Arthur J. Schmitt Professor of Chemical and Biomolecular Engineering; Professor of Chemistry & Biochemistry; Director, Advanced Diagnostics and Therapeutics
Electron transfer reactions are fundamental to life processes, such as respiration, vision, and energy catabolism, so it is critically important to understand the relationship between functional states of individual redox enzymes and the macroscopically observed phenotype, which results from averaging over all copies of the same enzyme, encompassing varying levels of catalytic activity. To address this problem, we are exploring a bifunctional nanoelectrochemical-nanophotonic architecture - the electrochemical zero mode waveguide (E-ZMW)) - that can couple biological electron transfer reactions and luminescence. The E-ZMW combines the capacity to trap electromagnetic radiation with the ability to control electrochemical potential in its sub-attoliter total volume. Single copies of redox enzyme molecules are immobilized in E-ZMW nanopores at the surface of a metal annulus that can function both as a working electrode, controlling the potential at the enzyme, and as the optical cladding layer of a ZMW. We are currently developing E-ZMW architectures capable of supporting potential controlled single molecule redox reactions with a variety of oxidoreductase enzymes. Parallel arrays of electrochemically-active single molecule "beakers" are being fabricated, and we then use the WE/OC for potential-control of enzyme redox state, and measure the effectiveness of doing so by characterizing the potential-dependent single molecule fluorescence dynamics, for example, measuring single reaction turnover events. Furthermore, the confined environment of the E-ZMW makes it possible to achieve in situ control over reaction conditions and delivery of reactants by elaborating the basic E-ZMW architecture to obtain a dual-electrode nanopore structure with the capacity to synthesize and deliver substrate molecules in situ and on-demand. We are exploiting this capability to characterize single enzyme reactions with reactive oxygen species. The unique capabilities being developed in our laboratory will open new avenues for coupled electrochemical and spectroscopic investigations of single enzyme molecules occurring under tightly controlled conditions.
- "Single Molecule Enzyme Dynamics of Monomeric Sarcosine Oxidase in a Au-Based Zero-Mode Waveguide" Zhao, J.; Branagan, S.P.; Bohn, P.W. Appl. Spectrosc., 2012, 66, 163-169.
- "Potential-Dependent Single Molecule Blinking Dynamics for Flavin Adenine Dinucleotide Covalently Immobilized in Zero-Mode Waveguide Array of Working Electrodes” Zhao, J.; Zaino, L.P.; Bohn, P.W. Faraday Disc. 2013, 164, 57-69.
- "Single Molecule Spectroelectrochemistry of Freely Diffusing Flavin Mononucleotide in Zero-Dimensional Nanophotonic Structures” Zaino, L.P. III; Grismer, D.A.; Han, D.; Crouch, G.M.; Bohn, P.W. Faraday Disc.2015, 184, 101-115.
- "Single-Molecule Spectroelectrochemical Cross-Correlation During Redox Cycling in Recessed Dual Ring Electrode Zero-Mode Waveguides” Han, D.; Crouch, G.M.; Fu, K.; Zaino, L.P. III; Bohn, P.W. Chem. Sci. 2017, 3, 5345-5355.