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Abstract：
Determining short-lived intermediate structures in chemical reactions is challenging. Although ultrafast spectroscopic methods can detect the formation of transient intermediates, real-space structures cannot be determined directly from such studies. Time-resolved serial femtosecond crystallography (TR-SFX) has recently proven to be a powerful method for capturing molecular changes in proteins on femtosecond timescales. However, the methodology has been mostly applied to natural proteins/enzymes and limited to reactions promoted by synthetic molecules due to structure determination challenges. This work demonstrates the applicability of TR-SFX for investigations of chemical reaction mechanisms of synthetic metal complexes. We fix a light-induced CO-releasing Mn(CO)3 reaction center in porous hen egg white lysozyme (HEWL) microcrystals. By controlling light exposure and time, we capture the real-time formation of Mn-carbonyl intermediates during the CO release reaction. The asymmetric protein environment is found to influence the order of CO release. The experimentally-observed reaction path agrees with quantum mechanical calculations. Therefore, our demonstration offers a new approach to visualize atomic-level reactions of small molecules using TR-SFX with real-space structure determination. This advance holds the potential to facilitate design of artificial metalloenzymes with precise mechanisms, empowering design, control and development of innovative reactions.
摘要：
在化学反应中确定短寿命的中间结构是具有挑战性的。尽管超快光谱方法可以检测到瞬态中间体的形成，真实空间结构不能直接从这些研究中确定。时间分辨串行飞秒晶体学（TR-SFX）最近被证明是一种在飞秒时间尺度上捕获蛋白质分子变化的强大方法。然而,该方法主要应用于天然蛋白质/酶，并且由于结构确定的挑战而仅限于由合成分子促进的反应。这项工作证明了TR-SFX可用于研究合成金属配合物的化学反应机理。我们将光诱导的CO释放Mn（CO）3反应中心固定在多孔鸡蛋清溶菌酶（HEWL）微晶中。通过控制曝光和时间，我们捕获了在CO释放反应过程中Mn-羰基中间体的实时形成。发现不对称蛋白质环境影响CO释放的顺序。实验观察到的反应路径与量子力学计算一致。因此,我们的演示提供了一种使用TR-SFX和实空间结构测定可视化小分子原子级反应的新方法。这一进展具有促进具有精确机制的人工金属酶设计的潜力，授权设计，控制和发展创新反应。
