Badiani Vivek M, Cobb Samuel J, Wagner Andreas, Oliveira Ana Rita, Zacarias Sónia, Pereira Inês A C, Reisner Erwin
Yusuf Hamied Department of Chemistry, University of Cambridge, Lensfield Road, Cambridge CB2 1EW, U.K.
Cambridge Graphene Centre, University of Cambridge, Cambridge CB3 0FA, U.K.
ACS Catal. 2022 Feb 4;12(3):1886-1897. doi: 10.1021/acscatal.1c04317. Epub 2022 Jan 20.
The immobilization of redox enzymes on electrodes enables the efficient and selective electrocatalysis of useful reactions such as the reversible interconversion of dihydrogen (H) to protons (H) and formate to carbon dioxide (CO) with hydrogenase (Hase) and formate dehydrogenase (FDH), respectively. However, their immobilization on electrodes to produce electroactive protein films for direct electron transfer (DET) at the protein-electrode interface is not well understood, and the reasons for their activity loss remain vague, limiting their performance often to hour timescales. Here, we report the immobilization of [NiFeSe]-Hase and [W]-FDH from Hildenborough on a range of charged and neutral self-assembled monolayer (SAM)-modified gold electrodes with varying hydrogen bond (H-bond) donor capabilities. The key factors dominating the activity and stability of the immobilized enzymes are determined using protein film voltammetry (PFV), chronoamperometry (CA), and electrochemical quartz crystal microbalance (E-QCM) analysis. Electrostatic and H-bonding interactions are resolved, with electrostatic interactions responsible for enzyme orientation while enzyme desorption is strongly limited when H-bonding is present at the enzyme-electrode interface. Conversely, enzyme stability is drastically reduced in the absence of H-bonding, and desorptive enzyme loss is confirmed as the main reason for activity decay by E-QCM during CA. This study provides insights into the possible reasons for the reduced activity of immobilized redox enzymes and the role of film loss, particularly H-bonding, in stabilizing bioelectrode performance, promoting avenues for future improvements in bioelectrocatalysis.
将氧化还原酶固定在电极上能够高效且选择性地电催化一些有用的反应，比如分别利用氢化酶（Hase）和甲酸脱氢酶（FDH）实现氢气（H₂）与质子（H⁺）以及甲酸与二氧化碳（CO₂）之间的可逆相互转化。然而，对于如何将它们固定在电极上以制备用于在蛋白质 - 电极界面进行直接电子转移（DET）的电活性蛋白质膜，我们还了解得不够透彻，而且它们活性丧失的原因也尚不明确，这常常将其性能限制在小时级的时间尺度内。在此，我们报道了将来自希尔德恩伯勒的[NiFeSe]-Hase和[W]-FDH固定在一系列具有不同氢键（H键）供体能力的带电和中性自组装单分子层（SAM）修饰的金电极上。利用蛋白质膜伏安法（PFV）、计时电流法（CA）和电化学石英晶体微天平（E - QCM）分析确定了决定固定化酶活性和稳定性的关键因素。解析了静电相互作用和氢键相互作用，其中静电相互作用决定酶的取向，而当酶 - 电极界面存在氢键时，酶的解吸受到强烈限制。相反，在没有氢键的情况下，酶的稳定性会急剧降低，并且通过CA期间的E - QCM证实解吸导致的酶损失是活性衰减的主要原因。这项研究深入探讨了固定化氧化还原酶活性降低的可能原因以及膜损失（特别是氢键）在稳定生物电极性能方面的作用，为未来生物电催化的改进提供了途径。
