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 20Å 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.