A simple method to engineer a protein-derived redox cofactor for catalysis

The 6 ×-Histidine tag which is commonly used for purification of recombinant proteins was converted to a catalytic redox-active center by incorporation of Co^{2 +}. Two examples of the biological activity of this engineered protein-derived cofactor are presented. After inactivation of the natural diheme cofactor of MauG, it was shown that the Co ^{2 +}-loaded 6 × His-tag could substitute for the hemes in the H₂O₂-driven catalysis of tryptophan tryptophylquinone biosynthesis. To further demonstrate that the Co^{2 +}-loaded 6 × His-tag could mediate long range electron transfer, it was shown that addition of H₂O₂ to the Co^{2 +}-loaded 6 × His-tagged Cu^{1 +} amicyanin oxidizes the copper site which is 20 Å away. These results provide proof of principle for this simple method by which to introduce a catalytic redox-active site into proteins for potential applications in research and biotechnology.