Key role of hydrogen in regulating hydrogenases and the reductive TCA cycle in a thermophilic, autotrophic sulfur-reducing bacterium

Many thermophilic autotrophs in deep-sea hydrothermal vents are anaerobes that require H 2 for growth. However, our understanding of how non-methanogenic thermophilic hydrogenotrophs adapt to low H 2 conditions is nascent. In this study, the thermophilic autotrophic thiosulfate-reducing bacterium Desulfurobacterium thermolithotrophum HR11 was grown at 72°C in a chemostat at 2 and 43 µM aqueous H 2 concentrations to quantify changes in growth and H 2 S production kinetics and in its proteome in response to H 2 availability. At 2 μM H 2 , cultures showed a decrease in specific growth rate, maximum cell concentration, and H 2 S production while cell yield (cells produced per mol of H 2 S produced) was unchanged. Differential proteomic analysis identified 79 and 66 proteins having more than twofold higher and lower abundances ( P adj < 0.01), respectively, under 2 μM H 2 conditions out of 887 total detected proteins. Among these, a membrane-bound ferredoxin-dependent [NiFe] hydrogenase and a cytoplasmic NAD(P) + -dependent [NiFe] hydrogenase increased in cells grown on 2 μM H 2 . Also, on 2 μM H 2 , a membrane-bound menaquinone-dependent [NiFe] hydrogenase increased and likely generated a proton motive force and drove ATP synthesis, while a membrane-bound menaquinone-dependent [FeFe] hydrogenase and a membrane-bound NADH dehydrogenase decreased. Four reductive TCA cycle enzymes used for CO 2 fixation were more abundant on 2 µM H 2 . These data suggest that hydrogenase and CO 2 fixation enzyme abundances are regulated by H 2 availability and support the idea of a shift in electron disposal toward CO 2 reduction and assimilation on low H 2 . IMPORTANCE Understanding how thermophilic, autotrophic sulfur-reducing bacteria such as Desulfurobacterium thermolithotrophum adapt to varying H 2 concentrations is important for understanding competition and survival strategies in energy-limited subseafloor hydrothermal environments. In this study, D. thermolithotrophum showed metabolic adaptations under low H 2 conditions that may provide them with a competitive growth advantage in hydrothermal vent ecosystems when H 2 is limited and other thermophilic hydrogenotrophs, such as methanogens, are also commonly found. This work also demonstrated the dynamic relationship between different types of hydrogenases and how they are coordinated with each other and other key pathways such as CO 2 fixation.