Interspecies hydrogen transfer in anoxic ecosystems is essential for the complete microbial breakdown of organic matter to methane. Acetogenic bacteria are key players in anaerobic food webs and have been considered as prime candidates for hydrogen cycling. We have tested this hypothesis by mutational analysis of the hydrogenase in the model acetogen Acetobacterium woodii. Hydrogenase-deletion mutants no longer grew on H 2 + CO 2 or organic substrates such as fructose, lactate, or ethanol. Heterotrophic growth could be restored by addition of molecular hydrogen to the culture, indicating that hydrogen is an intermediate in heterotrophic growth. Indeed, hydrogen production from fructose was detected in a stirredtank reactor. The mutant grew well on organic substrates plus caffeate, an alternative electron acceptor that does not require molecular hydrogen but NADH as reductant. These data are consistent with the notion that molecular hydrogen is produced from organic substrates and then used as reductant for CO 2 reduction. Surprisingly, hydrogen cycling in A. woodii is different from the known modes of interspecies or intraspecies hydrogen cycling. Our data are consistent with a novel type of hydrogen cycling that connects an oxidative and reductive metabolic module in one bacterial cell, "intracellular syntrophy."

Acetogenic bacteria have gained much attraction in recent years as they can produce different biofuels and biochemicals from H 2 plus CO 2 or even CO alone, therefore opening a promising alternative route for the production of biofuels from renewable sources compared to existing sugar-based routes. However, CO metabolism still raises questions concerning the biochemistry and bioenergetics in many acetogens. In this study, we focused on the two acetogenic bacteria Acetobacterium woodii and Thermoanaerobacter kivui which, so far, are the only identified acetogens harbouring a H 2-dependent CO 2 reductase and furthermore belong to different classes of 'Rnf'and 'Ech-acetogens'. Both strains catalysed the conversion of CO into the bulk chemical acetate and formate. Formate production was stimulated by uncoupling the energy metabolism from the Wood-Ljungdahl pathway, and specific rates of 1.44 and 1.34 mmol g −1 h −1 for A. woodii Δrnf and T. kivui wild type were reached. The demonstrated CO-based formate production rates are, to the best of our knowledge, among the highest rates ever reported. Using mutants of Δhdcr, ΔcooS, ΔhydBA, Δrnf and Δech2 with deficiencies in key enzyme activities of the central metabolism enabled us to postulate two different CO utilization pathways in these two model organisms.

No abstract