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

97enzyme product proved unequivocally ChaDH as NAD-dependentchanoclavine-I dehydrogenase like its homologue FgaDH.[5][1.] C. Wallwey, S.-M. Li, Nat. Prod. Rep. 2011, 28, 496-510.[2.] C. L. Schardl, D. G. Panaccione, P. Tudzynski, The Alkaloids, Chem. Biol. 2006, 63, 45-86.[3.] C. Wallwey, M. Matuschek, X.-L. Xie, S.-M. Li, Org. Biomol. Chem. 2010, 8, 3500-3508.[4.] M. Matuschek, C. Wallwey, X.-L. Xie, S.-M. Li, Org. Biomol. Chem. 2011, 9, 4328-4335.[5.] C. Wallwey, M. Matuschek, S.-M. Li, Arch. Microbiol. 2010, 192, 127-134.MEP021The interlocking between primary and secondary metabolismin the biosynthesis of the glycopeptide antibiotic BalhimycinV. Goldfinger*, M. Spohn, W. Wohlleben, E. StegmannEberhard-Karls-University, Microbiology/Biotechnology, Tübingen, GermanyBalhimycin is a glycopeptide antibiotic of vancomycin-type. Suchantibiotics are used for the treatment of serious infections caused by multiresistantgram-positive bacteria. To antagonize the consistently increasingnumber of the antibiotic resistance, it is important to understand thebiosynthetic pathway of antibiotic production in details to optimize itsproduction and advance its impact.As glycopeptide balhimycin consists of a glycosylated heptapeptidebackbone. Five of these seven amino acids derive from the shikimatepathway. The analysis of the gene cluster showed that in addition to thegenes encoding the biosynthetic enzymes, the balhimycin gene clusterincludes two genes (dahp, pdh) which encode the homologous keyenzymes of the shikimate pathway. The previous research showed that thedeletion and over expression of these additional genes inA.balhimycinaaffects the antibiotic production. The over expressionofdahpfrom the antibiotic gene cluster causes increased production ofbalhimycin. The deletion of the same gene causes the decreased antibioticproduction. In contrast the over expression ofpdhfrom the balhimycinbiosynthetic gene cluster leads to the lower antibiotic production and itsdeletion does not show any remarkable effects considering the antibioticproduction. This fact could be explained by cross-regulation betweentyrosine and phenyalanine biosynthetic pathway which was describedforA. methanolica. ByA. methanolicatyrosine functions as an activator forprephenate dehydratase (Pdt) which catalyzes the first step reaction on thebranching point from prephenate direction phenylalanine. Otherwise Pdt isfeedback inhibited by phenylalanine. The overexpression of Pdt inA.balhimycinain the current work resulted in the increased antibioticproduction what would explain the results of the previous research andconfirm the similar regulation mechanism byA. balhimycinaandA.methanolicaon the branching point between tyrosine and phenyalaninebiosynthesis.The other disputable question in the tyrosine biosynthesis is the substratespecificity of Pdh. In-silico analysis let to assume that Streptomyce’s Pdhis L-arogenate and not prephenate specific. The overexpression andpurification ofA. balhimycinaPdh which was used in enzyme assayshowed the prephenate specificity. The proof for L-arogenate specificity ofPdh fromA. balhimycinain an enzymassay has to be done.Thykaer J,Nielsen J,Wohlleben W,Weber T,Gutknecht M,Lantz AE,Stegmann E.,MetabEng.,2010,May 25, [Epub ahead of print]Jian Song, Carol A. Bonner, Murray Wolinsky, Roy A. Jensen,BMC Biol.,2005,e pub 3:13Carol A. Bonner, Terrence Disz, Kaitlyn Hwang, Jian Song, Veronika Vonstein, Ross Overbeek,Roy A. Jensen,Microbiol Mol Biol Rev.,2008,p. 13-53David H. Calhoun, Duane L. Pierson, Roy A. Jensen,J Bacteriol.,1973,p.241-251MEP022Identification of a phenazine gene cluster in Dermacoccus sp.MT1.2, isolated from a Mariana Trench sedimentM. Wagner* 1 , W. Abdel-Mageed 2 , M. Jaspars 2 , O. Saleh 3 , L. Heide 3 ,W. Pathom-aree 4 , M. Goodfellow 4 , H.