Sahin Selmihan, Brazard Johanna, Zuchan Kilian, Adachi Takuji B M, Mühlenhoff Ulrich, Milton Ross D, Stripp Sven T
University of Geneva, Department of Inorganic and Analytical Chemistry, Sciences II Quai Ernest-Ansermet 30 Geneva 4 1211 Switzerland
Suleyman Demirel University, Department of Chemistry, Faculty of Engineering and Natural Sciences Cunur Isparta 32260 Turkiye.
Chem Sci. 2025 Apr 30. doi: 10.1039/d5sc00550g.
Ferredoxins are small iron-sulfur proteins that engage in one-electron transfer with oxidoreductases across all domains of life. The catalyzed reactions often include multiple electrons, , in the two-electron reduction of NADP during photosynthesis or the reduction of protons to H by the metalloenzyme hydrogenase. To date, the microscopic details of how ferredoxins facilitate multiple electron redox chemistry are unknown. Ferredoxins of the subfamily contain two [4Fe-4S] clusters, which allows for two one-electron transfer reactions. However, the iron-sulfur clusters of conventional 2[4Fe-4S]-type ferredoxins have very similar reduction potentials and conclusive evidence for the transfer of two electrons during a single protein-protein interaction (PPI) has not been reported. In this work, the electron transfer complexes between the clostridial 2[4Fe-4S] ferredoxin, Fd, and [FeFe]-hydrogenases from both (I) and (HydA), were investigated. Introducing a non-canonical amino acid near to one of the iron-sulfur clusters of Fd permitted the quantification of electric field changes the vibrational Stark effect by Fourier-transform infrared (FTIR) spectroscopy. Upon reduction, FTIR difference spectroscopy reported on protein structural changes and microscale thermophoresis revealed that the affinity between ferredoxin and hydrogenase is modulated by redox-dependent PPIs. Prompted by these findings, we suggest a model how ferredoxin efficiently facilitates multiple electron redox chemistry based on individual one-electron transfer reactions.
铁氧化还原蛋白是一类小型铁硫蛋白，在生命的所有领域中，它们都能与氧化还原酶进行单电子转移。催化的反应通常涉及多个电子，例如在光合作用中NADP的双电子还原过程，或金属酶氢化酶将质子还原为H的过程。迄今为止，铁氧化还原蛋白如何促进多电子氧化还原化学反应的微观细节尚不清楚。亚家族的铁氧化还原蛋白包含两个[4Fe-4S]簇，这允许进行两个单电子转移反应。然而，传统的2[4Fe-4S]型铁氧化还原蛋白的铁硫簇具有非常相似的还原电位，并且尚未有关于在单个蛋白质-蛋白质相互作用（PPI）过程中转移两个电子的确凿证据报道。在这项工作中，研究了梭菌2[4Fe-4S]铁氧化还原蛋白Fd与来自（I）和（HydA）的[FeFe] -氢化酶之间的电子转移复合物。在Fd的一个铁硫簇附近引入一个非规范氨基酸，通过傅里叶变换红外（FTIR）光谱法能够量化电场变化——振动斯塔克效应。还原后，FTIR差光谱揭示了蛋白质结构变化，微尺度热泳显示铁氧化还原蛋白和氢化酶之间的亲和力受氧化还原依赖性PPI的调节。基于这些发现，我们提出了一个模型，说明铁氧化还原蛋白如何基于单个单电子转移反应有效地促进多电子氧化还原化学反应。
