• A systematic proteomic analysis of Listeria monocytogenes house-keeping protein secretion systems.

      Halbedel, Sven; Reiss, Swantje; Hahn, Birgit; Albrecht, Dirk; Mannala, Gopala Krishna; Chakraborty, Trinad; Hain, Torsten; Engelmann, Susanne; Flieger, Antje (2014-11)
      Listeria monocytogenes is a firmicute bacterium causing serious infections in humans upon consumption of contaminated food. Most of its virulence factors are secretory proteins either released to the medium or attached to the bacterial surface. L. monocytogenes encodes at least six different protein secretion pathways. Although great efforts have been made in the past to predict secretory proteins and their secretion routes using bioinformatics, experimental evidence is lacking for most secretion systems. Therefore, we constructed mutants in the main housekeeping protein secretion systems, which are the Sec-dependent transport, the YidC membrane insertases SpoIIIJ and YqjG, as well as the twin-arginine pathway, and analyzed their secretion and virulence defects. Our results demonstrate that Sec-dependent secretion and membrane insertion of proteins via YidC proteins are essential for viability of L. monocytogenes. Depletion of SecA or YidC activity severely affected protein secretion, whereas loss of the Tat-pathway was without any effect on secretion, viability, and virulence. Two-dimensional gel electrophoresis combined with protein identification by mass spectrometry revealed that secretion of many virulence factors and of enzymes synthesizing and degrading the cell wall depends on the SecA route. This finding was confirmed by SecA inhibition experiments using sodium azide. Analysis of secretion of substrates typically dependent on the accessory SecA2 ATPase in wild type and azide resistant mutants of L. monocytogenes revealed for the first time that SecA2-dependent protein secretion also requires the ATPase activity of the house-keeping SecA protein.
    • Metaproteomics to unravel major microbial players in leaf litter and soil environments: challenges and perspectives.

      Becher, Dörte; Bernhardt, Jörg; Fuchs, Stephan; Riedel, Katharina; Ernst-Moritz-Arndt-University of Greifswald, Institute of Microbiology, Greifswald, Germany. (2013-10)
      Soil- and litter-borne microorganisms vitally contribute to biogeochemical cycles. However, changes in environmental parameters but also human interferences may alter species composition and elicit alterations in microbial activities. Soil and litter metaproteomics, implying the assignment of soil and litter proteins to specific phylogenetic and functional groups, has a great potential to provide essential new insights into the impact of microbial diversity on soil ecosystem functioning. This article will illuminate challenges and perspectives of current soil and litter metaproteomics research, starting with an introduction to an appropriate experimental design and state-of-the-art proteomics methodologies. This will be followed by a summary of important studies aimed at (i) the discovery of the major biotic drivers of leaf litter decomposition, (ii) metaproteomics analyses of rhizosphere-inhabiting microbes, and (iii) global approaches to study bioremediation processes. The review will be closed by a brief outlook on future developments and some concluding remarks, which should assist the reader to develop successful concepts for soil and litter metaproteomics studies.