Mechanism-based tuning of a LOV domain photoreceptor.

Mechanism-based tuning of a LOV domain photoreceptor.
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DOI:
10.1038/nchembio.210
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发表时间:
2009-11
影响因子:
14.8
通讯作者:
Crane, Brian R.
Crane, Brian R.
中科院分区:
生物学1区
文献类型:
--
作者:
Zoltowski, Brian D.;Vaccaro, Brian;Crane, Brian R.

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类光促黄素LOV结构域在蓝光的响应下形成半胱氨基-黄素加合物,但在输出信号和光加合物信号态的寿命方面表现出显著的变化。对慢循环真菌LOV-感光体Vivid的机理研究揭示了反应性半胱氨酸构象、黄素电子环境和溶剂可及性对加合物断裂和热回复的重要性。质子库存、pH效应、碱催化和结构研究表明,黄素N5去质子化是回收的速度决定因素。在VVD中,活性位点残基Ile74、Ile85、Met135和Met165的替换使光加合物寿命延长四个数量级以上,其他LOV蛋白的类似变化也显示出类似的影响。加合物状态衰减率还与信号所需蛋白质的构象和寡聚体性质的变化相关。这些发现将LOV结构域的自然序列变异与功能联系起来,并为设计广泛反应的光敏探针提供了一种手段。
Phototropin-like LOV domains form a cysteinyl-flavin adduct in response to blue light, but show dramatic variation in output signal and the lifetime of the photo-adduct signaling state. Mechanistic studies of the slow-cycling fungal LOV-photoreceptor Vivid reveal the importance of reactive cysteine conformation, flavin electronic environment and solvent accessibility for adduct scission and thermal reversion. Proton inventory, pH-effects, base catalysis and structural studies implicate flavin N5 deprotonation as rate-determining for recovery. Substitutions of active site residues Ile74, Ile85, Met135 and Met165 alter photoadduct lifetimes by over four orders of magnitude in VVD and similar changes in other LOV proteins show analogous effects. Adduct state decay rates also correlate with changes in conformational and oligomeric properties of the protein necessary for signaling. These findings link natural sequence variation of LOV domains to function and provide a means to design broadly reactive light-sensitive probes.
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