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

network of different cellular appendices. The microcolonies wereremarkably stable with almost 100% of viable cells after three month ofincubation.Our analysis demonstrated that the ability of adhesion is widely distributedin haloarchaea and the multicellular communities detected represent biofilmstructures.[1] Tripepi M. et al (2010): J Bacteriol.192(12): 3093-102.ARV008Assessment of the predominant methanogenic pathwaysin anaerobic digesters by the combination of moleculartechniques with the isotopic fingerprinting of theproduced biogasM. Nikolausz* 1 , R.F.H. Walter 1 , H. Sträuber 1 , J. Liebetrau 2 , T. Schmidt 2 ,S. Kleinsteuber 3 , F. Bratfisch 4 ,U.Günther 4 , H.H. Richnow 41 Department of Bioenergy, Helmholtz Center for Environmental Research -UFZ, Leipzig, Germany2 Department of Biochemical Conversion, German Biomass Research Center(DBFZ), Leipzig, Germany3 Department of Environmental Microbiology, Helmholtz Center forEnvironmental Research (UFZ) , Leipzig, Germany4 Department of Isotope Biogeochemistry, Helmholtz Center forEnvironmental Research (UFZ), Leipzig, GermanyLaboratory scale continuously stirred tank reactors were run under variousconditions using either cereal distillers grains, a by-product from bioethanolindustry, maize silage or chicken manure as substrate. In addition to thestandard process parameters the stable hydrogen and carbon isotopiccomposition of the produced biogas (methane and CO 2) was also analysed toestimate the predominant methanogenic pathways (acetotrophic vs.hydrogenotrophic). The methanogenic communities in the reactors were alsoinvestigated for their phylogenetic composition by terminal restrictionfragment length polymorphism analysis and sequencing of the mcrA genescoding methyl-coenzyme M reductase. In addition, the expression of thegene was also studied as a better indicator of the metabolic activity. Thecarbon isotopic values (δ 13 C) of methane ranged between -68‰ and -35‰.This latter value of the maize silage reactor was probably influenced by theoriginal high value (-12‰) of this C4 plant substrate. The hydrogen isotopicvalues (δD) of methane were very low (-369 to -347‰) except the samplesfrom the maize silage reactor ranging from -292‰ to -281‰. Apparentfractionation factors (α CO2-CH4) suggested a hydrogenotrophic pathway in thechicken manure reactor, while probably both pathways influenced theisotopic signal of derived methane in the other reactors.According to the molecular biological investigations the reactors weredominated by hydrogenotrophic Methanomicrobiales with Methanoculleusas the predominant genus. Sequences affiliated with acetotrophicMethanosetaceae were found only in one cereal distillers grains reactor,while sequences affiliated with Methanosarcinaceae were frequently foundrepresenting less abundant members of the methanogenic communities. AtRNA level major changes in the relative abundance of the amplifiedsequences were observed compared to the results obtained from the isolatedDNA.ARP001Toxicity of methylated Bismuth produced by intestinalmicroorganisms to Bacteroides thetaiotaomicron, adominant member of human intestinal microbiotaB. Bialek*, D. Pieper, R. Diaz-Bone, R. HenselDepartment of Microbiology, University Duisburg-Essen, Essen, GermanyBismuth compounds have significant application in medicine. Bismuthsubcitrate is applied in a triple-therapy for the treatment of Helicobacterpylori which causes chronic inflammation of the stomach and is linked tothe development of duodenal, gastric ulcers and stomach cancer. Afteringestion of bismuth subcitrate, Bi 3+ is methylated to volatiletrimethylbismuth (TMBi) by the intestinal microbiota especially bymethanoarchaea.Here we investigate the influence of TMBi produced by Methanobrevibactersmithii on growth of Bacterioides thetaiotaomicron, an important member ofthe physiological intestinal microbiota. Transfer of TMBi from headspace ofMethanobrevibacter smithii cultures to B. thetaiotaomicron cultures resultsin a significant growth inhibition of this organism. Closer investigationsshowed that the volatile TMBi rapidly decays into soluble dimethyl- andmonomethylbismuth, which cause