VAAM-Jahrestagung 2012 18.–21. März in Tübingen

72CEP032Yeast mitochondria as a model system to study the biogenesisof Yersinia Adhesin A (YadA)T. Ulrich* 1 , J.E.N. Müller 1 , D. Papic 1 , I. Grin 2 , D. Linke 2 , K.S. Dimmer 1 ,I.B. Autenrieth 3 , D. Rapaport 31 University of Tübingen, Interfaculty Institute for Biochemistry, Tübingen,Germany2 Max-Planck Institute for Developmental Biology, Department of ProteinEvolution, Tübingen, Germany3 University of Tübingen, Interfaculty Institute of Microbiology andInfection Medicine, Tübingen, Germany-barrel proteins are found in the outer membranes of eukaryoticorganelles of endosymbiotic origin as well as in the outer membrane ofGram-negative bacteria. Precursors of mitochondrial -barrel proteins aresynthesized in the cytosol and have to be targeted to the organelle.Currently, the signal that assures their specific targeting to mitochondria ispoorly defined. To characterize the structural features needed for specificmitochondrial targeting and to test whether a full -barrel structure isrequired we expressed in yeast cells the -barrel domain of the trimericautotransporter Yersinia Adhesin A (YadA). Trimeric autotransporters arefound only in prokaryotes where they are anchored to the outer membraneby a single 12- stranded -barrel structure to which each monomer iscontributing 4 -strands. Importantly, we found that YadA is solelylocalized to the mitochondrial outer membrane where it exists in a nativetrimeric conformation. These findings demonstrate that rather than a linearsequence or a complete -barrel structure, four -strands are sufficient forthe mitochondria to recognize and assemble -barrel protein. Remarkably,the evolutionary origin of mitochondria from bacteria enables them toimport and assemble even proteins belonging to a class that is absent ineukaryotes.EMV1-FGDegradation of organic carbon by microorganisms - do weknow the 'rules' and limits?F. WiddelMax Planck Institute for Marine Microbiology, Bremen, GermanyThe postulate of 'microbial inerrancy' states that for every substancesynthesized by organisms there must be at least one type of microorganismable to degrade it. An undegradable biogenic substance would haveaccumulated in earth’s history. This postulate has significantly stimulatedbiodegradation research. For a long time, many compounds with lowchemical reactivity were thought to undergo biodegradation only in thepresence of oxygen. However, during the last two decades or so, metabolictypes of anaerobic microbes observed in habitats or enriched and isolatedin cultures were shown to degrade compounds that formerly wereconsidered recalcitrant under anoxic conditions; a class of such compoundsare, for instance, hydrocarbons in gas and oil. Microorganisms degradingchemically unreactive compounds in anoxic habitats are 'confronted' withtwo challenges, a mechanistic and (often) an energetic one: Bonds may bedifficult to activate, and the net energy gain may be very low, respectively.For experimental investigation, also slowness of the processes may presenta certain obstacle. Still, on a global scale and over geologically relevantperiods, even such slow processes are relevant.EMV2-FGCharacterising oligotrophic bacterial growth with flowcytometryF. HammesEawag, Microbiology, Dübendorf, SwitzerlandMost natural and engineered aquatic environments comprise a broaddiversity of both natural and anthropogenic organic carbon compounds,utilised by an equally broad diversity of indigenous bacterial species. Butcarbon concentrations are typically low. Total biodegradable organiccarbon concentrations below 1 mg/L is common in many lakes, rivers,groundwater and drinking water, and concentrations of individualsubstrates below 1 g/L are normal. Bacterial concentrations in suchenvironments are in direct correlation to available substrate concentrations, andtypically range from 10 2 to 10 6 cells/mL. These concentrations are severalorders of magnitude lower than those usually employed in laboratory basedstudies, and research is further complicated by the diversity in both the carbonresources and the utilising bacteria. Hence, improved methods for analysingbacterial growth are welcomed. Flow cytometry (FCM) is a method particularlysuited for analysis of bacterial growth in these conditions. Firstly, FCM detectsall bacteria, irrespective of cultivability. This allows the study of indigenousbacterial communities that do not grow on conventional nutrient media.Secondly, FCM analysis can provide sensitive data on cell concentrations, cellsize and nucleic acid content, allowing for detailed information on theorganisms in question. Finally, FCM analysis can be automated easily. Thisprovides the opportunity for extensive high resolution analysis of dynamicprocesses such as bacterial growth. This presentation will discuss the use ofFCM in studying (1) indigenous bacterial community growth on naturalassimilable organic carbon (AOC), (2) single species (pathogenic bacteria)growth on natural AOC, and (3) single species growth on specific organiccarbon compounds.EMV3-FGSubstrate use of extremely oligotrophic bacteriaA. Schwedt* 1 , M. Seidel 1,2 , T. Dittmar 1,2 , M. Simon 2 , V. Bondarev 1 ,S. Romano 1 , G. Lavik 1 , H.N. Schulz-Vogt 11 Max Planck Institute for Marine Microbiology, Microbiology, EcophysiologyGroup, Bremen, Germany2 Carl von Ossietzky University of Oldenburg, Institute of Chemistry andBiology of the Marine Environment, Oldenburg, GermanyMarine planktonic bacteria live in habitats that are extremely limited inavailable nutrients, especially the concentration of bioavailable dissolvedorganic compounds is very low and often close to the detection limit.Therefore, it is difficult to study the substrate use of these bacteria underoligotrophic conditions. Very sensitive methods are needed and it is crucialto keep equipment and medium contamination-free to study the physiologyof bacteria proliferating under extremely oligotrophic conditions. Thesubstrate use of Pseudovibriosp. strain FO-BEG1 was investigated inartificial and natural oligotrophic seawater on elemental (dissolved organiccarbon, DOC and total dissolved nitrogen, TDN) and molecular level. Themolecular composition of dissolved organic matter (DOM) wasdetermined by electrospray ionization Fourier transform ion cyclotronresonance mass spectrometry (ESI FT-ICR-MS) and molecular amino acidanalysis. Our data show that the investigated Pseudovibrio strain is able tomultiply from about 20 cells mL -1 to 20,000 cells mL -1 in artificial and to800,000 cells mL -1 in natural seawater. DOC concentrations in artificialseawater were < 5 mol C L -1 and 75 mol C L -1 in natural seawater.During growth no significant decrease in DOC and TDN concentrationswas detectable. Also N 2 and CO 2 fixation could be ruled out as majornitrogen or carbon source. Interestingly, amino acids were not the primarysubstrate for growth in both artificial and natural seawater. Among theseveral thousand compounds detected in seawater, the bacteria were ableto use different organic compounds simultaneously, such as organicsulfonates or aminosugars. Most of the metabolized compounds containednitrogen and thus might serve also as nitrogen source for the bacteria underoligotrophic conditions. Our data demonstrate that many differentsubstrates can be used under extremely oligotrophic conditions at originalconcentrations. Furthermore, growth in artificial seawater was observed,with DOC concentrations much lower than typically detected in naturaloligotrophic seawater.EMV4-FGMicrobial degradation of organic compounds (naturalcompounds, xenobiotics, and pesticides) and the formation ofsoil organic matter and biogenic non-extractable (or bound)residuesM. Kästner*, A. MiltnerHelmholtz-Centre for Environmental Research, EnvironmentalBiotechnology, Leipzig, GermanyDuring microbial degradation, carbon from any biodegradable organiccompound in soil is partitioned into parent compound, metabolites, nonextractableresidues (NER), CO 2, and microbial biomass. This distributionmust be known to assess the fate of the compound in soil, e.g. NER frompesticides are considered to consist of adsorbed and sequestered parentcompounds or metabolites and thus as hazardous residues. However, theymay also partly derive from bacterial biomass, resulting in harmlessbiogenic residues. In addition, the formation of soil organic matter (SOM)or humic compounds has long been a dominating topic in soil sciencebecause the amount and composition of SOM determines soil quality butthe processes are still not yet really understood. The so-called humicsubstances were regarded for a long time as a novel category of crosslinkedorganic materials. However, the genesis and microbial contributionis still poorly understood. In addition, due to decreasing soil organic matter(SOM) contents all over Europe, a proper management of SOM is neededfor maintaining soil fertility and for mitigation of the global increase of theatmospheric CO 2 concentration.Microbial biomass residues could be identified as a significant source forSOM. We incubated 13 C-labelled bacterial cells in an agricultural soil andtraced the fate of the 13 C label of bacterial biomass in soil by isotopicanalysis [1-5]. In the presentation, the mass balance data will besummarized and the microbial biomass and its residues by scanningelectron microscopy (SEM) will be visualized. The results indicate that ahigh percentage of the biomass-derived carbon (in particular fromproteins) remains in soil, mainly in the non-living part of SOM afterextended incubation. The SEM micrographs only rarely show intact cells.Instead, organic patchy fragments of 200-500 nm size are abundantindicating specific disintegration processes of cell walls. These fragmentsare associated with all stages of cell envelope decay and fragmentation.BIOspektrum | Tagungsband 2012