Mutational effects on protein structural changes and interdomain interactions in the blue-light sensing LOV protein YtvA
Mutational effects on protein structural changes and interdomain interactions in the blue-light sensing LOV protein YtvA
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DOI：
10.1562/2005-05-25-ra-541
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发表时间：
2005-09-01
影响因子：
3.3
通讯作者：
Gärtner, W
中科院分区：
文献类型：
--
作者：
Losi, A;Ghiraldelli, E;Gärtner, W
对来自枯草芽孢杆菌的向光蛋白相关蛋白YtvA的诱变研究揭示了所选结构元件在结构域间通信中的作用。YtvA的LOV（光、氧、电压）结构域经历与phot-LOV类似的光驱动反应，可逆地形成共价黄素-半胱氨酸加合物。用紫外荧光光谱和圆二色光谱对突变蛋白Ytva-E105 L和YtvA-E56 Q进行了研究。E105（光致伸缩蛋白中的L）位于LOV结构域中心P-片层的溶剂暴露表面，被证明参与光致伸缩蛋白中的结构域间相互作用。CD数据显示，YtvA-E105 L在黑暗中具有较低的α-螺旋含量，并且比YtvA-WT经历更大的光驱动构象变化。E56 Q突变破坏了E56-K97盐桥，这是LOV系列中高度保守的结构元件。在YtvA-E56 Q中，CD光谱与YtvA-WT中相同，尽管保守的W103变得更多地暴露于溶剂并且暗恢复动力学较慢。这些结果表明，E56-K97盐桥局部稳定蛋白质结构，并参与光循环的调节，但对整体结构的影响可以忽略不计。相反，E105 L突变突出了LOV蛋白中光驱动的构象变化中中心P-折叠的参与。
Mutagenesis studies on the phototropin-related protein YtvA from Bacillus subtilis have revealed the role of selected structural elements in interdomain communication. The LOV (light, oxygen, voltage) domain of YtvA undergoes light-driven reactions similar to that of phot-LOV, with reversible formation of a covalent flavin-cysteine adduct. The mutated proteins Ytva-E105L and YtvA-E56Q have been studied by UV fluorescence and circular dichroism (CD) spectroscopy. E105 (L in phototropin) is located at the solvent-exposed surface of the LOV domain central P-sheet, demonstrated to participate in interdomain interaction in phototropin. CD data show that YtvA-E105L has a lower a-helix content in the dark and undergoes larger light-driven conformational changes than YtvA-WT. The E56Q mutation breaks the E56-K97 salt bridge, a structural element highly conserved within the LOV series. In YtvA-E56Q the CD spectrum is the same as in YtvA-WT, although the conserved W103 becomes more exposed to the solvent and the dark-recovery kinetics is slower. These results indicate that the E56-K97 salt bridge stabilizes locally the protein structure and participates in the regulation of the photocycle but has negligible effects on the overall structure. The E105L mutation, instead, highlights the involvement of the central P-sheet in the light-driven conformational changes in LOV proteins.