| Summary: | <p>Abstract</p> <p>Background</p> <p>Conversion of industrial processes to more nature-friendly modes is a crucial subject for achieving sustainable development. Utilization of hydrogen-oxidation reactions by hydrogenase as a driving force of bioprocess reaction can be an environmentally ideal method because the reaction creates no pollutants. We expressed NAD-dependent alcohol dehydrogenase from <it>Kluyveromyces lactis</it> in a hydrogen-oxidizing bacterium: <it>Ralstonia eutropha</it>. This is the first report of hydrogen-driven <it>in vivo</it> coupling reaction of the alcohol dehydrogenase and indigenous soluble NAD-reducing hydrogenase. Asymmetric reduction of hydroxyacetone to (<it>R</it>)-1,2-propanediol, which is a commercial building block for antibacterial agents, was performed using the transformant as the microbial cell catalyst.</p> <p>Results</p> <p>The two enzymes coupled <it>in vitro</it> in vials without a marked decrease of reactivity during the 20 hr reaction because of the hydrogenase reaction, which generates no by-product that affects enzymes. Alcohol dehydrogenase was expressed functionally in <it>R. eutropha</it> in an activity level equivalent to that of indigenous NAD-reducing hydrogenase under the hydrogenase promoter. The hydrogen-driven <it>in vivo</it> coupling reaction proceeded only by the transformant cell without exogenous addition of a cofactor. The decrease of reaction velocity at higher concentration of hydroxyacetone was markedly reduced by application of an <it>in vivo</it> coupling system. Production of (<it>R</it>)-1,2-propanediol (99.8% e.e.) reached 67.7 g/l in 76 hr with almost a constant rate using a jar fermenter. The reaction velocity under 10% P<sub>H2</sub> was almost equivalent to that under 100% hydrogen, indicating the availability of crude hydrogen gas from various sources. The <it>in vivo</it> coupling system enabled cell-recycling as catalysts.</p> <p>Conclusions</p> <p>Asymmetric reduction of hydroxyacetone by a coupling reaction of the two enzymes continued in both <it>in vitro</it> and <it>in vivo</it> systems in the presence of hydrogen. The <it>in vivo</it> reaction system using <it>R. eutropha</it> transformant expressing heterologous alcohol dehydrogenase showed advantages for practical usage relative to the <it>in vitro</it> coupling system. The results suggest a hopeful perspective of the hydrogen-driven bioprocess as an environmentally outstanding method to achieve industrial green innovation. Hydrogen-oxidizing bacteria can be useful hosts for the development of hydrogen-driven microbial cell factories.</p>
|
|---|