Magnesium-dependent ATPase Activity and Cooperativity of Magnesium Chelatase from Synechocystis sp. PCC6803*
Magnesium-dependent ATPase Activity and Cooperativity of Magnesium Chelatase from Synechocystis sp. PCC6803*
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集胞藻镁螯合酶的镁依赖性 ATP 酶活性和协同性。
DOI：
10.1074/jbc.m400958200
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发表时间：
2004
影响因子：
4.8
通讯作者：
C. Hunter
中科院分区：
文献类型：
--
作者：
J. D. Reid;C. Hunter
叶绿素生物合成的第一个关键步骤是由镁螯合酶（一种至少具有三种底物的复合酶）、协同 Mg2+ 活化以及 ATP 水解和金属离子螯合之间的自由能耦合催化。对完整镁螯合酶的行为进行了详细的功能研究，包括镁协同性的表征以及与产生的镁卟啉相关的 ATP 消耗的化学计量。结果表明，在体外，这种催化反应需要〜15 MgATP2–的水解，并且在我们的测定条件下，螯合部分反应在能量上是不利的，ΔG°'为25–33 kJ mol–1。考虑到体内可能的代谢物浓度，这会导致螯合酶反应远离平衡。我们还确定了完整酶的稳态动力学行为，并将获得的动力学参数与单个亚基的部分反应观察到的动力学参数进行了比较。 KDIX（其中 DIX 代表次卟啉 IX）估计为 3.20 μm，KMgATP2– 为 0.45 mm。螯合的 kcat 估计为 0.8 min–1，表明在完整的螯合酶中，分离的 ChlI 亚基催化的 ATP 水解要慢得多。富镁形式的螯合酶是螯合反应更有效的催化剂；螯合酶的镁激活会增加 V 以及 MgATP2- 和 DIX 反应的特异性常数，这可能是镁触发的构象变化的结果。
The first committed step in chlorophyll biosynthesis is catalyzed by magnesium chelatase, a complex enzyme with at least three substrates, cooperative Mg2+ activation, and free energy coupling between ATP hydrolysis and metal-ion chelation. A detailed functional study of the behavior of the intact magnesium chelatase has been performed, including characterization of magnesium cooperativity and the stoichiometry of ATP consumption in relation to the magnesium porphyrin produced. It is demonstrated that, in vitro, this catalyzed reaction requires hydrolysis of ∼15 MgATP2– and that the chelation partial reaction is energetically unfavorable, under our assay conditions, with a ΔG°′ of 25–33 kJ mol–1. Given the likely metabolite concentrations in vivo, this results in the chelatase reaction operating far from equilibrium. We have also determined the steady-state kinetic behavior of the intact enzyme and have compared the kinetic parameters obtained with those observed for the partial reactions of individual subunits. KDIX (where DIX represents deuteroporphyrin IX) is estimated to be 3.20 μm, and KMgATP2– is 0.45 mm. kcat for chelation is estimated to be 0.8 min–1, suggesting that the ATP hydrolysis catalyzed by the isolated ChlI subunit is substantially slower in the intact chelatase. The magnesium-rich form of the chelatase is a more effective catalyst of the chelation reaction; magnesium activation of the chelatase increases V, as well as the specificity constant for the reaction of MgATP2– and DIX, possibly as a result of a magnesium-triggered conformational change.
影响因子：
2.9
作者：
GILL, SC;VONHIPPEL, PH
通讯作者：
VONHIPPEL, PH