ATP synthase catalyzes ATP synthesis at the expense of an electrochemical ion gradient across a membrane that can be generated by different exergonic reactions. Sulfur reduction is the main energyyielding reaction in the hyperthermophilic strictly anaerobic Crenarchaeon Ignicoccus hospitalis. This organism is unusual in having an inner and an outer membrane that are separated by a huge intermembrane compartment. Here we show, on the basis of immuno-EM analyses of ultrathin sections and immunofluorescence experiments with whole I. hospitalis cells, that the ATP synthase and H 2 :sulfur oxidoreductase complexes of this organism are located in the outer membrane. These two enzyme complexes are mandatory for the generation of an electrochemical gradient and for ATP synthesis. Thus, among all prokaryotes possessing two membranes in their cell envelope (including Planctomycetes, Gram-negative bacteria), I. hospitalis is a unique organism, with an energized outer membrane and ATP synthesis within the periplasmic space. In addition, DAPI staining and EM analyses showed that DNA and ribosomes are localized in the cytoplasm, leading to the conclusion that in I. hospitalis energy conservation is separated from information processing and protein biosynthesis. This raises questions regarding the function of the two membranes, the interaction between these compartments, and the general definition of a cytoplasmic membrane.Archaea | ATP synthase | ATPase | immunolabeling | sulfur reductase

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Bacterial motility is driven by the rotation of flagellar filaments which supercoil. The supercoiling involves the switching of coiled-coil protofilaments between two different states. In archaea, the flagellar filaments responsible for motility are formed by proteins with distinct homology in their N-terminal portion to bacterial Type IV pilins. The bacterial pilins have a single N-terminal hydrophobic α-helix, not the coiled-coil found in flagellin. We have used cryo-EM to study the adhesion filaments from the archaeon Ignicoccus hospitalis. While I. hospitalis is non-motile, these filaments make transitions between rigid stretches and curved regions, and appear morphologically similar to true archaeal flagellar filaments. A resolution of ~ 7.5 Å allows us to unambiguously build a model for the packing of these N-terminal α-helices, and this packing is different from several bacterial Type IV pili whose structure has been analyzed by electron microscopy and modeling. Our results show that the mechanism responsible for the supercoiling of bacterial flagellar filaments cannot apply to archaeal filaments.