NTP-binding properties of the blue-light receptor YtvA and effects of the E105L mutation
NTP-binding properties of the blue-light receptor YtvA and effects of the E105L mutation
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DOI：
10.1007/s00249-007-0155-1
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发表时间：
2007-09-01
影响因子：
2
通讯作者：
Losi, Aba
中科院分区：
文献类型：
--
作者：
Buttani, Valentina;Gaertner, Wolfgang;Losi, Aba
YtvA 是一种来自枯草芽孢杆菌的蓝光感应蛋白，与植物向光素相关。它携带一个 LOV（光、氧和电压）结构域，结合 FMN（黄素单核苷酸）作为发色团，以及一个功能尚不明确的 STAS（硫酸盐转运蛋白和抗因子拮抗剂）结构域。我们最近证明 YtvA 结合三磷酸核苷酸 (NTP)，并强调了 STAS 结构域和小 GTP 结合蛋白之间的结构相似性。在这项工作中，我们利用 GTP (GTP(TR)) 的荧光衍生物和诱变实验进一步研究了 YtvA 的 NTP 结合特性。主要结果如下：(a)竞争实验表明，YtvA对GTP的亲和力远高于对GDP和GMP的亲和力。 (b) 蓝光诱导的结构变化从 LOV 核心传递到 NTP 结合腔，建立可能的蛋白内信号转导途径。 (c) LOV 核心中央 β 支架 E105L 的突变会损害结合 GTP(TR) 的光驱动光谱变化。这一结果得到了圆二色性数据的支持，因为 YtvA-E105L 没有显示出表征 YtvA 的转角部分的光诱导构象变化，这意味着 E105 在功能上很重要。 (d) 在LOV-STAS复合体的结构模型中，基于对接算法，界面包括LOV核心上的I beta-H beta环，以及中央beta支架的部分。预计 E105 会与 LOV-STAS 连接区相互作用，表明其在向光素信号传导中发挥作用。
YtvA is a blue-light-sensing protein from Bacillus subtilis related to plant phototropins. It carries a LOV (light, oxygen and voltage) domain, binding FMN (flavin mononucleotide) as chromophore, and a STAS (sulphate transporters and antisigma-factor antagonists) domain with poorly characterized function. We have recently shown that YtvA binds triphosphate nucleotides (NTP) and highlighted a structural similarity between the STAS domain and small GTP-binding proteins. In this work we further investigated the NTP-binding properties of YtvA, employing a fluorescent derivative of GTP (GTP(TR)) and mutagenesis experiments. The main results are as follows: (a) competition experiments indicate that the affinity of YtvA for GTP is much higher than that for GDP and GMP. (b) Blue-light-induced structural changes are transmitted from the LOV core to the NTP-binding cavity, establishing a possible intraprotein signal-transduction pathway. (c) A mutation in the central beta-scaffold of the LOV core, E105L, impairs the light-driven spectroscopic changes of bound GTP(TR). This result is supported by circular dichroism data, in that YtvA-E105L does not show the light-induced conformational change in the turn fraction that characterizes YtvA, implying that E105 is functionally important. (d) In the structural model of the LOV-STAS complex, based on docking algorithms, the interface includes the I beta-H beta loop on the LOV core, as well as parts of the central beta-scaffold. E105 is predicted to interact with the LOV-STAS linker region, suggested to play a role in phototropin signaling.