Prof. Dr. Thomas Eitinger Humboldt-Universität zu Berlin Institut für Biologie/Mikrobiologie Chausseestraße 117 10115 Berlin Germany Phone: +49-30-20938103 e-mail: thomas.eitinger[a]cms.hu-berlin.de

 

 

 

Research Areas

 

A new class of membrane transporters

 

A novel class of solute transporters, frequently of vitamins, transition-metal ions and substrates of salvage pathways was predicted by comparative genomics and validated experimentally in an international collaboration with researchers from the United States, Russia and The Netherlands [see refs. Neubauer et al. (2009), Rodionov et al. (2009), Hebbeln et al. (2007), Rodionov et al. (2006)]. These systems, named ECF transporters, are widespread among archaea and bacteria and certain variants may also occur in plant organelles. They consist of unique substrate-capture (= S) proteins and an energy-coupling module composed of two ABC ATPase domains (= A) and a conserved transmembrane protein (= T). The majority of ECF transporters use shared AT-modules, an unprecedented property among membrane transporters. Since many human pathogens rely on vitamin uptake by ECF transporters, these systems represent a classic Achilles´ heel and a potential target for antibiotic development. Work on ECF transporters is supported by grants EI 374/3-1 and EI 374/4-1 from the Deutsche Forschungsgemeinschaft.

 

 

Modular architecture of energy- coupling factor (ECF) transporters. ECF transporters consist of a substrate-capture protein ("S") and a dedicated (left-hand part) or a shared (right-hand part) energy-coupling module composed of two ABC ATPase domains ("A") and a conserved transmembrane protein ("T"). Shared use of the AT-module is predominantly found in Gram-positive bacteria including many human pathogens. The genome-wide distribution of ECF transporters is listed in the SEED database under "ECF class transporters" [For details see Rodionov et al. (2009)].

 

 

  

 

Secondary transporters for Ni and Co ions: theme and variations

 

A major scientific interest lies in the field of cellular metal ion homeostasis. This research is devoted to structural and functional analyses of prokaryotic and eukaryotic transition-metal transporters that provide nickel or/and cobalt ions for (i) incorporation into various metalloenzymes including important pathogenicity determinants, and (ii) cobalamin biosynthesis. These permeases transport metal ions with very high affinity. Significantly,  members of this type of transporters  are extraordinarily selective and discriminate between closely related transition-metal ions. An interdisciplinary collaboration (applying site-directed mutagenesis, domain swapping, transport studies, overproduction, genomic screening for orthologa, genomic predictions of transcriptional regulatory elements, 2D and 3D structure determination) aims at (i) compiling information on substrate profiles of a multitude of transporters from various families and (ii) a detailed insight into the structural features controlling selectivity. This work has been and is funded by grants EI 374/1-1, EI 374/1-2, EI 374/1-3, EI 374/2-1, EI 374/2-2 and EI 374/2-3 from the Deutsche Forschungsgemeinschaft.

 

 

 

 

 

Topology of nickel/cobalt transporters (NiCoT) and some relatives in prokaryotes (UreH, HupE/UreJ, SodT) and in plants. Conserved segments within the NiCoT family, including the cytoplasmic loop between transmembrane domains (TMDs) IV and V, are highlighted. Dashed lines indicate sequence conservation in cytoplasmic loops. The motif in TMD II with the core sequence HX4DH is considered to be a signature sequence for NiCoTs. Very similar signatures in UreH, HupE/UreJ, SodT and in the plant relatives are shown. Another His-containing motif is conserved in TMD III of the NiCoT sequences, and in TMD II of the UreH, SodT and plant sequences (bold letters indicate strong conservation). The black cylinders in the HupE/UreJ and in the plant transporter models indicate a predicted cleavable leader peptide and the hydrophobic segment of a bipartite thylakoid transit peptide, respectively. Diagonal arrows pointing downwards indicate putative cleavage sites by bacterial signal peptidase I in HupE/UreJ proteins and by lumenal thylakoid processing peptidase in the plant transporters. The diagonal upward-pointing arrow indicates a putative processing site in the stroma. The shading of TMD IV of (i) UreH and the plant transporters and (ii) HupE/UreJ and SodT indicates a signature with one His residue or two His residues, respectively. "..H..H..H..H.." illustrates His motifs with up to 14 His residues in the cytoplasmic loops connecting TMDs III and IV in UreH and SodT proteins. [published in BioMetals 18:399-405]

 

 

Nickel-dependent superoxide dismutase

 

NiSOD is a novel type of superoxide dismutase originally identified in Gram-positive soil bacteria of the genus Streptomyces. Recently, open reading frames with significant similarity to NiSOD precursor proteins have been identified in the genome sequences of marine cyanobacteria (e.g. Crocosphaera watsonii, Prochlorococcus marinus, Synechococcus spec., Trichodesmium erythraeum) that occur in huge amounts in the oceans. In an initial study [see ref. Eitinger (2004)] the NiSOD precursor and a putative maturation peptidase of P. marinus MIT9313 were co-expressed in E. coli and conferred nickel-dependent superoxide dismutase activity on the recombinants. The maturation process of this ecologically very important enzyme is under investigation in our laboratory.

