Unravelling enzymatic hydrogen production mechanisms with ultrafast 2D-IR spectroscopy

氢酶是一类金属酶家族，能有效地催化氢的可逆裂解为质子和电子，因此为氢的可持续能源生产提供了理想的原型催化剂。然而，在我们能够开发基于氢酶的技术材料之前，需要详细了解氢气的产生和使用的机制。这个多学科项目将结合超快2D-IR光谱(Hunt)与电化学和生化方法(Parkin)，以了解结构变化和动态过程在这些重要酶的作用中的作用。2D-IR是一种新的超快激光光谱方法，它将分子的红外光谱扩展到第二个频率轴上[1]。这种多维性使人们能够获得丰富的新光谱信息，但2D-IR最强大的方面是能够以100fS的时间分辨率确定分子结构和结构中的变化(10-13 S)。这为实时观察氢酶机制中心的反应步骤、溶剂运动或氢键振动提供了可能性。这是对Parkin小组使用傅立叶变换大幅值交流伏安法获得的氢酶中超快电子运动的洞察的高度补充[2]。虽然氢酶的合成模型已经用2D-IR[3，4]进行了研究，但这次合作首次提供了将这些结果与完整的酶系统进行比较的独特机会。通过研究一系列的位点突变，并使用电化学方法制备催化中间体，我们将使用2D-IR来了解蛋白质支架与活性位点相互作用的方式，从而影响或控制酶的机制。这是一个特别重要的问题，因为缺乏蛋白质支架的仿生模型化合物表现出非常低的催化效率，强调了它的重要性[4]，但它在酶循环中所起的确切分子功能是未知的。(1)Hunt，N.T.Chem Soc Rev 2009，38,1837.(2)Adamson，H.；Robinson，M.；Wright，J.；Flanagan，L.A.；Walton，J.；Elton，D.；Gavaghan，D.；Bond，A.M.；Roessler，M.M.；(3)Fritzsch，R.；Brady，O.；Adair，E.；Wright，J.A.；Pickett，C.J.；Hunt，N.T.物理化学快报，2016年，7,2838.(4)Frederx，P.W.J.M.；Adamczyk，K.；Wright，J.A.；Tuttle，T.；Ulijn，R.V.；Pickett，C.J.；Hunt，N.T.有机金属学报，2014年，33,5888。

The hydrogenases are families of metalloenzymes that efficiently catalyse the reversible cleavage of H2 into protons and electrons and so offer ideal prototype catalysts for sustainable energy generation involving H2. Before we can develop technological materials based upon the hydrogenases however, a detailed understanding of the mechanism of H2 production and usage is required. This multidisciplinary project will combine ultrafast 2D-IR spectroscopy (Hunt) with electrochemical and biochemical methods (Parkin) to understand structural changes and the role of dynamic processes in the action of these important enzymes. 2D-IR is a new ultrafast laser spectroscopy method that spreads the IR spectrum of a molecule over a second frequency axis [1]. This multidimensionality allows access to a wealth of new spectral information but the most powerful aspect of 2D-IR is the ability to determine molecular structure and changes in that structure with a time resolution of 100 fs (10-13 s). This opens up the possibility of observing reaction steps, solvent motion or H-bond vibrations that are central to the hydrogenase mechanism in real time. This is highly complementary to the insight into ultrafast electron movement in hydrogenases which can has been obtained in the Parkin group using Fourier transform large amplitude alternating current voltammetry [2]. Although synthetic models of the hydrogenases have been studied with 2D-IR [3,4], this collaboration offers the unique opportunity to compare these results with the full enzyme systems for the first time. By studying a range of site mutations and using electrochemical methods to prepare catalytic intermediates we will use 2D-IR to understand the way in which the protein scaffold interacts with the active site to influence or control the enzyme mechanism. This is a particularly important issue since biomimetic model compounds lacking the protein scaffold show very low catalytic efficiency, emphasising its importance [4], but the precise molecular function that it plays in the enzyme cycle is unknown.(1) Hunt, N. T. Chem Soc Rev 2009, 38, 1837.(2) Adamson, H.; Robinson, M.; Wright, J. J.; Flanagan, L. A.; Walton, J.; Elton, D.; Gavaghan, D. J.; Bond, A. M.; Roessler, M. M.; Parkin, A. Journal of the American Chemical Society 2017, 139, 10677.(3) Fritzsch, R.; Brady, O.; Adair, E.; Wright, J. A.; Pickett, C. J.; Hunt, N. T. Journal of Physical Chemistry Letters 2016, 7, 2838.(4) Frederix, P. W. J. M.; Adamczyk, K.; Wright, J. A.; Tuttle, T.; Ulijn, R. V.; Pickett, C. J.; Hunt, N. T. Organometallics 2014, 33, 5888.