TY - JOUR A1 - Kraegeloh, A. A1 - Amendt, B. A1 - Kunte, Hans-Jörg T1 - Potassium Transport in a Halophilic Member of the Bacteria Domain: Identification and Characterization of the K+ Uptake Systems TrkH and TrkI from Halomonas elongata DSM 2581T N2 - The halophilic bacterium Halomonas elongata accumulates K+, glutamate, and the compatible solute ectoine as osmoprotectants. By functional complementation of Escherichia coli mutants defective in K+ uptake, we cloned three genes that are required for K+ uptake in H. elongata. Two adjacent genes, named trkA (1,374 bp) and trkH (1,449 bp), were identified on an 8.5-kb DNA fragment, while a third gene, called trkI (1,479 bp), located at a different site in the H. elongata chromosome, was found on a second 8.5-kb fragment. The potential protein expressed by trkA is similar to the cytoplasmic NAD+/NADH binding protein TrkA from E. coli, which is required for the activity of the Trk K+ uptake system. The deduced amino acid sequences of trkH and trkI showed significant identity to the transmembrane protein of Trk transporters. K+ transport experiments with trkH and trkI mutants of H. elongata revealed that TrkI exhibits a Km value of 1.12 mM, while the TrkH system has a half-saturation constant of 3.36 mM. Strain KB12, relying on TrkH alone, accumulated K+ with a lower Vmax and required a higher K+ concentration for growth in highly saline medium than the wild type. Strain KB15, expressing only TrkI, showed the same phenotype and the same K+ transport kinetics as the wild type, proving that TrkI is the main K+ transport system in H. elongata. In the absence of both transporters TrkH and TrkI, K+ accumulation was not detectable. K+ transport was also abolished in a trkA deletion mutant, indicating that TrkI and TrkH depend on one type of TrkA protein. Reverse transcriptase PCR experiments and Northern hybridization analyses of the trkAH locus revealed cotranscription of trkAH as well as a monocistronic transcript with only trkA. KW - Potassium transport KW - Osmoregulation KW - Halophilic Bacteria KW - Halomonas elongata KW - TrkH KW - TrkA PY - 2005 U6 - https://doi.org/10.1128/JB.187.3.1036-1043.2005 SN - 0021-9193 SN - 1098-5530 VL - 187 IS - 3 SP - 1036 EP - 1043 PB - American Society for Microbiology CY - Washington, DC AN - OPUS4-6294 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - GEN A1 - Kunte, Hans-Jörg ED - Gunde-Cimerman, N. ED - Oren, A. ED - Plemenitas, A. T1 - K+ Transport and its Role for Osmoregulation in a Halophilic Member of the Bacteria Domain: Characterization of the K+ Uptake Systems from Halomonas elongata KW - Osmoregulation KW - Potassium KW - Glutamate KW - Potassium transport KW - TrkH KW - TrkA KW - Trkl KW - Halomonas elongata KW - Halophilic Bacteria PY - 2005 SN - 1-402-03632-9 N1 - Serientitel: Cellular Origin, Life in Extreme Habitats and Astrobiology – Series title: Cellular Origin, Life in Extreme Habitats and Astrobiology IS - 9 SP - 289 EP - 300 PB - Springer CY - Berlin AN - OPUS4-11668 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kunte, Hans-Jörg A1 - Davies, N. W. A1 - Galinski, E. A. A1 - Witt, K. J. A1 - Grammann, K. A1 - Volke, A. A1 - Sauer, T. T1 - How to shift and exploit the unfavourable equilibrium of ADPC formation T2 - VAAM Jahrestagung CY - Bremen, Germany DA - 2013-03-10 PY - 2013 AN - OPUS4-28375 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Weinhart, M. A1 - Becherer, T. A1 - Schnurbusch, N. A1 - Schwibbert, Karin A1 - Kunte, Hans-Jörg A1 - Haag, R. T1 - Linear and hyperbranched polyglycerol derivatives as excellent bioinert glass coating materials N2 - The non-specific adsorption of proteins to surfaces in contact with biofluids constitutes a major problem in the biomedical and biotechnological field, due to the initiation of biofilm formation and the resulting improper function of devices. Therefore, non-fouling surfaces modified with poly(ethylene