 

 

 

 

 

Genetic localization and putative role of SodT and SodX in maturation of [Ni] superoxide dismutase in many marine cyanobacteria. sodN encodes the NiSOD precursor which undergoes N-terminal proteolysis, catalyzed by SodX, to release the nickel-binding amino group of His-1. SodT presumably acts as a Ni²⁺ transporter in the cytoplasmic membrane. Intracellular nickel trafficking and incorporation into the NiSOD subunits has not yet been analyzed. By analogy to the hexameric Streptomyces enzymes (Barondeau et al 2004; Wuerges et al. 2004), it is likely that marine cyanobacterial NiSOD has an oligomeric structure. [published in BioMetals 18:399-405]

 

 

Relevant Publications

 

Eitinger, T. 2013. Transport of nickel and cobalt in prokaryotes. In Metals and Cells (Encyclopedia of Inorganic and Bioinorganic Chemistry series) (Culotta, V. and Scott, R.A., Eds.) Wiley, in press.

 

Eitinger, T. 2013. Cobalt transporters. Chapter 74. In Encyclopedia of Metalloproteins (Kretsinger, R.H., Uversky, V.N. and Permyakov, E.A., Eds.): SpringerReference (www.springerreference.com). Springer-Verlag Berlin Heidelberg. DOI: 10.1007/SpringerReference_309393 2012-02-23 14:50:27 UTC . [Summary]

 

Eitinger, T. 2013. Nickel transporters. Chapter 85. In Encyclopedia of Metalloproteins (Kretsinger, R.H., Uversky, V.N. and Permyakov, E.A., Eds.): SpringerReference (www.springerreference.com). Springer-Verlag Berlin Heidelberg. DOI: 10.1007/SpringerReference_307412 2012-02-06 10:48:53 UTC. [Summary]

 

Ziomkowska, J., F. Kirsch, A. Herrmann and T. Eitinger. 2012. FRET in lebenden Bakterien. Analyse eines unkonventionellen Vitamin-Transporters. BIOspektrum 18:493-496. [BIOspektrum]

 

Kirsch, F., S. Frielingsdorf, A. Pohlmann, J. Ziomkowska, A. Herrmann and T. Eitinger. 2012. Essential amino acid residues of BioY reveal that dimers are the functional S unit of the Rhodobacter capsulatus biotin transporter. Journal of Bacteriology 194:4505-4512. [J. Bacteriol.]

 

Neubauer, O., C. Reiffler, L. Behrendt and T. Eitinger. 2011. Interactions among the A and T units of an ECF-type biotin transporter analyzed by site-specific crosslinking. PLoS ONE 6:e29087. [PLoS ONE]

 

Eitinger, T., D.A. Rodionov, M. Grote and E. Schneider. 2011. Canonical and ECF-type ATP-binding cassette importers in prokaryotes: diversity in modular organization and cellular functions. FEMS Microbiology Reviews 35:3-67. [FEMS Microbiol. Rev.]

 

Finkenwirth, F., O. Neubauer, J. Gunzenhäuser, J. Schoknecht, S. Scolari, M. Stöckl, T. Korte, A. Herrmann and T. Eitinger. 2010. Subunit composition of an energy-coupling-factor-type biotin transporter analysed in living bacteria. Biochemical Journal 431:373-380. [Biochem. J.]

 

Siche, S., O. Neubauer, P. Hebbeln and T. Eitinger. 2010. A bipartite S unit of an ECF-type cobalt transporter. Research in Microbiology 161:824-829. [Res. Microbiol.]

 

Neubauer, O., A. Alfandega, J. Schoknecht, U. Sternberg, A. Pohlmann and T. Eitinger. 2009. Two essential arginine residues in the T components of energy-coupling factor transporters. Journal of Bacteriology 191:6482-6488. [J. Bacteriol.]

 

Rodionov, D.A., P. Hebbeln, A. Eudes, J. ter Beek, I.A. Rodionova, G.B. Erkens, D.J. Slotboom, M.S. Gelfand, A.L. Osterman, A.D. Hanson and T. Eitinger. 2009. A novel class of modular transporters for vitamins in prokaryotes. Journal of Bacteriology 191:42-51. [J. Bacteriol.]

