PASTEUR, Département de chimie , École normale supérieure, PSL University, Sorbonne Université, CNRS , 75005 Paris , France.
Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS , Univ. Paris-Sud, Université Paris-Saclay , 91198 , Gif-sur-Yvette cedex , France.
J Am Chem Soc. 2019 Aug 28;141(34):13394-13409. doi: 10.1021/jacs.9b03680. Epub 2019 Aug 19.
The animal-like cryptochrome of (aCRY) is a recently discovered photoreceptor that controls the transcriptional profile and sexual life cycle of this alga by both blue and red light. aCRY has the uncommon feature of efficient formation and longevity of the semireduced neutral form of its FAD cofactor upon blue light illumination. Tyrosine Y plays a crucial role by elongating , as fourth member, the electron transfer (ET) chain found in most other cryptochromes and DNA photolyases, which comprises a conserved tryptophan triad. Here, we report the full mechanism of light-induced FADH formation in aCRY using transient absorption spectroscopy from hundreds of femtoseconds to seconds. Electron transfer starts from ultrafast reduction of excited FAD to FAD by the proximal tryptophan (0.4 ps) and is followed by delocalized migration of the produced WH radical along the tryptophan triad (∼4 and ∼50 ps). Oxidation of Y by coupled ET to WH and deprotonation then proceeds in ∼800 ps, without any significant kinetic isotope effect, nor a pH effect between pH 6.5 and 9.0. The FAD/Y pair is formed with high quantum yield (∼60%); its intrinsic decay by recombination is slow (∼50 ms), favoring reduction of Y by extrinsic agents and protonation of FAD to form the long-lived, red-light absorbing FADH species. Possible mechanisms of tyrosine oxidation by ultrafast proton-coupled ET in aCRY, a process about 40 times faster than the archetypal tyrosine-Z oxidation in photosystem II, are discussed in detail.
(aCRY) 的动物样隐色体是最近发现的光受体，它通过蓝光和红光控制藻类的转录谱和有性生命周期。aCRY 具有一个不寻常的特征，即在蓝光照射下，其 FAD 辅因子有效地形成并长时间保持半还原的中性形式。酪氨酸 Y 通过延长其长度，作为第四个成员，加入到大多数其他隐色体和 DNA 光裂合酶中发现的电子转移 (ET) 链中，该 ET 链包含一个保守的色氨酸三联体。在这里，我们使用从几百飞秒到几秒钟的瞬态吸收光谱报告了 aCRY 中光诱导 FADH 形成的完整机制。电子转移始于激发态 FAD 被邻近色氨酸快速还原为 FAD（0.4 ps），随后产生的 WH 自由基沿着色氨酸三联体发生离域迁移（∼4 和 ∼50 ps）。然后，通过与 WH 的偶联 ET 和去质子化，Y 的氧化在∼800 ps 内进行，没有明显的动力学同位素效应，也没有在 pH 6.5 和 9.0 之间的 pH 效应。FAD/Y 对以高量子产率（∼60%）形成；其通过复合的缓慢衰减（∼50 ms），有利于外部试剂还原 Y 和质子化 FAD 以形成长寿命、吸收红光的 FADH 物种。在 aCRY 中，超快质子偶联 ET 过程中酪氨酸氧化的可能机制，这一过程比光合作用 II 中典型的酪氨酸-Z 氧化快约 40 倍，被详细讨论。
