The Laser-Induced Potential Jump: A Method for Rapid Electron Injection into Oxidoreductase Enzymes
The Laser-Induced Potential Jump: A Method for Rapid Electron Injection into Oxidoreductase Enzymes
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DOI：
10.1021/acs.jpcb.0c05718
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发表时间：
2020-10-08
影响因子：
3.3
通讯作者：
Dyer, R. Brian
中科院分区：
文献类型：
--
作者：
Sanchez, Monica L. K.;Konecny, Sara E.;Dyer, R. Brian
氧化还原酶经常利用丰富的金属辅因子在环境条件下高效地进行技术上有用的化学转化。然而，对这些酶的催化机制的理解高度依赖于具有良好特征和优化的时间分辨分析技术的可用性。我们已经开发了一种方法，可以将电子快速注入到催化系统中，使用光活化纳米材料与一系列氧化还原介质相结合，在溶液中产生潜在的跃迁，然后通过电子转移(ET)到催化剂来启动周转。然而，纳米材料-介体催化剂界面上的ET事件对实验条件高度敏感，如光子通量、体系组分的相对浓度和pH。在这里，我们针对一个特定的催化体系--莱茵衣藻[FeFe]氢酶(CrHydAl)对这些实验参数进行了系统的优化。然而，所开发的策略可以应用于各种氧化还原酶的研究。我们的电势跃迁体系由作为光敏剂的CdSe/CDS核壳纳米棒和作为介体的一系列取代的联吡啶盐组成，氧化还原电位在-550 mV到-670 mV(Vs Se)之间。利用这些成分，我们筛选了pH、介体浓度、蛋白质浓度、光敏剂浓度和光子通量对稳态光还原和产氢以及ET和潜在跳跃效率的影响。通过操作这些实验条件，我们展示了简单修改的潜力，以提高潜在跳跃的可调性，应用于研究氧化还原酶。
Oxidoreductase enzymes often perform technologically useful chemical transformations using abundant metal cofactors with high efficiency under ambient conditions. The understanding of the catalytic mechanism of these enzymes is, however, highly dependent on the availability of well-characterized and optimized time-resolved analytical techniques. We have developed an approach for rapidly injecting electrons into a catalytic system using a photoactivated nanomaterial in combination with a range of redox mediators to produce a potential jump in solution, which then initiates turnover via electron transfer (ET) to the catalyst. The ET events at the nanomaterial-mediatorcatalyst interfaces are, however, highly sensitive to the experimental conditions such as photon flux, relative concentrations of system components, and pH. Here, we present a systematic optimization of these experimental parameters for a specific catalytic system, namely, [FeFe] hydrogenase from Chlamydomonas reinhardtii (CrHydAl). The developed strategies can, however, be applied in the study of a wide variety of oxidoreductase enzymes. Our potential jump system consists of CdSe/CdS core-shell nanorods as a photosensitizer and a series of substituted bipyridinium salts as mediators with redox potentials in the range from -550 to -670 mV (vs SHE). With these components, we screened the effect of pH, mediator concentration, protein concentration, photosensitizer concentration, and photon flux on steady-state photoreduction and hydrogen production as well as ET and potential jump efficiency. By manipulating these experimental conditions, we show the potential of simple modifications to improve the tunability of the potential jump for application to study oxidoreductases.