We developed a self-assembling protein hydrogel for enzyme immobilization on electrodes for enzymatic biofuel cell applications.  The building blocks of this hydrogel are two protein block copolymers (PC1 and PC2). PC1 contains a trimer protein fused to an engineered PDZ protein, and PC2 contains the same trimer protein and an engineered peptide ligand of PDZ, as well as a repetitive non-structural S peptide [((AG)3PEG)10] and a docking station peptide SH3lig. The S peptide facilitates the retention of water in the hydrogel and SH3lig enables stable non-covalent immobilization of the SH3 (Src homology 3 domain from the adaptor protein Crk)-modified target protein. Individually, PC1 and PC2 exist as viscous solutions composed of trimeric proteins. Mixing solutions of PC1 and PC2 triggers the physical association of PDZ with its peptide ligand, joining individual PC1 and PC2 trimers and giving rise to the formation of a highly crosslinked protein network (hydrogel). This self-assembled protein hydrogel is highly stable in aqueous solution with <20% total mass loss after 7 days in room temperature. To demonstrate the utility of this hydrogel for enzyme immobilization, two recombinant proteins, PTDH (phosphite dehydrogenase) and Slac (laccase), were purified as fusions to the SH3 protein and immobilized individually in the hydrogel via interaction with the SH3lig. A very low leaching rate of the target protein from the hydrogel in solution was observed with <15% enzyme leaching after 7 days in room temperature. The Slac-containing hydrogel was subsequently rendered conductive by incorporation of carbon nanotube and was able to generate a high current in the mA range. The potential of this hydrogel for power generation will be discussed.

Self-assembling protein hydrogel for enzyme incorporation onto electrodes in biofuel cells

245th National Meeting (2013)

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