Division of Pharmaceutical Sciences, Okayama University, Okayama 700-8530, Japan.
Atmosphere and Ocean Research Institute, The University of Tokyo, Chiba 277-8564, Japan.
Sci Rep. 2017 Mar 14;7:44427. doi: 10.1038/srep44427.
Rhodopsins are proteins that contain seven transmembrane domains with a chromophore retinal and that function as photoreceptors for light-energy conversion and light-signal transduction in a wide variety of organisms. Here we characterized a phylogenetically distinctive new rhodopsin from the thermophilic eubacterium Rubrobacter xylanophilus DSM 9941 that was isolated from thermally polluted water. Although R. xylanophilus rhodopsin (RxR) is from Actinobacteria, it is located between eukaryotic and archaeal rhodopsins in the phylogenetic tree. Escherichia coli cells expressing RxR showed a light-induced decrease in environmental pH and inhibition by a protonophore, indicating that it works as a light-driven outward proton pump. We characterized purified RxR spectroscopically, and showed that it has an absorption maximum at 541 nm and binds nearly 100% all-trans retinal. The pK values for the protonated retinal Schiff base and its counterion were estimated to be 10.7 and 1.3, respectively. Time-resolved flash-photolysis experiments revealed the formation of a red-shifted intermediate. Of note, RxR showed an extremely high thermal stability in comparison with other proton pumping rhodopsins such as thermophilic rhodopsin TR (by 16-times) and bacteriorhodopsin from Halobacterium salinarum (HsBR, by 4-times).
视紫红质是一种含有七个跨膜结构域的蛋白质，其发色团为视黄醛，在多种生物中作为光能量转换和光信号转导的光感受器。在这里，我们从嗜热真细菌红杆菌属（Rubrobacter）的木聚糖嗜热菌（Rubrobacter xylanophilus）DSM 9941 中鉴定了一种新的具有系统发育特征的视紫红质，该菌是从热污染水中分离出来的。尽管 R. xylanophilus 视紫红质（RxR）来自放线菌，但它在系统发育树中位于真核生物和古菌视紫红质之间。表达 RxR 的大肠杆菌细胞表现出光诱导的环境 pH 值下降和质子载体抑制，表明它是一种光驱动的外向质子泵。我们对纯化的 RxR 进行了光谱特性表征，并表明它在 541nm 处具有最大吸收峰，并结合了近 100%的全反式视黄醛。质子化视黄醛 Schiff 碱及其抗衡离子的 pK 值分别估计为 10.7 和 1.3。时间分辨闪光光解实验揭示了形成了一个红移的中间产物。值得注意的是，与其他质子泵视紫红质（如嗜热视紫红质 TR（高出 16 倍）和来自盐生盐杆菌的菌视紫红质 HsBR（高出 4 倍））相比，RxR 表现出极高的热稳定性。
