Department of Life and Coordination-Complex Molecular Science, Institute for Molecular Science, 38 Nishigo-Naka, Myodaiji, Okazaki 444-8585, Japan.
J Phys Chem B. 2013 Apr 4;117(13):3449-58. doi: 10.1021/jp308765t. Epub 2013 Mar 26.
Rhodopsin contains retinal as the chromophore within seven transmembrane helices. Recently, we found a unique rhodopsin (middle rhodopsin, MR), which is evolutionarily located between the well-studied bacteriorhodopsin and sensory rhodopsin II, and which accommodates three retinal isomers in its ground state (the all-trans, the 13-cis, and, uniquely, the 11-cis isomers). In this study, we investigated structural changes of both the protein moiety and the retinal chromophore during photocycles of MR by time-resolved Fourier-transform infrared spectroscopy. Three photointermediates with decay time constants of 95 μs, 0.9 ms, and >~10 ms were identified by the global exponential fitting analysis. The first and third intermediates were attributed to the all-trans photocycle, in accordance with recently published results, whereas the second intermediate was likely one that was spectroscopically silent in the visible region and that was formed between the first and third states or resulted from the activation of the 13-cis isomer. By comparing light-induced difference spectra with various isotope labels in either the retinal or the protein moiety, we concluded that a β-sheet structure in the hydrophilic part was significantly altered during the all-trans photocycle of MR, which may involve an active state of the protein. This feature is characteristic of MR among microbial (type-1) rhodopsins.
视紫红质包含视黄醛作为发色团，位于七个跨膜螺旋内。最近，我们发现了一种独特的视紫红质（中视紫红质，MR），它在研究充分的菌视紫红质和感光视紫红质 II 之间进化，并且在其基态中容纳三种视黄醛异构体（全反式、13-顺式和独特的 11-顺式异构体）。在这项研究中，我们通过时间分辨傅里叶变换红外光谱研究了 MR 光循环过程中蛋白质部分和视黄醛发色团的结构变化。通过全局指数拟合分析，确定了三个衰减时间常数为 95μs、0.9ms 和>~10ms 的光中间产物。第一个和第三个中间产物归因于全反式光循环，这与最近发表的结果一致，而第二个中间产物可能是在可见光区域光谱上沉默的，它是在第一和第三状态之间形成的，或者是由 13-顺式异构体的激活引起的。通过将光诱导的差光谱与视黄醛或蛋白质部分的各种同位素标记进行比较，我们得出结论，在 MR 的全反式光循环过程中，亲水性部分的β-折叠结构发生了显著变化，这可能涉及蛋白质的活性状态。这一特征是 MR 在微生物（I 型）视紫红质中的特征。
