Laser-induced transient grating analysis of dynamics of interaction between sensory rhodopsin II D75N and the HtrII transducer
Laser-induced transient grating analysis of dynamics of interaction between sensory rhodopsin II D75N and the HtrII transducer
复制标题
DOI：
10.1529/biophysj.106.097493
复制
发表时间：
2007-03-15
影响因子：
3.4
通讯作者：
Terazima, Masahide
中科院分区：
文献类型：
--
作者：
Inoue, Keiichi;Sasaki, Jun;Terazima, Masahide
使用 SRII 的 D75N 突变体，通过时间分辨激光诱导瞬态光栅方法研究了感觉视紫红质 II (SRII) 与其传感器 HtrII 之间的相互作用，该突变体在其光循环过程中表现出最小的可见光吸收变化，但介导正常的趋光反应。无传感器的 D75N 和连接到 SRII 传感器蛋白 HtrII (Delta HtrII) N 端部分 120 个氨基酸残基的 D75N 的闪光诱导瞬态吸收光谱显示，其光循环中仅存在一种光谱上不同的 K 样中间体，但瞬态光栅方法解析了四种体积不同的中间体 (K-1 -K-4)。与 HtrII 结合的 D75N 表现出另一种较慢的动力学物质，通过紫外-可见光区域的吸收变化评估，该物质在完全恢复初始状态后仍然存在。动力学表明换能器部分（指定为 Tr*）的构象变化形式，在光感受器返回到未光解状态后仍然存在。 DHtrII 部分的最大构象变化被发现导致 K-4 状态下 8 μs 内体积上升的 Delta HtrII 依赖性增加，以及 K-4 的扩散系数 (D) 相对于未光解状态和 Tr* 的扩散系数 (D) 急剧下降。 D 减少的幅度表明存在较大的结构变化，可能是在 Delta HtrII 暴露于溶剂的 HAMP 结构域中，其中溶剂中相互作用分子的重排将显着改变蛋白质和溶剂之间的摩擦力。
The interaction between sensory rhodopsin II (SRII) and its transducer HtrII was studied by the time-resolved laser-induced transient grating method using the D75N mutant of SRII, which exhibits minimal visible light absorption changes during its photocycle, but mediates normal phototaxis responses. Flash-induced transient absorption spectra of transducer-free D75N and D75N joined to 120 amino-acid residues of the N-terminal part of the SRII transducer protein HtrII (Delta HtrII) showed only one spectrally distinct K-like intermediate in their photocycles, but the transient grating method resolved four intermediates (K-1 -K-4) distinct in their volumes. D75N bound to HtrII exhibited one additional slower kinetic species, which persists after complete recovery of the initial state as assessed by absorption changes in the UV-visible region. The kinetics indicate a conformationally changed form of the transducer portion (designated Tr*), which persists after the photoreceptor returns to the unphotolyzed state. The largest conformational change in the DHtrII portion was found to cause a Delta HtrII-dependent increase in volume rising in 8 mu s in the K-4 state and a drastic decrease in the diffusion coefficient (D) of K-4 relatively to those of the unphotolyzed state and Tr*. The magnitude of the decrease in D indicates a large structural change, presumably in the solventexposed HAMP domain of Delta HtrII, where rearrangement of interacting molecules in the solvent would substantially change friction between the protein and the solvent.