HYDRIDE · HYDRIDE: Hydrogenase Driven H2 production through Design and Evolution

Climate change is shaping up to be the greatest existential threat that humanity has ever faced. To combat climate change and the corresponding energy crisis, greenhouse gas emissions must be substantially reduced by 2030 while developing non-fossil future fuel alternatives, such as hydrogen. Hydrogen is highly versatile as it can be used in both fuel cells and electricity production. However, currently >95% of the global hydrogen production is fossil fuel-based and not sustainable. Biotechnological hydrogen production is realized through microorganisms harbouring hydrogenases and represents a promising alternative to expand the proportion of sustainable hydrogen within the global budget. However, the utilization of these enzymes is limited by various mechanisms, including inhibition by oxygen, making biohydrogen in its current state not economically sustainable.HYDRIDE is an interdisciplinary study aiming to overcome the oxygen sensitivity of [FeFe]- hydrogenases by designing and evolving high-performance hydrogenase enzymes. This will be achieved by 2 major steps: 1) Using sequence data, ancestral enzymatic scaffolds (AEC) of [FeFe]-hydrogenases will be designed and characterized towards their ability to produce hydrogen, their oxygen sensitivity and active site characteristics. AECs have been shown to provide good starting points for laboratory evolution. Thus, these ACEs will be 2) evolved to overcome low hydrogen production under oxygen exposure using a high-throughput selection system. Consequently, HYDRIDE will tackle one of the major bottlenecks for the utilization of [FeFe]-hydrogenases, paving the way for sustainable and economical biotechnological hydrogen production. While addressing two societal UN sustainability goals, the knowledge gained from this study will substantially advance other scientific fields, aiding e.g., the development and design of artificial catalysts.