Modified redox proteins for biopolymer modification. The biopolymer lignin is naturally digested under ambient conditions by lignin peroxidases (LiPs), a transformation yet unrealized on an industrial scale. Application of LiPs on large scales is hampered by low yielding LiP expression. We are developing cytochrome c peroxidase (CcP)-based analogs of LiPs; the two enzymes show similar tertiary structure and some sequence homology, but standard CcP expressions yield large amounts of soluble protein. We will explore heme redox chemistry and reactions with organic models and with natural lignin, to develop new systems for applications in lignin digestion or treatment of contaminated waters.
Redox proteins and interfaces. Capture and conversion of energy from sunlight is the most promising way to meet global energy demand. Interfacing light collection machinery with catalysts for fuel production or substrate modification is a highly desirable. Few catalysts are as selective as proteins and new applications continue to emerge. We are developing of methods for attaching photosensitizer-modified redox proteins to transparent conducting electrodes and controlling electron/hole flow to the active sites. Initial work focuses on functionalizing nano-indium tin oxide (nanoITO) with sensitizer-labeled proteins whose intramolecular electron transfer (ET) reactivity is established. Other efforts focus on devices functionalized with cytochrome P450s for hydrocarbon oxidation and reductive dehalogenation. The ultimate goal is to produce robust and efficient photoelectrochemical devices that exploit Nature’s best catalysts.
Artificially tuning protein redox properties. Enzymes tune metal ion reduction potentials over a range of >1 V and catalyze an array of challenging redox transformations, but we are still learning about how this is possible. Taking a page from small molecule inorganic chemistry, we envision small protein scaffolds as “ligands.” Our initial efforts are focuses on active site mutations in three proteins: Cu-containing rusticyanin from T. ferrooxidans; [FeS]-containing rubredoxin from P. furiosus; and heme-containing cytochrome c from yeast, which are all simple to express, tolerate a variety of mutations, and can be crystallographically characterized. Redox potential tuning, reactivity with small molecules at exposed sites, and the effects of metal substitution are being explored. Emphasis will be placed on frustrated coordination geometries reactivity with substrates such as O2, CO, CO2, and Cl2O–.
Biological proton-coupled electron transfer in modified proteins. Biological redox chains rely on long-range proton-coupled electron transfer (PCET) through amino acids, but there are few simple models based on natural systems that are being used to bridge the gap between small molecule, biological, and theoretical work. We are developing modified protein scaffolds for investigation of PCET reactions of protein-bound amino acids. Artificial phenol-Brønsted base (PhOH-B) moieties will be introduced into modified blue-copper proteins using covalent modification of a surface cysteine residue. We will use time-resolved spectroscopies to probe the kinetics of PhOH-B oxidation by surface-attached Ru oxidants. The kinetic effects of distance and redox pathways will be probed. Finally, apo-cupredoxins, which have Cys hydrogen bonded to His in the metal binding site, will enable unprecedented investigations of the redox properties of Cys in a controllable protein environment.