glycol) (PEG) are usually applied. In this study, we report the synthesis of triethoxysilane modified glycerol based polymers of linear and branched architecture for the preparation of covalently attached monolayers on glass. Evaluation of the biocompatibility of these surfaces was performed in comparison to bare non-coated glass, hydrophobic hexadecane modified glass, and mPEG modified glass as the controls. Protein adsorption of BSA and fibrinogen (1 mg · mL-1 in PBS) after 4 and 24 h immersion was reduced by more than 96 and 90%, respectively, compared to the adsorption on bare glass substrates. In addition, mouse NIH-3T3 fibroblast cells showed only marginal adhesion on the polyglycerol and mPEG coated slides after 3 and 7 d incubation in cell suspension, which demonstrates the long-term stability of the applied glass coatings. The non-adhesive properties of these coatings were further reflected in bacterial adhesion tests of Escherichia coli K12 and three clinically relevant Gram-positive and negative strains (Staphylococcus aureus, Pseudomonas aeruginosa, and Aeromonas hydrophila), since linear polyglycerol (LPG(OH)), linear poly(methyl glycerol) (LPG(OMe)), and hyperbranched polyglycerol (HPG) reduced the adhesion for all tested strains by more than 99% compared to bare glass. Therefore, polyglycerol derivatives present an excellent non-fouling surface coating as an alternative to PEG with feasibility for surface modification of various substrates. KW - Protein adsorption KW - Non-fouling-surfaces KW - Linear polyglycerol KW - Hyperbranched polyglycerol KW - Bacterial adhesion PY - 2011 U6 - https://doi.org/10.1002/adem.201180012 SN - 1438-1656 VL - 13 IS - 12 SP - B501 EP - B510 PB - Wiley-VCH Verl. CY - Weinheim AN - OPUS4-25221 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Marinelli, F. A1 - Kuhlmann, S.I. A1 - Grell, E. A1 - Kunte, Hans-Jörg A1 - Ziegler, C. M. A1 - Faraldo-Gómez, J.D. T1 - Evidence for an allosteric mechanism of substrate release from membrane-transporter accessory binding proteins N2 - Numerous membrane importers rely on accessory water-soluble proteins to capture their substrates. These substrate-binding proteins (SBP) have a strong affinity for their ligands; yet, substrate release onto the low-affinity membrane transporter must occur for uptake to proceed. It is generally accepted that release is facilitated by the association of SBP and transporter, upon which the SBP adopts a conformation similar to the unliganded state, whose affinity is sufficiently reduced. Despite the appeal of this mechanism, however, direct supporting evidence is lacking. Here, we use experimental and theoretical methods to demonstrate that an allosteric mechanism of enhanced substrate release is indeed plausible. First, we report the atomic-resolution structure of apo TeaA, the SBP of the Na+-coupled ectoine TRAP transporter TeaBC from Halomonas elongata DSM2581T, and compare it with the substrate-bound structure previously reported. Conformational free-energy landscape calculations based upon molecular dynamics simulations are then used to dissect the mechanism that couples ectoine binding to structural change in TeaA. These insights allow us to design a triple mutation that biases TeaA toward apo-like conformations without directly perturbing the binding cleft, thus mimicking the influence of the membrane transporter. Calorimetric measurements demonstrate that the ectoine affinity of the conformationally biased triple mutant is 100-fold weaker than that of the wild type. By contrast, a control mutant predicted to be conformationally unbiased displays wild-type affinity. This work thus demonstrates that substrate release from SBPs onto their membrane transporters can be facilitated by the latter through a mechanism of allosteric modulation of the former. KW - Binding thermodynamics KW - Periplasmic binding protein KW - Secondary transporter