Conformational Heterogeneity and Propagation of Structural Changes in the LOV2/Jα Domain from Avena sativa Phototropin 1 as Recorded by Temperature-Dependent FTIR Spectroscopy
Conformational Heterogeneity and Propagation of Structural Changes in the LOV2/Jα Domain from Avena sativa Phototropin 1 as Recorded by Temperature-Dependent FTIR Spectroscopy
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DOI：
10.1016/j.bpj.2009.03.047
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发表时间：
2009-07-08
影响因子：
3.4
通讯作者：
Kennis, John T. M.
中科院分区：
文献类型：
--
作者：
Alexandre, Maxime T. A.;van Grondelle, Rienk;Kennis, John T. M.
趋光素控制植物的向光性、叶绿体运动、气孔开放和叶片扩张。Phototropin 1 （phot1）由一个激酶结构域组成，该结构域与两个结合黄素单核苷酸的蓝光感应结构域LOV2和LOV1相连。LOV2结构域和一个名为J α的螺旋片段之间的相互作用被破坏，将LOV连接到激酶结构域，诱导随后的趋光蛋白1激酶活性和进一步的下游信号转导。本文研究了温度和水合作用对Avena sativa (AsLOV2/J α) phot1 LOV2域中光触发信号传播的影响，利用傅里叶变换红外光谱揭示了趋光素1的部分分子机制。我们报道AsLOV2/J α在酰胺I和酰胺II区显示出强烈的信号，主要是由于β -片的改变和Ja螺旋从Per-ARNT-Sim核上的解结合及其随后的部分展开引起的。重要的是，这些结构变化只发生在完全水化和温度高于280 K的条件下。我们表征了一种新分离的低水合中间体，它显示出高频酰胺I信号的下移，可能对应于环收紧，没有大的β -片或Ja结构变化。此外，我们报道了AsLOV2/J α的异质性，涉及两个不同的C(4)=O构象种群，共存于暗态，并以C(4)=O羰基频率在1712 cm（-1）和1694 cm-1为特征，分别可归因为该位点的单个氢键和两个氢键。这些构象显示出轻微的吸收光谱偏移，并在低温下引起LOV域紫外/可见光谱中475 nm波段的分裂。
Phototropins control phototropism, chloroplast movement, stomatal opening, and leaf expansion in plants. Phototropin 1 (phot1) is composed of a kinase domain linked to two blue light-sensing domains, LOV2 and LOV1, which bind flavin mononucleotide. Disruption of the interaction between the LOV2 domain and a helical segment named J alpha, joining LOV to the kinase domain, induces the subsequent kinase activity of phototropin 1 and further-downstream signal transduction. Here we study the effects of temperature and hydration on the light-triggered signal propagation in the phot1 LOV2 domain of Avena sativa (AsLOV2/J alpha), using Fourier transform infrared spectroscopy to unravel part of the molecular mechanism of phototropin 1. We report that AsLOV2/J alpha shows an intense signal in the amide I and II regions, arising mainly from beta-sheet changes and the unbinding of the Ja helix from the Per-ARNT-Sim core and its subsequent partial unfolding. Importantly, these structural changes only occur under conditions of full hydration and at temperatures above 280 K. We characterized a newly isolated low-hydration intermediate that shows a downshift of high-frequency amide I signals and that possibly corresponds to loop tightening, without large beta-sheet or Ja structural changes. In addition, we report a heterogeneity in AsLOV2/J alpha involving two different C(4)=O conformer populations, coexisting in the dark state and characterized by C(4)=O carbonyl frequencies at 1712 cm(-1) and 1694 cm-1 that are attributable to a single H-bond and two H-bonds at this site, respectively. Such conformers display slightly shifted absorption spectra and cause a splitting of the 475-nm band in the ultraviolet/visible spectra of LOV domains at low temperature.