Tsunoda Satoshi P, Ewers David, Gazzarrini Sabrina, Moroni Anna, Gradmann Dietrich, Hegemann Peter
Experimentelle Biophysik, Fachbereich für Biologie, Humboldt-Universität zu Berlin, 10115 Berlin, Germany.
Biophys J. 2006 Aug 15;91(4):1471-9. doi: 10.1529/biophysj.106.086421. Epub 2006 May 26.
An opsin-encoding cDNA was cloned from the marine alga Acetabularia acetabulum. The cDNA was expressed in Xenopus oocytes into functional Acetabularia rhodopsin (AR) mediating H+ carried outward photocurrents of up to 1.2 microA with an action spectrum maximum at 518 nm (AR518). AR is the first ion-pumping rhodopsin found in a plant organism. Steady-state photocurrents of AR are always positive and rise sigmoidally from negative to positive transmembrane voltages. Numerous kinetic details (amplitudes and time constants), including voltage-dependent recovery of the dark state after light-off, are documented with respect to their sensitivities to light, internal and external pH, and the transmembrane voltage. The results are analyzed by enzyme kinetic formalisms using a simplified version of the known photocycle of bacteriorhodopsin (BR). Blue-light causes a shunt of the photocycle under H+ reuptake from the extracellular side. Similarities and differences of AR with BR are pointed out. This detailed electrophysiological characterization highlights voltage dependencies in catalytic membrane processes of this eukaryotic, H+ -pumping rhodopsin and of microbial-type rhodopsins in general.
从海洋藻类伞藻中克隆出一种编码视蛋白的cDNA。该cDNA在非洲爪蟾卵母细胞中表达，形成功能性的伞藻视紫红质（AR），介导高达1.2微安的外向质子光电流，其作用光谱峰值在518纳米（AR518）。AR是在植物生物体中发现的首个离子泵视紫红质。AR的稳态光电流始终为正，并且在跨膜电压从负到正的过程中呈S形上升。记录了许多动力学细节（幅度和时间常数），包括光照后暗态的电压依赖性恢复，以及它们对光、内部和外部pH值以及跨膜电压的敏感性。使用细菌视紫红质（BR）已知光循环的简化版本，通过酶动力学形式对结果进行分析。蓝光会导致在从细胞外侧重新摄取质子的情况下光循环出现分流。指出了AR与BR的异同。这种详细的电生理特性突出了这种真核的、质子泵视紫红质以及一般微生物型视紫红质在催化膜过程中的电压依赖性。