KW - ABC transporter KW - Replica-exchange metadynamics PY - 2011 U6 - https://doi.org/10.1073/pnas.1112534108 SN - 0027-8424 SN - 1091-6490 VL - 108 IS - 49 SP - E1285 EP - E1292 PB - National Academy of Sciences CY - Washington, DC AN - OPUS4-25049 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kunte, Hans-Jörg T1 - Osmoregulation in halophilic bacteria: Synthesis and transport of osmoprotectants in halomonas elongata T2 - Max-Planck-Institut für Biochemie CY - Planegg, Germany DA - 2006-04-26 PY - 2006 AN - OPUS4-14156 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kunte, Hans-Jörg T1 - Mechanismen der Anpassung an Wasserstress: Synthese und Tansport von Zellschutzstoffen in halophilen Bacteria T2 - Humboldt Universität CY - Berlin, Germany DA - 2006-12-19 PY - 2006 AN - OPUS4-14157 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Kunte, Hans-Jörg T1 - Osmoregulation in Bacteria: Compatible Solute Accumulation and Osmosensing N2 - Bacteria and Archaea have developed two basic mechanisms to cope with osmotic stress, the salt-in-cytoplasm mechanism, and the organic-osmolyte mechanism. Organic osmolytes or so-called compatible solutes can be accumulated in molar concentration in the cytoplasm and allow for the adaptation of bacterial cells to varying salt concentrations. The biosynthetic pathways of compatible solutes and different compatible solute transport systems are described. A model for osmoregulatory compatible solute accumulation is introduced. KW - Archaea KW - Compatible solutes KW - Halophilic KW - Halotolerant KW - Osmoregulated transporter PY - 2006 U6 - https://doi.org/10.1071/EN06016 SN - 1448-2517 SN - 1449-8979 VL - 3 IS - 2 SP - 94 EP - 99 PB - CSIRO Publishing CY - Collingwood, Vic. AN - OPUS4-14258 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Kuhlmann, S. A1 - Terwisscha van Scheltinga, A. C. A1 - Bienert, Ralf A1 - Kunte, Hans-Jörg A1 - Ziegler, C. M. T1 - 1.55 Å Structure of the Ectoine Binding Protein TeaA of the Osmoregulated TRAP-Transporter TeaABC from Halomonas elongata N2 - TeaABC from the moderate halophilic bacterium Halomonas elongata belongs to the tripartite ATP-independent periplasmic transporters (TRAP-T), a family of secondary transporters functioning in conjunction with periplasmic substrate binding proteins. TeaABC facilitates the uptake of the compatible solutes ectoine and hydroxyectoine that are accumulated in the cytoplasm under hyperosmotic stress to protect the cell from dehydration. TeaABC is the only known TRAP-T activated by osmotic stress. Currently, our knowledge on the osmoregulated compatible solute transporter is limited to ABC transporters or conventional secondary transporters. Therefore, this study presents the first detailed analysis of the molecular mechanisms underlying substrate recognition of the substrate binding protein of an osmoregulated TRAP-T. In the present study we were able to demonstrate by isothermal titration calorimetry measurements that TeaA is a high-affinity ectoine binding protein (Kd = 0.19 μM) that also has a significant but somewhat lower affinity to hydroxyectoine (Kd = 3.8 μM). Furthermore, we present the structure of TeaA in complex with ectoine at a resolution of 1.55 Å and hydroxyectoine at a resolution of 1.80 Å. Analysis of the TeaA binding pocket and comparison of its structure to other compatible solute binding proteins from ABC transporters reveal common principles in compatible solute binding but also significant differences like the solvent-mediated specific binding of ectoine to TeaA. KW - Halophilic Bacteria KW - Halomonas elongata KW - Substrate binding protein KW - TRAP-transporter KW - TeaABC KW - osmoregulation KW - compatible solutes KW - ectoine KW - hydroxyectoine PY - 2008 U6 - https://doi.org/10.1021/bi8006719 SN - 0006-2960 SN - 1520-4995 VL - 47 IS - 36 SP - 9475 EP - 9485 PB - ACS Publ. CY - Washington, DC AN - OPUS4-18560 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schwibbert, Karin A1 - Marin-Sanguino, A. A1 - Bagyan, I. A1 - Heidrich, Gabriele A1 - Lentzen, G. A1 - Seitz, H. A1 - Rampp, M. A1 - Schuster, S.C. A1 - Klenk, H.