Hydrogenases catalyse the reversible oxidation and production of H₂. They have been the subject of intense interest in recent years since these enzymes, or catalysts inspired by them, may greatly enhance our exploitation of H₂ as an energy carrier in a future ‘green’ H₂-based economy. However, a major challenge to the future use of these catalysts is their reactions with small molecule inhibitors, such as O₂ or CO. This thesis presents studies using Protein Film Electrochemistry, in which an enzyme is adsorbed onto an electrode to give an electroactive film. Although most hydrogenases are inhibited or irreversibly damaged by even trace O₂, certain O₂-tolerant hydrogenases are unusual in that they are able to sustain H₂ oxidation activity in the presence of O₂. Results outlined in this thesis suggest that the O₂ tolerance of the membrane-bound [NiFe]-hydrogenase from Ralstonia eutropha relies upon O₂ attack generating exclusively the ‘Ready’ inactive state (formed by complete, four-electron reduction of O₂), which subsequently reactivates both rapidly and at high potential. The results contributed to a new explanation for how hydrogenases in certain microbes survive O₂. Electrochemical studies performed on a variant enzyme suggest that a modified proximal FeS cluster plays a role in conferring this O₂ tolerance. Studies of an enzymatic H₂/O₂ fuel cell employing the O₂-tolerant [NiFe]-hydrogenase Hyd1 from Escherichia coli as the anodic catalyst highlight the subtle influence of the reactions of the hydrogenase with O₂ on the power characteristics of the fuel cell under various operating conditions. This research also identifies straight-chain aldehydes as unprecedented inhibitors of H₂ production by the [FeFe]-hydrogenases. However, some of these results cannot currently be made freely available as they are to be published at a later date in academic journals.

Parmi les options en cours d’investigation, le stockage souterrain de l'hydrogène dans les formations sédimentaires comme les grès pourrait offrir un potentiel unique pour stocker de grandes quantités d'énergie. L'évaluation des modalités de stockage souterrain de l'hydrogène nécessite donc à la fois une connaissance précise des transformations minéralogiques dues à la présence de l'hydrogène et l’acquisition de données sur le comportement hydrodynamique des fluides. Par conséquent, cette étude se composera de trois parties : 1- Etude des interactions géochimiques de l’hydrogène dans des formations sédimentaires gréseuses : Les produits expérimentaux portent la marque d'une réaction très limitée entre les minéraux du grès et l'hydrogène. Si les résultats expérimentaux sont combinés aux résultats numériques, l’étude démontre que l'hydrogène, une fois injecté, peut être considéré comme relativement inerte. De façon globale, nos résultats renforcent la faisabilité du confinement de l'hydrogène dans des réservoirs géologiques comme les grès. 2- Etude de la migration de l'hydrogène dans les grès : détermination de la perméabilité relative et de la pression capillaire du système hydrogène-eau : Afin de fournir des données quantitatives pour le développement du stockage souterrain de l'hydrogène, la pression capillaire et la perméabilité relative ont été mesurées pour le système hydrogène-eau en deux conditions potentielles. Les résultats indiquent que les données obtenues sont applicables à l’ensemble des conditions de stockage de l'hydrogène. 3- Modélisation numérique d’un site de stockage géologique d’hydrogène : La simulation numérique a été effectuée pour caractériser l'évolution dynamique d’un site de stockage d'hydrogène pur. Une fluctuation saisonnière du fonctionnement du réservoir et l'effet des fuites d'hydrogène dus aux réactions ont été pris en compte. / Underground hydrogen storage has been introduced as storage solution for renewable energy systems as it offers a unique potential to store large amounts of energy, especially in sedimentary formations such as sandstones. However, evaluating the underground hydrogen storage requires a precise knowledge of the hydrodynamic behavior of the fluids and of mineralogical transformations due to the presence of hydrogen that may affect the storage properties. Therefore, this study is consists in three parts: 1- Study of geochemical reactivity of hydrogen in sandstone sedimentary formations: The experimental products bear the mark of only very limited reaction between sandstone minerals and hydrogen. Taken together with the numerical results, this study demonstrates that hydrogen, once injected, can be considered as relatively inert. Overall, our results support the feasibility of hydrogen confinement in geological reservoirs such as sandstones. 2- Study of the migration of hydrogen in sandstone: determination of relative permeability and capillary pressure of hydrogen-water system: To provide quantitative data for the development of underground hydrogen storage, capillary pressures and relative permeabilities of hydrogen-water system have been measured at two potential conditions. The interpretation of the results would suggest that the obtained data are applicable for the entire range of hydrogen storage conditions. Interfacial tensions and contact angles for the hydrogen-water system have been also derived. 3- Numerical simulation of a geological hydrogen storage site: The numerical simulation was performed to characterize the evolution of pure hydrogen storage, by considering the seasonal fluctuation of renewable energy and the effect of hydrogen loses due to the biotic reactions.