VAAM-Jahrestagung 2011 Karlsruhe, 3.–6. April 2011

ARV004Subcellular organization and energy conservation ofIgnicoccus hospitalisL. Kreuter* 1 , U. Küper 1 , T. Heimerl 2 , A. Röhl 1 , F. Mayer 1,3 , V. Müller 3 ,R. Rachel 2 , H. Huber 11 Institute for Microbiology and Archaeal Center, University of Regensburg,Regensburg, Germany2 Center for Electron Microscopy, University of Regensburg, Regensburg,Germany3 Institute for Molecular Biosciences, Goethe-University, Frankfurt amMain, GermanyIgnicoccus hospitalis is a chemolithoautotrophic Crenarchaeote that obtainsenergy from the reduction of elemental sulfur with molecular hydrogen aselectron donor (1). It is able to carry out CO 2 fixation via a new pathway,named dicarboxylate/ 4-hydroxybutylate cycle. Acetyl-CoA is the primaryacceptor molecule and is regenerated via the characteristic intermediate 4-hydroxybutyrate (2). I. hospitalis, like all identified Ignicoccus species,exhibits a unique cell architecture that differs from all other Archaea knownso far. Its cell envelope consists of two membranes enclosing a hugeintermembrane compartment (IMC) (3). In its lipid composition, the outermembrane of I. hospitalis significantly differs from the cytoplasmicmembrane, as it comprises only archaeol and its derivatives, but nocaldarchaeol. In addition, there are unique and abundant proteins only foundin the outer membrane of I. hospitalis, like the pore-forming Ihomp1.Recently, it was shown that the outer membrane contains the H 2:sulphuroxidoreductase as well as the ATP synthase. Thus, I. hospitalis is the firstorganism with an energized outer membrane and ATP synthesis within theIMC. DAPI staining and EM analyses showed that DNA and ribosomes arelocalized in the cytoplasm, leading to the conclusion that in I. hospitalisenergy conservation is separated from information processing and proteinbiosynthesis (4). In addition, we were able to demonstrate that the acetyl-CoA synthetase that activates acetate to acetyl-CoA in an ATP consumingprocess is associated to the outer membrane. This is the first energyconsumingprocess proven to take place in the intermembrane compartment.These results raise questions on other metabolic reactions that are likely tooccur in the IMC, e.g. the first steps in CO 2 fixation, and on the existence oftransporters that convey ATP from the site of its synthesis to the cytoplasmwhere DNA replication and transcription take place. The findings may alsoshed light on the nature of the intimate association between I. hospitalis andNanoarchaeum equitans (5). It is known that N. equitans receives aminoacids and lipids from its host. However, it is still unclear at present if N.equitans is able to synthesize ATP or if it obtains this form of energydirectly from I. hospitalis, too.[1] Paper W. et al (2007): Int. J. Syst. Evol. Microbiol. 57:803-808.[2] Huber H. et al (2008): PNAS 105: 7851-7856.[3] Junglas B. et al (2008): Arch. Microbiol. 190: 395-408.[4] Kueper U. et al (2010): PNAS 107: 3152-3156.[5] Jahn U. et al (2008): J. Bacteriol. 190: 1743-1750.ARV005Full speed ahead: analysis of the assembly of the archaealflagellumK. Lassak* 1 , A. Ghosh 1 , R. Wirth 2 , S.-V. Albers 11 Department of Molecular Microbiology, Max Planck Institute forTerrestrial Microbiology, Marburg, Germany2 Department of Microbiology, University of Regensburg, Regensburg,GermanyMicroorganisms move towards optimum locations and escape fromunfavorable conditions by use of motility structures like flagella.The archaeal flagellum, which is distinct from the bacterial one, was studiedintensively in Euryarchaeota. However, the crenarchaeal flagellar assemblysystem is not well understood. We study the thermoacidophilic crenarchaeonSulfolobus acidocaldarius to analyse the assembly and function of theirflagella system. Markerless in-frame deletion strains were constructed for allseven genes of the fla operon. To exclude polar effects, both at thetranscriptional and translational level, we performed q-PCRs and WesternBlots. Motility assays and electron microscopy analysis of all Δfla strainsrevealed non-motile and non-flagellated S. acidocaldarius cells. Takentogether, these results indicate the involvement of all seven genes of the flaoperon in the correct flagellar assembly.In a parallel approach pH, osmotic pressure, temperature and starvation weretested to stimulate flagellar biosynthesis and