-P. Fiedler 11 Universität Tübingen, IMIT, Tübingen, Germany2 University of Aberdeen, Department of Chemistry, Aberdeen, United Kingdom3 Universität Tübingen, Pharmazeutische Biologie, Tübingen, Germany4 University of Newcastle, School of Biology, Newcastle, United KingdomA sediment sample was taken from the Mariana Trench at the third deepestpoint of earth, the Challenger Deep (10,898 m), in the western PacificOcean (11°19‘911“ N; 142°12‘372“ E) on 21 May 1998 by the remotelyoperated submersible Kaiko, using sterilized mud samplers during divenumber 74. The sediment sample (approximately 2 ml) was stored at -20°C until analyzed for actinomycetes. 38 actinomycetes were isolatedusing marine and raffinose-histidine agar, and were characterized byphylogenetic analysis on 16S rRNA gene sequencing [1]. The strains wereassigned to the genera Dermacoccus (19 isolates), Kocuria (1 isolate),Micromonospora (1 isolate), Streptomyces (5 isolates), Tsukamurella (11isolates) and Williamsia (1 isolate).The Dermacoccus isolates showed unusual secondary metabolite profilesdetermined by HPLC-DAD analysis. Strains MT1.1 and MT1.2 exhibitedthe highest productivity and were therefore selected for fermentationstudies using ISP2 and 410 media, respectively. This led to the productionof seven novel phenazine metabolites, the dermacozines. Structureelucidation was performed by 13 C and 1 H NMR spectroscopic methods,electronic structure calculations and CD spectroscopy. The biologicaleffects of the dermacozines compromised antitumor, antiparasitic andantioxidative activities [2].We show the identification of the phenazine gene cluster in Dermacoccussp. MT1.2. A genome library of strain MT1.2 was screened by colonyPCR. On cosmid MW_A9 a possible gene cluster was found that containedthe essential phenazine core genes and some more genes involved in themodification of the intermediate product phenazine-1,6-dicarboxylic acid.We also show a proposed biosynthesis of dermacozines with respect to thepathway already known from other phenazine producing bacteria.Pathom-aree, W., Stach, J.E.M., Ward, A.C., Horikoshi, K., Bull, A.T. & Goodfellow, M.,Extremophiles, 2006, 10, 181-189.Abdel-Mageed, W.M., Milne, B.F., Wagner, M., Schumacher, M., Sandor, P., Pathom-aree, W.,Goodfellow, M., Bull, A.T., Horikoshi, K., Ebel, R., Diedrich, M., Fiedler, H.-P. and Jaspars, M.,Org. Biomol. Chem., 2010, 8, 2352-2362.MEP023On the way to unravel a novel biosynthetic pathway for theunique volatile 'sodorifen' of Serratia odoriferaT. Weise* 1 , M. Kai 1,2 , S.H. von Reuß 3,4 , W. Francke 4 , B. Piechulla 11 University of Rostock, Institute of Biological Sciences, Rostock, Germany2 Max Planck Institute for Chemical Ecology, Jena, Germany3 Cornell University, Boyce Thompson Institute, Ithaca, United States4 University of Hamburg, Organic Chemistry, Hamburg, GermanyBacteria are a profound source of secondary metabolites, e.g. antibioticsand toxins (1). Unexpectedly large and diverse is also the spectrum ofvolatile secondary compounds. Octamethyl bicycle (3.2.1) octadiene(`sodorifen´) a volatile secondary metabolite of Serratia odorifera 4Rx13was recently found and structurally elucidated (2). `Sodorifen´ (C 16H 