Studies On Biochemical Properties And Inhibition Kinetics Of Wheat Leaf Phosphatase And Thylakoid Membrane Phosphatase

The enzymatic properties, heat stability, activation and inhibition kineticsof wheat leaf phosphatase and thylakoid membrane phosphatase were studiedusing wheat pH82-2-2, anticipating to supply theoretical base for regulatingphotosynthesis, improving phosphorus nutrition by regulating enzyme activity.The results were as follows: 1. The Km value for pNPP of leaf phosphatase was 6.554 mmol/L. Thephosphatase had a pH optimum of 6.8 and a temperature optimum of 45℃incubated for 10 min. The half-life of phosphatase at 50℃, 60℃ and 70℃were 12 min, 3.75 min and 1.8 min, respectively. 2. 1-5 mmol/LMg2+and Ca2+had effects of activation on the phosphataseand 0.1-0.5 mmol/L Zn2+, Cu2+ and Mn2+ had effects of inhibition. Methanol,ethanol and glycol inhibited the activity of leaf phosphatase. Fluoride andphosphate also inhibited the activity of leaf phosphatase and the inhibitiontype was noncompetitive and competitive according to Lineweaver-Burk plot,and the inhibition constants were 1.076 mmol/L and 2.47 mmol/L respectively. 3费美娟:小麦叶片磷酸酯酶和类囊体膜磷酸酯酶生化性质及抑制动力学研究 3. The thylakoid membrane phosphatase had a Km value for pNPP of3.914 mmol/L. It had a pH optimum of 4.6 belonging to acid phosphatase anda temperature optimum of 55℃. The half-life of phosphatase at 60℃, 70℃and 80℃ were 4.5 min, 2.8 min and 2 min respectively. The activity was lostcompletely when incubated at 80℃ for 9 min. 4. The thylakoid membrane phosphatase activity was activated by Mg2+,Ca2+and Fe2+, inhibited by Mn2+ and Cu2+. Methanol, ethanol and glycol allcould activate the activity, the activation degree of ethanol was the strongestand its activation type was uncompetitive. The activity of thylakoid membranephosphatase was inhibited by molybdate, vanadate, phosphate and fluoride,the inhibition type of them were uncompetitive, noncompetitive, competitiveand mixed, respectively. 5. The treatment of light/dark can regulate the activity of wheat thylakoidmembrane phosphatase, the activity of thylakoid membrane phosphatase underlight was higher than that of under darkness; the change trend of the activity ofwheat thylakoid membrane phosphatase were "V"and "W"form when treatedfrom light to darkness and from darkness to light. 6. The wheat leaf phosphatase and thylakoid membrane phosphatase hadsome common biochemical properties. In addition to the main pH activitypeak, there was a smaller peak; the activities were activated by Mg2+and Ca2+,and inhibited by molybdate and vanadate. All these results indicated that theremay be multiple phosphatases in wheat leaf and thylakoid membrane.