Department of Chemistry and Biochemistry and Center for Bioenergy and Photosynthesis, Arizona State University, Tempe, Arizona 85287-1604, United States.
Biochemistry. 2011 Dec 27;50(51):11034-46. doi: 10.1021/bi201131r. Epub 2011 Dec 1.
In Photosystem 1 (PS1), phylloquinone (PhQ) acts as a secondary electron acceptor from chlorophyll ec(3) and also as an electron donor to the iron-sulfur cluster F(X). PS1 possesses two virtually equivalent branches of electron transfer (ET) cofactors from P(700) to F(X), and the lifetime of the semiquinone intermediate displays biphasic kinetics, reflecting ET along the two different branches. PhQ in PS1 serves only as an intermediate in ET and is not normally fully reduced to the quinol form. This is in contrast to PS2, in which plastoquinone (PQ) is doubly reduced to plastoquinol (PQH(2)) as the terminal electron acceptor. We purified PS1 particles from the menD1 mutant of Chlamydomonas reinhardtii that cannot synthesize PhQ, resulting in replacement of PhQ by PQ in the quinone-binding pocket. The magnitude of the stable flash-induced P(700)(+) signal of menD1 PS1, but not wild-type PS1, decreased during a train of laser flashes, as it was replaced by a ~30 ns back-reaction from the preceding radical pair (P(700)(+)A(0)(-)). We show that this process of photoinactivation is due to double reduction of PQ in the menD1 PS1 and have characterized the process. It is accelerated at lower pH, consistent with a rate-limiting protonation step. Moreover, a point mutation (PsaA-L722T) in the PhQ(A) site that accelerates ET to F(X) ~2-fold, likely by weakening the sole H-bond to PhQ(A), also accelerates the photoinactivation process. The addition of exogenous PhQ can restore activity to photoinactivated PS1 and confer resistance to further photoinactivation. This process also occurs with PS1 purified from the menB PhQ biosynthesis mutant of Synechocystis PCC 6803, demonstrating that it is a general phenomenon in both prokaryotic and eukaryotic PS1.
在光系统 1（PS1）中，叶绿醌（PhQ）作为叶绿素 ec(3)的二次电子受体起作用，同时也是铁硫簇 F(X)的电子供体。PS1 具有来自 P(700)到 F(X)的两条几乎等效的电子转移（ET）辅助因子分支，半醌中间态的寿命呈现出两相动力学，反映了两条不同分支的 ET。PS1 中的 PhQ 仅作为 ET 的中间物，通常不会完全还原为醌形式。这与 PS2 形成对比，在 PS2 中，质体醌（PQ）被双重还原为质体氢醌（PQH(2))作为末端电子受体。我们从不能合成 PhQ 的莱茵衣藻 menD1 突变体中纯化 PS1 颗粒，导致 PhQ 在醌结合口袋中被 PQ 取代。menD1 PS1 的稳定闪光诱导的 P(700)(+)信号的幅度，而不是野生型 PS1，在激光闪光的一系列过程中减小，因为它被来自前一个自由基对（P(700)(+)A(0)(-))的30 ns 反向反应所取代。我们表明，这个光失活过程是由于 menD1 PS1 中 PQ 的双重还原引起的，并对该过程进行了表征。在较低的 pH 值下，它会加速，这与限速质子化步骤一致。此外，在 PhQ(A)位的点突变（PsaA-L722T），通过削弱与 PhQ(A)的唯一氢键，将 ET 加速到 F(X)2 倍，也加速了光失活过程。添加外源性 PhQ 可以恢复光失活 PS1 的活性并赋予其对进一步光失活的抗性。这个过程也发生在从集胞藻 PCC 6803 的 menB PhQ 生物合成突变体中纯化的 PS1 中，表明它是原核和真核 PS1 中的普遍现象。
