Understanding the local chemical environment of bioelectrocatalysis.

Bioelectrochemistry employs an array of high-surface-area meso- and macroporous electrode architectures to increase protein loading and the electrochemical current response. While the local chemical environment has been studied in small-molecule and heterogenous electrocatalysis, conditions in enzyme electrochemistry are still commonly established based on bulk solution assays, without appropriate consideration of the nonequilibrium conditions of the confined electrode space. Here, we apply electrochemical and computational techniques to explore the local environment of fuel-producing oxidoreductases within porous electrode architectures. This improved understanding of the local environment enabled simple manipulation of the electrolyte solution by adjusting the bulk pH and buffer pKa to achieve an optimum local pH for maximal activity of the immobilized enzyme. When applied to macroporous inverse opal electrodes, the benefits of higher loading and increased mass transport were employed, and, consequently, the electrolyte adjusted to reach -8.0 mA ⋅ cm-2 for the H2 evolution reaction and -3.6 mA ⋅ cm-2 for the CO2 reduction reaction (CO2RR), demonstrating an 18-fold improvement on previously reported enzymatic CO2RR systems. This research emphasizes the critical importance of understanding the confined enzymatic chemical environment, thus expanding the known capabilities of enzyme bioelectrocatalysis. These considerations and insights can be directly applied to both bio(photo)electrochemical fuel and chemical synthesis, as well as enzymatic fuel cells, to significantly improve the fundamental understanding of the enzyme-electrode interface as well as device performance.