assembly. Interestingly, onlystarvation conditions induced the production of flagellar assembly proteins.Moreover, under these conditions thermo microscopy revealed highly motilecells, reaching swimming velocities up to 60 μm/s. Thus, we speculate thatthe crenarchaeal flagellum plays a role in escaping from nutrient limitedenvirons.Further experiments like pull-down assays and yeast two-hybridexperiments will elucidate Fla protein interactions. These findings will bethe basis to understand the molecular mechanism of the crenarchaealflagellar assembly system.ARV006Microorganisms, Peregrine Falcons and Cheetahs - Whois the fastest?R. Wirth*, B. HerzogInstitute for Microbiology and Archaeal Center, University of Regensburg,Regensburg, GermanyAn often asked question in Biology concerns velocity: who is the fastest?An answer to this question has to take into account the different body sizesof organisms to be compared. For this reason the term bps was introduced as‘bodies per second'; i. e. relative velocities are defined in movements ofbody size per second. By this definition man runs with ca. 5 bps, cheetahsrun with a maximum speed of 30 bps; the peregrine falcon flies at ca. 100bps and accelerates by dives into the air to a maximal 400 bps. This lattervelocity often is referred to as maximum relative speed in nature (but is notreached by active movement).Using a so-called thermomicroscope, allowing analyses at up to 95° C underanaerobic conditions we analyzed the swimming behavior ofhyperthermophilic Archaea (some bacteria were used for controls). Our dataclearly show that certain microorganisms are the fastest organisms on earth.E. coli swims with an average speed of ca. 45 μm/sec = ca. 30 bps. ForArchaea we measured the following speeds: Halobacterium salinarum: 3μm/sec = ca. 1 bps; Pyrococcus furiosus 60 μm/sec = 60 bps;Methanococcus voltae: 80 μm/sec = 80 bps; Methanocaldococcus villosus:290 μm/sec = 290 bps; Methanocaldococcus jannaschii: 380 μm/sec = 380bps. The latter two species did swim with maximal relative velocities of 470and 590 bps, respectively - they for sure, thereby extend the maximumrelative speed in nature by at least a factor of 5.Very interestingly, the swimming behavior of hyperthermophilic Archaeadiffers from that of mesophilic Bacteria. Whilst the latter swim in more orless smooth runs, the former exhibit a ‘seek and stay' behavior, which mightbe explained by the conditions they life in. Examples of those differentswimming modes will be presented.ARV007Screening and characterization of biofilm formation inhalophilic ArchaeaS. Fröls*, F. PfeiferInstitute of Microbiology and Genetics, Unviersity of Technology,Darmstadt, GermanyBiofilm formation is described for some hyperthermophilic andmethanogenic Archaea, only one example of surface adhesion is reported forhaloarchaea [1]. We developed a fluorescence-based adhesion assay toscreen and quantify this property in haloarchaea. Eight extremely halophilicHalobacterium salinarum strains, four moderately halophilic Haloferaxstrains, one haloalkaliphilic strain and eight halopsychrophilic archaea weretested. Twelve of them showed adhesion, that were categorized in fourgroups. Members of group I did not adhere (0%-10%), group II exhibited alow to moderate (>10%-30%), group III a strong (>30%-70%), and group IVa very strong ability for adhesion (>70%). The latter group contains oneAntarctic isolate and the gas vesicle producing Hbt. salinarum DSM3754,whereas the gas vesicle producing wildtype strains PHH1 and NRC-I did notadhere. Among the environmental isolates only half of them were adhesiveand adhesion could also get lost after several rounds of incubation.Biofilm producing strains cultivated on glass and plastic surfaces wereanalyzed by microscopy. Different growth parameters or variations of mediasupplements had almost no effect on biofilm formation. Biofilms of Hbt.salinarum DSM3754 and the Antarctic isolate were composed of flat celllayers with additional three-dimensional microcolonies. In contrast, biofilmsof Haloferax and Halorubrum mainly consisted of large cellular aggregatesthat loosely attached to the surface. Extracellular polymeric substances(EPS) were composed of free nucleic acids and glycoconjugates. In the caseof Hbt. salinarum DSM3754 attachment started one day after incubation.Electron microscopic studies showed that adherent cells were connected by aspektrum | Tagungsband 2011