PICOSECOND TIME-RESOLVED CRYSTALLOGRAPHY

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 我们的目标是使用皮秒时间分辨劳厄结晶学技术来研究蛋白质中的生物物理过程。具体地说，我们将使用强度高、持续时间短的激光脉冲来触发蛋白质晶体的结构变化，并将使用一系列X射线快门从同步加速器脉冲序列中分离出单个X射线脉冲来探索其结构演变。通过获取激光光解后特定时刻的衍射数据，我们可以构建出蛋白质S结构的快照，其时间分辨率仅受X射线脉冲持续时间的限制，该脉冲的持续时间约为100ps。我们的目标是解决大量的生物物理问题，包括蛋白质中高度保守残基的功能作用，配体进出蛋白质S活性部位的途径，以及调节或控制变构调节的相关结构变化。为了获得尽可能高质量的数据，我们首先将重点放在结构变化是可逆的蛋白质系统上。配体结合的血红素蛋白，包括肌红蛋白和血红蛋白，是这些生物物理研究的理想模型系统。当CO被用作O2的替代物时，配体可以高量子效率地从血红素上光解离，从而触发一系列可以从结构上研究的事件。由于解离的配体在ms时间尺度上以双分子的方式重新结合到血红素上，结构返回到其起始状态，这个过程可以重复数千次。因此，这些模型系统是进行这些生物物理研究的理想选择。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We aim to employ the technique of picosecond time-resolved Laue crystallography to study biophysical processes in proteins. Specifically, we will use intense, short-duration laser pulses to trigger a structural change in a protein crystal, and will probe its structural evolution with single X-ray pulses isolated from the synchrotron pulse train using a sequence of X-ray shutters. By acquiring diffraction data at a well-defined instant in time after laser photolysis, we can construct a snapshot of the protein?s structure with time resolution limited only by the duration of the X-ray pulse, which is of the order of 100 ps. We aim to address numerous biophysical questions including the functional role of highly conserved residues in proteins, pathways for ligand migration to and from the protein?s active site, and the correlated structural changes that mediate or control allosteric regulation. To obtain the highest quality data possible, we will focus initially on protein systems whose structural changes are reversible. Ligand-binding heme proteins, including myoglobin and hemoglobin, are ideal model systems for these biophysical investigations. When CO is used as a surrogate for O2, the ligand can be photodissociated from the heme with high quantum efficiency, thereby triggering a sequence of events that can be studies structurally. Because the dissociated ligands rebind to the heme bimolecularly on the ms time scale, the structure returns to its starting state and the process can be repeated thousands of times. Thus, these model systems are ideal for pursuing these biophysical studies.