Mechanistic Reappraisal of Early Stage Photochemistry in the Light-Driven Enzyme Protochlorophyllide Oxidoreductase
Mechanistic Reappraisal of Early Stage Photochemistry in the Light-Driven Enzyme Protochlorophyllide Oxidoreductase
复制标题
光驱动酶原叶绿素内酯氧化还原酶早期光化学的机理重新评估
作者：
D. Heyes;S. Hardman;David J Mansell;J. Gardiner;N. Scrutton
光驱动酶原叶绿素氧化还原酶（POR）催化原叶绿素（Pchlide）还原为叶绿素（Chlide）。该反应是叶绿素生物合成的关键步骤。Pchlide分子内的超快光化学过程是催化所必需的，先前的研究表明，一种短寿命的激发态物质，称为I675*，是反应中的第一个催化中间体，对于捕获激发能以驱动随后的氢化物和质子转移至关重要。I675* 激发态物质的化学性质及其在催化中的作用尚不清楚。在这里，我们报告的时间分辨泵浦-探测光谱测量研究的I675* 中间POR光化学的参与。我们表明，I675* 不是唯一的POR催化的光还原Pchlide，因为它也形成在POR酶的情况下。I675* 物质仅在含有Pchlide底物和Chlide产物的样品中产生，并且其形成取决于泵浦激发波长。Pchlide和Chlide在50∶50的混合物中形成的速率和量子产率最大，这是由于Pchlide和相邻的Chlide分子之间的直接能量转移引起的，在极性溶剂甲醇中被抑制。因此，我们重新评估了Pchlide的光驱动还原中早期光化学的机制，并提出I675* 代表Pchlide-Chlide二聚体中形成的激发态物种，可能是激基缔合物。与以前的报道相反，我们得出结论，这种激发态物种没有直接的机械相关性的POR催化还原Pchlide。
The light-driven enzyme protochlorophyllide oxidoreductase (POR) catalyzes the reduction of protochlorophyllide (Pchlide) to chlorophyllide (Chlide). This reaction is a key step in the biosynthesis of chlorophyll. Ultrafast photochemical processes within the Pchlide molecule are required for catalysis and previous studies have suggested that a short-lived excited-state species, known as I675*, is the first catalytic intermediate in the reaction and is essential for capturing excitation energy to drive subsequent hydride and proton transfers. The chemical nature of the I675* excited state species and its role in catalysis are not known. Here, we report time-resolved pump-probe spectroscopy measurements to study the involvement of the I675* intermediate in POR photochemistry. We show that I675* is not unique to the POR-catalyzed photoreduction of Pchlide as it is also formed in the absence of the POR enzyme. The I675* species is only produced in samples that contain both Pchlide substrate and Chlide product and its formation is dependent on the pump excitation wavelength. The rate of formation and the quantum yield is maximized in 50∶50 mixtures of the two pigments (Pchlide and Chlide) and is caused by direct energy transfer between Pchlide and neighboring Chlide molecules, which is inhibited in the polar solvent methanol. Consequently, we have re-evaluated the mechanism for early stage photochemistry in the light-driven reduction of Pchlide and propose that I675* represents an excited state species formed in Pchlide-Chlide dimers, possibly an excimer. Contrary to previous reports, we conclude that this excited state species has no direct mechanistic relevance to the POR-catalyzed reduction of Pchlide.