-P. A1 - Pfeiffer, F. A1 - Oesterheit, D. A1 - Kunte, Hans-Jörg T1 - A blueprint of ectoine metabolism from the genome of the industrial producer Halomonas elongata DSM 2581T N2 - The halophilic γ-proteobacterium Halomonas elongata DSM 2581T thrives at high salinity by synthesizing and accumulating the compatible solute ectoine. Ectoine levels are highly regulated according to external salt levels but the overall picture of its metabolism and control is not well understood. Apart from its critical role in cell adaptation to halophilic environments, ectoine can be used as a stabilizer for enzymes and as a cell protectant in skin and health care applications and is thus produced annually on a scale of tons in an industrial process using H. elongata as producer strain. This paper presents the complete genome sequence of H. elongata (4 061 296 bp) and includes experiments and analysis identifying and characterizing the entire ectoine metabolism, including a newly discovered pathway for ectoine degradation and its cyclic connection to ectoine synthesis. The degradation of ectoine (doe) proceeds via hydrolysis of ectoine (DoeA) to Nα-acetyl-L-2,4-diaminobutyric acid, followed by deacetylation to diaminobutyric acid (DoeB). In H. elongata, diaminobutyric acid can either flow off to aspartate or re-enter the ectoine synthesis pathway, forming a cycle of ectoine synthesis and degradation. Genome comparison revealed that the ectoine degradation pathway exists predominantly in non-halophilic bacteria unable to synthesize ectoine. Based on the resulting genetic and biochemical data, a metabolic flux model of ectoine metabolism was derived that can be used to understand the way H. elongata survives under varying salt stresses and that provides a basis for a model-driven improvement of industrial ectoine production. KW - Genom KW - Ectoin KW - Biotechnologie KW - Halophile Bakterien PY - 2011 U6 - https://doi.org/10.1111/j.1462-2920.2010.02336.x SN - 1462-2912 SN - 1462-2920 VL - 13 IS - 8 SP - 1973 EP - 1994 PB - Blackwell Science CY - Oxford AN - OPUS4-22344 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - GEN A1 - Osterloh, Kurt A1 - Wrobel, Norma A1 - Kunte, Hans-Jörg A1 - Zscherpel, Uwe A1 - Ewert, Uwe T1 - Making the world a safer place - some know-how already exists T2 - MATEST 2009 CY - Cavtat/Dubrovnik, Croatia DA - 2009-09-23 KW - Existing hazards KW - Threat detection KW - Interdisciplinary approach KW - Existing safety rules and procedures KW - Bridging safety and security KW - Common definitions PY - 2009 SN - 978-953-7283-03-2 IS - #10 SP - 1 EP - 7 AN - OPUS4-20710 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Kunte, Hans-Jörg A1 - Lentzen, G. A1 - Galinski, E.A. T1 - Industrial production of the cell protectant ectoine: protection mechanisms, processes, and products N2 - Bacteria, Ärchaea and Eukatya can adapt to saline environments by accumulating compatible solutes in order to maintain an osmotic equilibrium. Compatible solutes are of diverse Chemical structure (sugars, polyols, amino acid derivatives) and are beneficial for bacterial cells not only as osmoregulatory solutes, but also as protectants of proteins by mitigating detrimental effects of ffeezing, drying and high temperatures. The aspartate derivative ectoine is a wide spread compatible solute in Bacteria and possesses additional protective properties compared with other compatible solutes, and stabilizes even vvhole cells against stresses such as UV radiation or cytotoxins. The protective properties of ectoine for proteins can be explained by its strong (kosmotropic) interaction with water and subsequent exclusion ffom Protein surface, the decrease of the solubility of the peptide