comparable growth inhibition effects,suggesting that these derivatives are the actual agents of growth inhibition.Analyses are presented, which give insight into possible mechanismsresponsible for the toxic effects of the various methylbismuth derivatives.ARP002The fimbriae of MethanothermobacterthermoautotrophicusC. Sarbu* 1 , R. Rachel 2 , R. Wirth 11 Institute of Microbiology, University of Regensburg, Regensburg, Germany2 Institute of Microbiology and Electron Microscopy, University ofRegensburg, Regensburg, GermanyThe fimbriae of the euryarchaeon Methanothermobacterthermoautotrophicus were among the first detailed characterized archaealfimbriae. We have shown that these cell appendages with a diameter of 5 nm(mainly) consist of 16 kDa glycoprotein Mth60 and function as adhesins.For further analyses Mth60 fimbriae were enriched from the supernatant of a100 liter fermentor by PEG/NaCl induced precipitation, CsCl-gradientcentrifugation and dialysis. Electron microscopy demonstrated that soprepared fimbriae are pure and well-structured. At some ends knob-likepatterns could be detected. SDS-PAGE indicated four different proteins inthe fimbriae fractions; the prominent band at about 16 kDa correspondsMth60. The identification of the other proteins failed up to now.Reverse transcription PCR and Northern Blots revealed the main fimbringene mth60 to be part of two operons: it is co-transcribed with mth58 andmth59, a further operon comprises mth60 and mth61. It is well known frombacteria that all genes necessary for fimbriae formation are clustered.Bioinformatical investigations showed Mth59 to have a significant similarityto bacterial chaperone proteins. Chaperones play an important role infimbriae assembly (chaperone-usher-pathway) of some bacteria. Thehomology of Mth59 to bacterial chaperones might indicate that archaeal andbacterial fimbriae have a related mode of assembly. The now availableMth59 antibodies are applied in co-immunoprecipitation experiments, thusanalyzing the function of this protein. Immunolabeling of ultrathin sectionsof M. thermoautotrophicus will hopefully allow to clarify the localization ofMth59.ARP003Hot protein phosphorylation: carbon source dependentphospho-proteom mapping from Sulfolobus solfataricusP2D. Esser* 1 , T.K. Pham 2 , J. Reimann 3 , S.-V. Albers 3 , P. Wright 2 , B. Siebers 11 Department of Molecular Enzymetechnology and Biochemistry, UniversityDuisburg-Essen, Essen, Germany2 Department of Chemical and Biological Engineering, University ofSheffield, Sheffield, United Kingdom3 Max Planck Institut for Molecular Biology of Archaea, Marburg, GermanyPosttranslational modifications (PTMs) are of major interest for theregulation of cellular processes. Reversible protein phosphorylation is themain mechanism, which is applied to control the functional properties ofproteins in response to environmental stimuli [1]. In the 80’s proteinphosphorylation has been demonstrated in the third domain of life, theArchaea. However, so far only few phospho proteins were identified andfew protein kinases and protein phosphatases were investigated. A hugeprogress was achieved only recently by the determination of the completephospho-proteom of the extremhalophilic Euryarchaeon Halobacterimsalinarium, which was analyzed via MS with prior TiO 2 phospho-peptideenrichment [2].Model organism of this study is the thermoacidophilic CreanarchaeonSulfolobus solfataricus. Bioinformatic investigation revealed that S.solfataricus only harbors eukaryal protein kinases and classical twocomponent systems are absent. Until now, only little is known about proteinphosphorylation in this organism. So far, only six possible target proteinswere reported. In addition three eukaryal type serine/threonine specificprotein kinases as well as two protein phosphatases were characterized (PP1-arch1 and PTP) [1; 3]. In order to analyze the phospho-proteom of S.solfataricus in more detail, we applied a gel and enrichment free proteomeapproach by using the precursor acquisition independent from ion count(PAcIFIC) method [4]. The detailed pospho-proteom mapping with a specialfocus on the central carbohydrate metabolism (CCM) will be presented.spektrum | Tagungsband 2011