Redox reactions are at the heart of numerous biological functions and chemical transformations, from electron transfer (ET) in photosynthesis and respiration to catalytic activations of C-H bonds and other molecules. The redox potential (E°) is one of the most important parameter in determining the efficiency of reactions.  For example, with the global energy crisis, there is growing interest among all fields in science in making bio-inspired redox reagents for applications ranging from fuel cells to solar energy conversion. A critical obstacle to success is the availability of redox centers with tunable redox potentials to lower overpotentials and increase catalytic efficiency. Areas such as fuel cell research also often require redox agents that are stable in aqueous solution, agents which are relatively rare.

  For millions of years, biology has operated within the range of physiological E°s, defined by the highest E° (~ +1 V vs. SHE) at which water is oxidized, and the lowest E° (~ -1V) at which protons are reduced to H2.  Amazingly, nature has found a way to cover this wide range using a strikingly limited set of not only metal cofactors, but also protein folds. Despite many years of research into these proteins and efforts to change the E°, how the E°s can be tuned systematically in a wide range using the limited metal cofactors is still not well understood. An ultimate test of our understanding of this process is to design redox centers with the minimum number of metal cofactors and mutations that can cover the entire 2V range of physiological E°. We reported in this PNAS paper (doi: 10.1073/pnas.1515897112) the design of the protein azurin to cover a range from +970 mV to −954 mV vs. standard hydrogen electrode (SHE) by mutating only five residues and using two metal ions. Spectroscopic methods have revealed geometric parameters important for the high E°′.

  This ability to cover the entire range, which even surpasses the observed E°′ range for all ET proteins combined, using two metal ions and mutating only five residues in a single protein scaffold (Fig. 1) is a testimony to how much we now understand the structural features responsible for tuning E°′ of metalloproteins. Given the wide range of potentials attainable from a single protein possessing the same overall fold and surface properties, the azurin variants reported in this study may enable scientists and engineers to take advantage of these water-soluble redox agents for biochemical and biotechnological applications such as solar energy transfer and other alternative energy conversions. Because tuning the potentials of many inorganic, bioinorganic, and organometallic catalysts can result in catalysts with different oxidation states with dramatically different catalytic efficiency for different substrates, the knowledge gained from this study may also allow others to use the same principle to tune redox properties of numerous catalysts for even wider applications, such as small molecule activation and synthesis of important intermediates or products for pharmaceutical applications.

   This research has been supported by NSF through CHE 14-13328 to Yi Lu group at the University of Illinois at Urbana-Champaign, in collaboration with Ninian J. Blackburn’s group from Oregon Health and Science University, who provided x-ray absorption spectral analysis.

  What makes this work unique is we have achieved the ultimate challenge both highest E°′ and lowest E°′ under physiological conditions. Applying the principles demonstrated in this work in other systems can results in novel redox agents and catalysts with unprecedented properties.

   Furthermore, while a number of redox agents are available for applications in organic solvents, water-soluble and stable redox agents are quite limited. To overcome this limitation, we have made many azurin mutants with a wide range of Eº´ available as redox agents to the general scientific community through Kerafast.com (https://www.kerafast.com/product/2721/redox-potential-tuned-azurin-proteins). This NSF-funded research has expanded the number of water-soluble and stable redox agents significantly and will help many other chemists, biochemists and biophysics in their studies to advance many other areas of science and technology.

Last Modified: 08/18/2017
Modified by: Yi Lu