不同生境两种生态型芦苇叶片质膜H+-ATPase的比较

利用两相法化纯化质膜微囊,研究了分布西北沙地区的两种生态型芦苇(Phragmites communis trih.)水生芦苇和重度盐化草甸芦苇,分别简称为水芒和盐芦)叶片质膜H - ATPase的部分性质.结果显示,与水芦相比,盐芦质膜H -ATPase的ATP水解活性升高,Km值由1.27mmoll降至Vmax没有显著差异.并且该酶活性对温度的敏感必玫PH谱型也发生了变化.以对硝基苯磷酸盐为底物,低浓度时盐芦的的质膜H -ATPase水解活性有差异.钡酸盐抑制实验表明,两种生态的质膜H -ATPase磷酸-酶区的催化性质不同.胰酶对质膜H -ATPase活性的活化谱型也存在差异,说明该酶C末端的结构或性质发生了变化.此外,与水芦相比,盐芦质膜H -ATPase的质子泵活性的耦联程度也升高了.以上结果明,当芦苇从水生环境向盐渍环境过渡时,质膜H -ATPase的催化性质发生了变化,这些变化可能是由酶结构的修饰和不同的同工酬酶谱引起的.H -ATPase催化性质的变化可能是对盐渍生境的适应性反应.

Comparison of Plasma Membrane H+-ATPase Activity in Two Ecotypes of Reed (Phragmites communis) Leaves from Different Habitats

Plasma membrane (PM) vesicles of the leaves of two ecotypes of reed (Phragmites communis Trin.), swamp reed (SR) and heavy salt meadow reed (HSMR) growing in the desert region of Northwest China, were purified by two-phase partitioning and the properties of their PM H⁺-ATPases (EC 3.6.1.35) were compared. The specific activity of this enzyme was greater in HSMR than in SR and the K_m lower (1.27 mmol/L in SR and 0.30 mmol/L in HSMR), and the V_max of ATP hydrolysis activity showed no significant difference between the two ecotypes. The PM H⁺-ATPase was more sensitive to denaturing temperatures in HSMR than in SR, and the pH profile also showed a slight difference, suggesting that the catalytic mechanism of this enzyme was different in HSMR compared with that in SR. The p -nitrophenyl phosphate (PNPP) hydrolysis activity of H⁺-ATPase was higher in HSMR than in SR at low concentrations of PNPP, but showed no difference at high PNPP concentration. The K_m for PNPP hydrolysis was 3.61 mmol/L and 1.92 mmol/L in SR and HSMR, respectively. And the V_max of PNPP hydrolysis showed no significant difference in the two reed ecotypes. An experiment with the inhibitor vanadate showed that the catalytic mechanisms of the phosphatase domain of the ATPase were different in the two ecotypes. The data obtained following trypsin treatment showed a difference in the enzyme activity pattern, suggesting that there existed a possible change in the C-terminus of the ATPase, either in the structure or in the property or both of them. In addition, compared with SR, the ATP-dependent H⁺ pumping activity of ATPase and the coupling between proton transport and ATP hydrolysis in HSMR were increased. These results indicated that the properties of PM H⁺-ATPase were changed in HSMR with compared to those in SR, which might include enzyme modifications and different isoforms expressed. The alterations of the properties of this enzyme might be an adaptive response to the habitat.