme was stable at pH8.3 only if stored at about 20 degrees in the presence of Mg(2+) and a thiol-reducing reagent, care being taken to prevent the oxidation of the thiol. 3. The apparent Michaelis constants for phosphoenolpyruvate and PP(i) were 0.11mm and 0.04mm respectively and that for AMP was less than 4mum. 4. At pH8.3 the initial velocity of the reaction was about 6 times as fast in the direction towards phosphoenolpyruvate synthesis as in the reverse direction. 5. With the exception of ATP, all the products of the reaction in both directions were inhibitory. 6. The phosphate groups of PP(i) were derived from P(i) and from the terminal phosphate of ATP. 7. Isotope-exchange studies indicated that the reaction proceeds in the following steps:Enzyme+ATP+P(i) right harpoon over left harpoon Enzyme-P+AMP+PP(i)Enzyme-P+pyruvate right harpoon over left harpoon Enzyme+phosphoenolpyruvate

制备了甘蔗叶丙酮酸、磷酸（Pi）二激酶，使其不含会干扰对其催化反应的化学计量和机制研究的酶。明确表明该反应涉及等摩尔量的丙酮酸、ATP和磷酸（Pi）转化为磷酸烯醇丙酮酸、AMP和焦磷酸（PPi）。
纯化后的酶仅在pH8.3时稳定，前提是在镁离子（Mg2+）和一种硫醇还原试剂存在的情况下于约20摄氏度储存，要注意防止硫醇氧化。
磷酸烯醇丙酮酸和焦磷酸（PPi）的表观米氏常数分别为0.11毫摩尔和0.04毫摩尔，而AMP的表观米氏常数小于4微摩尔。
在pH8.3时，反应朝着磷酸烯醇丙酮酸合成方向的初始速度约为反向的6倍。
除了ATP外，反应在两个方向上的所有产物均具有抑制作用。
焦磷酸（PPi）的磷酸基团来自磷酸（Pi）和ATP的末端磷酸基团。
同位素交换研究表明反应按以下步骤进行：酶+ATP+磷酸（Pi）⇌酶-P+AMP+焦磷酸（PPi）；酶-P+丙酮酸⇌酶+磷酸烯醇丙酮酸

Properties and mechanism of action of pyruvate, phosphate dikinase from leaves.