Delocalized hole transport coupled to sub-ns tryptophanyl deprotonation promotes photoreduction of class II photolyases
Delocalized hole transport coupled to sub-ns tryptophanyl deprotonation promotes photoreduction of class II photolyases
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DOI：
10.1039/c8cp04548h
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发表时间：
2018-10-21
影响因子：
3.3
通讯作者：
Essen, Lars-Oliver
中科院分区：
文献类型：
--
作者：
Lacombat, Fabien;Espagne, Agathe;Essen, Lars-Oliver
II类光裂合酶利用与大多数其他光裂合酶和隐花色素完全不同的色氨酸三联体来光还原其黄素辅因子（FAD）。为了对抗加速的反向电子转移，他们进化出了一种异常快速的去质子化的远端羟烷基自由基阳离子（WH+），这是激发黄素后产生的。我们研究了氧化FAD光还原的超快瞬态吸收光谱的主要方面，使用II类光解酶从甲烷八叠球菌Mazei。随着9.2 ps的时间常数，初始还原步骤的激发黄素由近端W-381色氨酸进行几乎20倍慢于其他光解酶携带氧化FAD，最有可能是因为较大的距离之间的黄素和近端色氨酸。由此形成的W381 H+自由基被跟踪的瞬态各向异性测量迁移在29 ps与离域超过几个成员的色氨酸三联体。这29 ps阶段还包括一小部分激发黄素的衰变，在较慢的时间尺度上反应，和FAD（-）/WH+自由基对的部分重组。在230 ps的最后一个动力学阶段被分配到去质子化的W388 H+发生在竞争与部分电荷重组。有趣的是，我们通过与Y345 F突变体的比较表明，这最后一个阶段还涉及通过W388 H+氧化Y-345酚基，产生少量的中性酪氨酰自由基（YO）。该电子转移步骤的速率比去质子化的W-388自由基对Y-345的相应氧化快约6个数量级。与传统的光解酶，电子空穴积累在远端色氨酸之前慢得多的dichanyl去质子化，我们的数据表明，离域空穴传输伴随着由W388 H+的超快去质子化结束。
Class II photolyases utilize for the photoreduction of their flavin cofactor (FAD) a completely different tryptophan triad than most other photolyases and cryptochromes. To counter sped-up back electron transfer, they evolved an unusually fast deprotonation of the distal tryptophanyl radical cation (WH+) that is produced after excitation of the flavin. We studied the primary aspects of oxidized FAD photoreduction by ultrafast transient absorption spectroscopy, using the class II photolyase from Methanosarcina mazei. With a time constant of 9.2 ps, the initial reduction step of the excited flavin by the proximal W-381 tryptophan proceeds almost twentyfold slower than in other photolyases carrying oxidized FAD, most likely because of the larger distance between the flavin and the proximal tryptophan. The thus formed W381H+ radical is tracked by transient anisotropy measurements to migrate in 29 ps with delocalization over several members of the tryptophan triad. This 29 ps phase also includes the decay of a small fraction of excited flavin, reacting on a slower timescale, and partial recombination of the FAD(-)/WH+ radical pair. A final kinetic phase in 230 ps is assigned to the deprotonation of W388H+ that occurs in competition with partial charge recombination. Interestingly, we show by comparison with the Y345F mutant that this last phase additionally involves oxidation of the Y-345 phenolic group by W388H+, producing a small amount of neutral tyrosyl radical (YO). The rate of this electron transfer step is about six orders of magnitude faster than the corresponding oxidation of Y-345 by the deprotonated W-388 radical. Unlike conventional photolyases, where the electron hole accumulates on the distal tryptophan before the much slower tryptophanyl deprotonation, our data show that delocalized hole transport is concomitantly concluded by ultrafast deprotonation of W388H+.