Visualizing reaction pathways in photoactive yellow protein from nanoseconds to seconds
Visualizing reaction pathways in photoactive yellow protein from nanoseconds to seconds
复制标题
DOI：
10.1073/pnas.0409035102
复制
发表时间：
2005-05-17
影响因子：
11.1
通讯作者：
Moffat, K
中科院分区：
文献类型：
--
作者：
Ihee, H;Rajagopal, S;Moffat, K
在环境条件下，实时确定蛋白质生物作用过程中的三维中间结构对于理解蛋白质的功能至关重要。在这里，我们使用定时器分辨的劳厄结晶学来提取短暂的中间结构，从而揭示盐生盐生螺旋体蓝光感光器光活性黄色蛋白(PYP)的信号转导。通过分析PYP光周期(从1纳秒到1秒的47个时间点)中的一组完整的Laue数据，我们跟踪了PYP光周期中的所有原子，并直接观察到蓝光光子被其p-香豆酸生色团吸收如何触发可逆光循环。我们确定了一个复杂的化学机制，其特征是五个不同的结构中间体。早期红移中间体中生色团的结构变化通过最初的“体积守恒”异构化和生色团与其周围结合口袋之间氢键的逐渐破坏，在晚期蓝移中间体中被传递到蛋白质的外部。根据以前对该系统的生物物理研究，这些结果提供了对PYP光循环的全面看法。
Determining 3D intermediate structures during the biological action of proteins in real time under ambient conditions is essential for understanding how proteins function. Here we use timer-esolved Laue crystallography to extract short-lived intermediate structures and thereby unveil signal transduction in the blue light photoreceptor photoactive yellow protein (PYP) from Halorhodospira halophila. By analyzing a comprehensive set of Laue data during the PYP photocycle (forty-seven time points from one nanosecond to one second), we track all atoms in PYP during its photocycle and directly observe how absorption of a blue light photon by its p-coumaric acid chromophore triggers a reversible photocycle. We identify a complex chemical mechanism characterized by five distinct structural intermediates. Structural changes at the chromophore in the early, red-shifted intermediates are transduced to the exterior of the protein in the late, blue-shifted intermediates through an initial "volume-conserving" isomerization of the chromophore and the progressive disruption of hydrogen bonds between the chromophore and its surrounding binding pocket. These results yield a comprehensive view of the PYP photocycle when seen in the light of previous biophysical studies on the system.