It has become relatively easy to design proteins which are extremely rigid but lack the dynamism required for catalytic activity. Here we report the rational redesign of DF2, an extremely rigid, computationally designed homodimeric diiron protein, into the functional hydrogen peroxide oxidase CDM3, a single-chain dimanganese-containing four-helix bundle, by simply increasing the dynamic motion of the protein via the reversal of the amino acid sequence of two of the helices, greatly reducing the packing complementarity in the protein core. The resulting enzyme oxidizes hydrogen peroxide by two electrons forming molecular oxygen at a rate of at least 400 s-1 while only catalyzing the reduction of hydrogen peroxide to water at a rate of 0.08 s-1. Most natural enzymes with peroxide as a substrate are catalases - hydrogen peroxide dismutases which catalyze both reactions at high rates in order to lower the cellular concentration of hydrogen peroxide. Because it favors the oxidizing half-reaction over the other by a factor of 5000, CDM3 promises to serve as a unique alternate version of an oxygen evolving cluster in the design of new photoautotrophic organisms which perform carbon fixation utilizing light and low energy electrons originating from what is typically considered to be a toxic intercellular waste product. Time permitting, we will discuss our demonstration of this in vitro via the utilization of CDM3 as an electron source for our designed light-activated charge separation domain.

Rational design of an artificial hydrogen peroxide oxidase and its use as an electron source for artificial reaction centers

254th National Meeting (2017)

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