Selected for Faculty of 1000 Biology: http://www.f1000biology.com/article/id/1126825/evaluation

 

Hebbeln, P., D.A. Rodionov, A. Alfandega and T. Eitinger. 2007. Biotin uptake in prokaryotes by solute transporters with an optional ATP-binding cassette-containing module. Proceedings of the National Academy of Sciences of the U.S.A. 104:2909-2914. [Proc. Natl. Acad. Sci. USA]

 

Pohlmann, A., W.F. Fricke, F. Reineke, B. Kusian, H. Liesegang, R. Cramm, T. Eitinger, C. Ewering, M. Pötter, E. Schwartz, A. Strittmatter, I. Voß, G. Gottschalk, A. Steinbüchel, B. Friedrich and B. Bowien. 2006. Genome sequence of the bioplastic-producing "Knallgas" bacterium Ralstonia eutropha H16. Nature Biotechnology 24:1257-1262. [Nat. Biotechnol.]

 

Rodionov, D.A., P. Hebbeln, M.S. Gelfand and T. Eitinger. 2006. Comparative and functional genomic analysis of prokaryotic nickel and cobalt uptake transporters: evidence for a novel group of ATP-binding cassette transporters. Journal of Bacteriology 188:317-327. [J. Bacteriol.]

 

Eitinger, T., J. Suhr, L. Moore and J.A.C. Smith. 2005. Secondary transporters for nickel and cobalt ions: theme and variations. BioMetals 18:399-405. [PubMed]

 

Eitinger, T. 2004. In vivo production of active nickel superoxide dismutase from Prochlorococcus marinus MIT9313 is dependent on its cognate peptidase. Journal of Bacteriology 186:7821-7825. [J. Bacteriol.]

 

Hebbeln, P. and T. Eitinger. 2004. Heterologous production and characterization of bacterial nickel/cobalt permeases. FEMS Microbiology Letters 230:129-135. [FEMS Microbiol. Lett.]

 

Schwartz, E., A. Henne, R. Cramm, T. Eitinger, B. Friedrich and G. Gottschalk. 2003. Complete nucleotide sequence of pHG1: A Ralstonia eutropha H16 megaplasmid encoding key enzymes of H₂-based lithoautotrophy and anaerobiosis. Journal of Molecular Biology 332:369-383. [PubMed]

 

Degen, O. and T. Eitinger. 2002. Substrate specificity of nickel/cobalt permeases: Insights from mutants altered in transmembrane domains I and II. Journal of Bacteriology 184:3569-3577. [J. Bacteriol.]

 

Eitinger, T. 2001. Microbial Nickel Transport. In Microbial Transport Systems (Winkelmann, G., Ed.), pp. 397-417, Wiley-VCH, Weinheim, Germany. [Summary]

 

Eitinger, T. 2000. Mikrobielle Transporter für Ni²⁺-Ionen. BIOspektrum 6:456.

 

Eitinger, T., O. Degen, U. Böhnke and M. Müller. 2000. Nic1p, a relative of bacterial transition metal permeases in Schizosaccharomyces pombe, provides nickel ion for urease biosynthesis. Journal of Biological Chemistry 275:18029-18033. [J. Biol. Chem.]

 

Eitinger, T. and M.-A. Mandrand-Berthelot. 2000. Nickel transport systems in microorganisms. Archives of Microbiology 173:1-9. [PubMed]

 

Degen, O., M. Kobayashi, S. Shimizu and T. Eitinger. 1999. Selective transport of divalent cations by transition metal permeases: The Alcaligenes eutrophus HoxN and the Rhodococcus rhodochrous NhlF. Archives of Microbiology 171:139-145. [PubMed]

 

Eitinger, T., L. Wolfram, O. Degen and C. Anthon. 1997. A Ni²⁺ binding motif is the basis of high affinity transport of the Alcaligenes eutrophus nickel permease. Journal of Biological Chemistry 272:17139-17144. [J. Biol. Chem.]

 

Eitinger, T. and B. Friedrich. 1997. Microbial nickel transport and incorporation into hydrogenases. In Transition Metals in Microbial Metabolism. (Winkelmann, G. and Carrano, C.J., Eds.), pp. 235-256, Harwood Academic Publishers, Amsterdam, The Netherlands.

 

Wolfram, L., B. Friedrich and T. Eitinger. 1995. The Alcaligenes eutrophus protein HoxN  mediates nickel transport in Escherichia coli. Journal of Bacteriology  177:1840-1843. [J. Bacteriol.]

 

Eitinger, T. and B. Friedrich. 1994. A topological model for the high-affinity nickel transporter of Alcaligenes eutrophus. Molecular Microbiology 12:1025-1032. [PubMed]

 

Eitinger, T. and B. Friedrich. 1991. Cloning, nucleotide sequence, and heterologous expression of a high-affinity nickel transport gene from Alcaligenes eutrophus. Journal of Biological Chemistry 266:3222-3227. [J. Biol. Chem.]

 

Last modified: August 2009