26) iscomposed of a new and unusual type of carbon skeleton. Each carbon atomof the bicyclic structure is methylated resp. methylenated. As the structureis new to science also the biosynthesis of this compound is still a mystery.A multi strategy approach including physiological experiments, genome,proteome, and metabolome analysis is presently conducted to unravel thebiosynthesis and regulation of `sodorifen´. Feeding experiments withdifferent carbon sources, e.g. amino acids, organic acids and sugars, wereperformed. The carbon compounds, which resulted in highest `sodorifen´emission, were subsequently used in [ 13 C] isotope feeding experiments andincorporation into `sodorifen´ was analysed by GC/MS and NMR. Besidethe results of the feeding experiments we will present the accompaniedproteome and genome approaches.Acknowledgement: We thank our collaborators G. Gottschalk, R. Daniel, A. Thürmer, J. Voss, R.Lehmann (University of Göttingen, D), M. Glocker and S. Mikkat (University of Rostock, D).1 Wenke K., Kai M., Piechulla B. (2010). Planta 231: 499-5062 Von Reuß S., Kai M., Piechulla B., Francke W. (2009). Angewandte Chemie 122: 2053-2054MEP024New elaiomycins produced by Streptomyces strainsN. Manderscheid* 1 , S. Helaly 2 , A. Kulik 1 , B.-Y. Kim 3 , M. Goodfellow 3 ,J. Wiese 4 , J.F. Imhoff 4 , R.D. Süssmuth 2 , H.-P. Fiedler 11 Universität Tübingen, IMIT, Tübingen, Germany2 TU Berlin, Institut für Chemie, Berlin, Germany3 University of Newcastle, School of Biology, Newcastle, United Kingdom4 Leibniz Institut für Meereswissenschaften, Kieler Wirkstoffzentrum, Kiel,GermanyIn our search for novel secondary metabolites by HPLC-DAD screening,strains Streptomyces sp. BK 190 and Streptomyces sp. Tü 6399 weresubjected to a closer scrutiny because of interesting peaks in their HPLCprofile of a culture filtrate extract. Strain BK 190 was isolated from a haymeadow soil taken from Cockle Park Experimental Farm inNorthumberland, UK. Strain Tü 6399 was isolated from a rhizospheric soilcollected in a spruce stand located in the Rammert Forest near Tübingen,Germany. Both strains were assigned to the genus Streptomyces by theirmorphological and chemotaxonomic features and by the sequence of thealmost complete 16S rRNA gene.It was shown by Kim et al. that strain BK 190 produces two novelalkylhydrazide antibiotics, named elaiomycin B and C, which showedinhibitory activities against Staphylococcus lentus DSM 6672 and towardsthe enzymes acetylcholinesterase and phosphodiesterase [1].Strain Tü 6399 produced two novel azoxy antibiotics, named elaiomycin Dand E, which showed an inhibitory activity against Bacillus subtilis DSM10, Staphylococcus lentus DSM 6672, Xanthomonas campestris DSM1706 and a slight activity towards the enzyme phosphodiesterase 4;elaiomycin E showed a slight activity against acetylcholinesterase.The new compounds are similar in structure to elaiomycin, which was firstdescribed by Stevens et al. [2] containing a unique aliphatic ,unsaturatedazoxy group. Elaiomycin exhibits an unusual inhibitoryactivity against Mycobacterium tuberculosis.1 Kim, B.-Y., Willbold, S., Kulik, A., Helaly, S. E., Zinecker, H., Wiese, J., Imhoff, J. F.,Goodfellow, M., Süssmuth, R. D. & Fiedler, H.-P. Elaiomycins B and C, novel alkylhydrazidesproduced by Streptomyces sp. BK 190. J. Antibiot. 64, 595-597 (2011).2 Haskell, T. H., Ryder, A. & Bartz, Q. R. Elaiomycin, a new tuberculostatic antibiotic; isolationand chemical characterization. Antibiot. Chemother. 4, 141-144 (1954).BIOspektrum | Tagungsband 2012