• 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Journal of Molecular Biology >Elucidation of the Nature of the Conformational Changes of the EF-interhelical Loop in Bacteriorhodopsin and of the Helix VIII on the Cytoplasmic Surface of Bovine Rhodopsin: A Time-resolved Fluorescence Depolarization Study.
【24h】

Elucidation of the Nature of the Conformational Changes of the EF-interhelical Loop in Bacteriorhodopsin and of the Helix VIII on the Cytoplasmic Surface of Bovine Rhodopsin: A Time-resolved Fluorescence Depolarization Study.

机译：阐明细菌视紫红质中的EF-螺旋间环的构象变化性质以及牛视紫红质的细胞质表面上的螺旋VIII的构象变化：时间分辨荧光去极化研究。

获取原文
获取原文并翻译 | 示例
           
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

The conformation of the AB-loop and EF-loop of bacteriorhodopsin and of the fourth cytoplasmic loop (helix VIII) of bovine rhodopsin were assessed by a combination of time-resolved fluorescence depolarization and site-directed fluorescence labeling. The fluorescence anisotropy decays were measured employing a tunable Ti:sapphire laser/microchannel plate based single-photon counting apparatus with picosecond time resolution. This method allows measurement of the diffusional dynamics of the loops directly on a nanosecond time-scale. We implemented the method to study model peptides and two-helix systems representing sequences of bacteriorhodopsin. Thus, we systematically analyzed the anisotropic behavior of four different fluorescent dyes covalently bound to a single cysteine residue on the protein surface and assigned the anisotropy decay components to the modes of motion of the protein and its segments. We have identified two mechanisms of loop conformational changes in the functionally intact proteinsbacteriorhodopsin and bovine rhodopsin. First, we found a surface potential-dependent transition between two conformational states of the EF-loop of bacteriorhodopsin, detected with the fluorescent dye bound to position 160. A transition between the two conformational states at 150mM KCl and 20 degrees C requires a surface potential change that corresponds to Deltasigma approximately -1.0e(-)/bacteriorhodopsin molecule. We suggest, that the surface potential-based switch of the EF-loop is the missing link between the movement of helix F and the transient surface potential change detected during the photocycle of bacteriorhodopsin. Second, in the visual pigment rhodopsin, with the fluorescent dye bound to position 316, a particularly striking pH-dependent conformational change of the fourth loop on the cytoplasmic surface was analyzed. The loop mobility increased from pH 5 to 8. The midpoint of this transition is at pH 6.2 and correlates with the midpoint of the pH-dependent equilibrium between the active metarhodopsin II and the inactive metarhodopsin I state.
机译：通过时间分辨荧光去极化和定点荧光标记相结合，评估了细菌视紫红质的AB环和EF环以及牛视紫红质的第四个胞质环（螺旋VIII）的构象。使用具有皮秒时间分辨率的基于可调谐的Ti：蓝宝石激光器/微通道板的单光子计数设备测量荧光各向异性衰减。这种方法可以直接在纳秒级的时间尺度上测量环路的扩散动力学。我们实施了该方法来研究代表细菌视紫红质序列的模型肽和两个螺旋系统。因此，我们系统地分析了共价结合到蛋白质表面上单个半胱氨酸残基的四种不同荧光染料的各向异性行为，并将各向异性衰减成分分配给蛋白质及其片段的运动模式。我们已经确定功能完整的蛋白质细菌视紫红质和牛视紫红质中环构象变化的两种机制。首先，我们发现细菌视紫红质的EF环的两个构象状态之间存在依赖于表面电势的转变，荧光染料与位置160结合检测。在150mM KCl和20摄氏度下两个构象状态之间的转变需要表面电势大约对应于-1.0e（-）/细菌视紫红质分子的Δ变化。我们建议，EF环的基于表面电势的开关是螺旋F的运动与细菌视紫红质的光循环过程中检测到的瞬时表面电势变化之间缺少的联系。第二，在可视色素视紫红质中，荧光染料结合到位置316，分析了细胞质表面第四环的一个特别显着的pH依赖性构象变化。环的迁移率从pH 5增加到8。该转变的中点在pH 6.2处，与活动性视紫红蛋白II状态和非活动性视紫红蛋白I状态之间的pH依赖性平衡的中点相关。

著录项

相似文献

  • 外文文献
  • 中文文献
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号