Direct observation of the structural change of Tyr174 in the primary reaction of sensory rhodopsin II.
Direct observation of the structural change of Tyr174 in the primary reaction of sensory rhodopsin II.
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DOI：
10.1021/bi101817y
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发表时间：
2011-03
期刊：
影响因子：
2.9
通讯作者：
M. Mizuno;Y. Sudo;M. Homma;Y. Mizutani
中科院分区：
文献类型：
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作者：
M. Mizuno;Y. Sudo;M. Homma;Y. Mizutani
感觉视紫红质II（SRII）是一种负性趋光性受体，其发色团为视黄醛，介导对蓝光的回避。信号转导是由视网膜发色团的光异构化引发的，导致蛋白质的构象变化，这些构象变化被传递到转导蛋白。为了深入了解SRII传感机制，我们采用时间分辨紫外共振拉曼光谱监测光异构化后皮秒时间范围内蛋白质结构的变化。我们使用了450 nm的泵浦脉冲启动SRII光循环和两种探测脉冲的波长为225和238 nm的色氨酸和酪氨酸波段，分别检测光谱的变化。观察到的拉曼谱带的光谱变化很可能是由于位于视网膜发色团附近的色氨酸和酪氨酸残基，即，Trp 76、Trp 171、Tyr 51或Tyr 174。225 nm的UVRR光谱表现出漂白的仪器响应时间内的所有的色氨酸带的强度，然后部分恢复与30 ps的时间常数，并没有进一步的变化高达1 ns。在238 nm的UVRR光谱中，除了30 ps的时间常数分量外，还观察到快速恢复分量。SRII的WT和Y174 F突变体的光谱之间的比较表明，Tyr 174改变其结构和/或发色团光异构化后的环境。这些数据代表了Tyr 174结构变化的首次实时观察，其功能重要性先前已指出。
Sensory rhodopsin II (SRII) is a negative phototaxis receptor containing retinal as its chromophore, which mediates the avoidance of blue light. The signal transduction is initiated by the photoisomerization of the retinal chromophore, resulting in conformational changes of the protein which are transmitted to a transducer protein. To gain insight into the SRII sensing mechanism, we employed time-resolved ultraviolet resonance Raman spectroscopy monitoring changes in the protein structure in the picosecond time range following photoisomerization. We used a 450 nm pump pulse to initiate the SRII photocycle and two kinds of probe pulses with wavelengths of 225 and 238 nm to detect spectral changes in the tryptophan and tyrosine bands, respectively. The observed spectral changes of the Raman bands are most likely due to tryptophan and tyrosine residues located in the vicinity of the retinal chromophore, i.e., Trp76, Trp171, Tyr51, or Tyr174. The 225 nm UVRR spectra exhibited bleaching of the intensity for all the tryptophan bands within the instrumental response time, followed by a partial recovery with a time constant of 30 ps and no further changes up to 1 ns. In the 238 nm UVRR spectra, a fast recovering component was observed in addition to the 30 ps time constant component. A comparison between the spectra of the WT and Y174F mutant of SRII indicates that Tyr174 changes its structure and/or environment upon chromophore photoisomerization. These data represent the first real-time observation of the structural change of Tyr174, of which functional importance was pointed out previously.