Hydrogenase-driven ATP synthesis from air

All cells require a continuous supply of the universal energy currency, adenosine triphosphate (ATP), to drive countless cellular reactions. The universally conserved F1Fo-ATP synthase regenerates ATP from ADP and Pi by harnessing a transmembrane electrochemical proton gradient (pmf). Bacteria have evolved a wide diversity of pmf-forming proteins to make ATP using light, organic, and inorganic energy sources. Recently, we proposed that many bacteria survive using atmospheric trace gases to produce ATP when limited for other energy sources. However, there is no direct proof that atmospheric energy sources are sufficient to generate pmf or drive ATP synthesis. Here, we show that the membrane-associated hydrogen:quinone oxidoreductase Huc from Mycobacterium smegmatis enables ATP synthesis from air. Purified Huc couples H2 oxidation to the reduction of various ubiquinone and menaquinone analogues. We designed and optimised a minimal respiratory chain in which Huc is reconstituted into liposomes with a pmf-generating terminal oxidase and the ATP-generating F1Fo-ATP synthase. Our experiments show that passive hydrogen exchange from air to solution is sufficient for the electron transfer and pmf generation required to accumulate ATP. Finally, by combining continuous culture bioenergetics measurements with theoretical calculations, we show this process is sufficient for mycobacteria to sustain pmf and ATP synthesis (two ATP molecules per H2 oxidised) for maintenance energy requirements during nutrient starvation. These findings prove that atmospheric energy sources are dependable lifeline substrates that enable continuous energy conservation during nutrient starvation. In addition, this work provides a new tool for ATP production in synthetic applications, which unlike other approaches is traceless without by-product accumulation.