Department of Biochemistry, Faculty of Science, Charles University, Hlavova 2030, Prague 2 128 40, Czech Republic.
Biochimie. 2010 Oct;92(10):1362-70. doi: 10.1016/j.biochi.2010.06.019. Epub 2010 Jun 30.
Phosphoenolpyruvate carboxylase (PEPC, EC 4.1.1.31) from mature maize seeds (Zea mays L.) was purified to homogeneity and a final specific activity of 13.3 μmol min⁻¹ mg⁻¹. Purified PEPC was treated with phosphatase from bovine intestinal mucosa or protein kinase A to study its apparent phosphorylation level. Kinetic parameters of the enzyme reaction catalyzed by phosphorylated and dephosphorylated forms under different conditions were compared, as well as an effect of modulators. The enzyme dephosphorylation resulted in the change of hyperbolic kinetics to the sigmoidal one (with respect to PEP), following with the decrease of maximal reaction rate and the increase of sensitivity to L-malate inhibition. The hyperbolic kinetics of native PEPC present in dry maize seeds was not changed after the protein kinase A treatment, while it was converted to the sigmoidal one after dephosphorylation. Level of PEPC phosphorylation was not affected during seed imbibition.
从成熟的玉米种子（Zea mays L.）中纯化得到磷酸烯醇式丙酮酸羧化酶（PEPC，EC 4.1.1.31），并达到了纯品的水平，最终比活性为 13.3 μmol min⁻¹ mg⁻¹。用来自牛肠黏膜的磷酸酶或蛋白激酶 A 处理纯化的 PEPC，以研究其明显的磷酸化水平。比较了不同条件下磷酸化和去磷酸化形式催化的酶反应的动力学参数，以及调节剂的影响。酶的去磷酸化导致酶促反应动力学从双曲线型转变为（相对于 PEP）的 S 型，最大反应速率降低，对 L-苹果酸抑制的敏感性增加。经过蛋白激酶 A 处理后，存在于干燥玉米种子中的天然 PEPC 的双曲线型动力学没有改变，而在去磷酸化后则转变为 S 型。在种子吸胀过程中，PEPC 的磷酸化水平没有受到影响。
