Serial Femtosecond Crystallography of Optogenetic Function

设计可遗传编码的合成光敏材料的能力为生物学家提供了感知和控制生物功能和环境的手段和机会。为了了解这些材料是如何发挥作用的，最近已经有可能使用X射线结晶学对光致运动进行非常快速的“快照”，这甚至可能在非常短的时间尺度上仍然涉及激发的电子(在飞秒时间尺度上)。新的第四代光源现在可以用来录制这样的分子电影，这是我们最近才展示的一项突破性技术。在不久的将来，我们预计能够录制质量更好的电影，因为汉堡的欧洲XFEL将开始用户操作，它允许记录更多的数据。以前我们只能使用激光光谱学技术，现在我们实际上可以直接‘观察’蛋白质在激活后的第一次运动。众所周知，涉及感知或激活(通过‘激活’)的生物反应的‘结果’是在这些发生在时间尺度上的最早的运动中决定的，这些运动通常小于皮秒。例如，在双向光致变色荧光蛋白的情况下，单向开关的效率很好，为30%，而反向开关的效率较差，低于0.05%，潜在地限制了光遗传学的应用。因此，我们能够提出这样一个问题：是什么决定了反应结果的这种显著差异，并提供了结构和动力学信息，这些信息将对合理的光遗传学设计的反馈非常有价值。

The ability to design synthetic light-sensitive materials that can be genetically encoded provides biologists the means and opportunity to sense and control biological function and environment. In order to learn how these materials function it has very recently become possible to make very fast 'snapshots' of the light-induced motions using X-ray crystallography, which is even possible for very short time scales that still involves the excited electrons (on femtosecond time scale). Novel 4th generation light sources can now be used to record such 'molecular movies' which is a breakthrough technology that we only very recently demonstrated. In the very near future we anticipate being able to record even better quality movies, as the European XFEL in Hamburg that allows much more data to be recorded will start user operation. Where previously we could only use laser spectroscopy techniques we can now actually 'watch' the very first motions of proteins directly after activation. It is known that the 'outcome' of biological reactions involving either sensing or activation (via 'actuation') is determined in these very first motions that occur on time scales typically less than picoseconds. For example, in the case of a bi-directional photochromic fluorescent protein the efficiency of switching in one direction is excellent at 30%, while the efficiency of switching in the reverse direction is poor at less than 0.05%, potentially limiting optogenetics applications. We are thus in a position to ask the question what determines this dramatic difference in the outcome of the reactions, and provide structural and dynamical information that will be highly valuable for feeding back for rational optogenetics design.