Blue light-excited LOV1 and LOV2 domains cooperatively regulate the kinase activity of full-length phototropin2 from Arabidopsis
Blue light-excited LOV1 and LOV2 domains cooperatively regulate the kinase activity of full-length phototropin2 from Arabidopsis
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DOI：
10.1074/jbc.ra117.000324
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发表时间：
2018-01-19
影响因子：
4.8
通讯作者：
Nakasako, Masayoshi
中科院分区：
文献类型：
--
作者：
Oide, Mao;Okajima, Koji;Nakasako, Masayoshi
向光蛋白2（phototropin 2，phot 2）是一种蓝光（blue-light，BL）受体，在植物中调节BL依赖的活性以实现高效的光合作用。phot 2包含两个BL-接收光-氧-电压-敏感结构域（LOV 1和LOV 2）和激酶结构域。BL-激发的LOV 2被认为是主要负责BL依赖性激活的激酶。然而，小BL诱导的LOV 2结构域的构象变化传递到激酶的分子机制仍不清楚。在这里，我们使用全长野生型和突变体phot 2蛋白从拟南芥研究其在黑暗中的分子特性和BL照射下。磷酸化分析和吸收测量表明，LOV 1域有助于BL-激发LOV 2的热弛豫，反之亦然。使用小角X射线散射和电子显微镜，我们观察到，phot 2形成二聚体，并具有最大长度为188埃，回转半径为44埃的棒状。在BL下，phot 2显示出大的构象变化，使棒状弯曲。通过叠加的晶体结构的LOV 1二聚体，LOV 2，和一个同源模型的激酶所观察到的变化，我们推断，BL-依赖的变化包括的LOV 2和激酶相对于LOV 1的位置偏移。此外，在LOV 1或LOV 2中缺乏光循环的phot 2突变体在BL下仍然表现出构象变化，这表明LOV 1和LOV 2协同促进激活激酶的构象变化。这些结果表明，BL激活的LOV 1有助于phot 2的激酶活性。我们讨论了可能的分子内相互作用和信号转导机制的光2。
Phototropin2 (phot2) is a blue-light (BL) receptor that regulates BL-dependent activities for efficient photosynthesis in plants. phot2 comprises two BL-receiving light-oxygen-voltage-sensing domains (LOV1 and LOV2) and a kinase domain. BL-excited LOV2 is thought to be primarily responsible for the BL-dependent activation of the kinase. However, the molecular mechanisms by which small BL-induced conformational changes in the LOV2 domain are transmitted to the kinase remain unclear. Here, we used full-length wild-type and mutant phot2 proteins from Arabidopsis to study their molecular properties in the dark and under BL irradiation. Phosphorylation assays and absorption measurements indicated that the LOV1 domain assists the thermal relaxation of BL-excited LOV2 and vice versa. Using small-angle X-ray scattering and electron microscopy, we observed that phot2 forms a dimer and has a rod shape with a maximum length of 188 angstrom and a radius of gyration of 44 angstrom. Under BL, phot2 displayed large conformational changes that bent the rod shape. By superimposing the crystal structures of the LOV1 dimer, LOV2, and a homology model of the kinase to the observed changes, we inferred that the BL-dependent change consisted of positional shifts of both LOV2 and the kinase relative to LOV1. Furthermore, phot2 mutants lacking the photocycle in LOV1 or LOV2 still exhibited conformational changes under BL, suggesting that LOV1 and LOV2 cooperatively contribute to the conformational changes that activate the kinase. These results suggest that BL-activated LOV1 contributes to the kinase activity of phot2. We discuss the possible intramolecular interactions and signaling mechanisms in phot2.