backbone and the strengthening of intramolecular hydrogen bonds (secondary structures). The stabilizing and UV-protective properties of ectoine attracted industry, which saw the potential to market ectoine as a novel active component in health care products and cosmetics. In joint efforts of industry and research large-scale fermentation procedures have been developed with the halophilic bacterium Halomonas elongata used as a producer strain. The two key technologies that allow for the annual production of ectoine on a scale of tons are the bacterial milking procedure and the development and application of ectoine-excreting mutants (“leaky” mutant). The details of these two procedures including the strain development and fermentation processes will be introduced and current and future applications of ectoine will be discussed. KW - Bacterial milking KW - Batch fermentation KW - Continuous culture KW - Ectoine excretion KW - Hofmeister effect KW - Kosmotrope KW - Leaky mutant KW - Osmophobic effect KW - Protein protection KW - Preferential exclusion PY - 2014 SN - 2211-5501 SN - 2211-551X VL - 3 IS - 1 SP - 1 EP - 16 PB - Bentham Science CY - Sharjah [u.a.] AN - OPUS4-30078 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schneider, Beate A1 - Pfeiffer, F. A1 - Dyall-Smith, M. A1 - Kunte, Hans-Jörg T1 - Genome Sequence of Pseudomonas veronii Strain G2, a Member of a Bacterial Consortium Capable of Polyethylene Degradation N2 - Nine different bacterial isolates were recovered from landfills. Each isolate was obtained in pure culture. As a consortium, the bacteria degrade polyethylene. The complete genome sequence of strain G2 was determined by PacBio sequencing. Using the TYGS server for taxonomic classification, strain G2 was assigned to the species Pseudomonas veronii. KW - Polyethylene KW - Next generation sequencing KW - Pseudomonas veronii PY - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-548973 SP - 1 EP - 2 PB - ASM AN - OPUS4-54897 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schneider, Beate A1 - Pfeiffer, F. A1 - Dyall-Smith, M. A1 - Kunte, Hans-Jörg T1 - Genome Sequence of Micromonospora aurantiaca Strain G9, a Member of a Bacterial Consortium Capable of Polyethylene Degradation N2 - Nine different bacterial isolates were recovered from landfills. Each isolate was obtained in pure culture. As a consortium, the bacteria degrade polyethylene. The complete genome sequence of strain G9 was determined by PacBio sequencing. Using the TYGS server for taxonomic classification, strain G9 was assigned to the species Micromonospora aurantiaca. KW - Polyethylene KW - Micromonospora aurantiaca KW - Degradation KW - iChip PY - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-547181 SN - 2576-098X VL - 11 IS - 5 SP - 1 EP - 2 PB - American Society for Microbiology CY - Washington, DC AN - OPUS4-54718 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Wittmar, J. A1 - Ohle, Corina A1 - Kunte, Hans-Jörg A1 - Brand, I. T1 - Effect of Ectoine on the Conformation and Hybridization of dsDNA in Monolayer Films: A Spectroelectrochemical Study N2 - Lack of long-time stability of dsDNA-based supramolecular assemblies is an important issue that hinders their applications. In this work, 20 base pairs long dsDNA fragments [(dCdG)20 65%] composed of 65% dCdG and 35% dAdT nucleotides were tethered via a thiol to the surface of a gold electrode. The selfassembled (dCdG)20 -65% monolayer was immersed in solutions containing ectoine, a compatible solute. Electrochemical results showed that these monolayers were stable for one month. In situ IR spectroscopy indicated that ectoine interacts weakly with the phosphate-ribose backbone, dehydrating the phosphate groups and stabilizing the A-DNA conformation. This structural reorganization led to a reorientation of nucleic acid base pairs and a local disruption of the double-helix structure. However, the conformation and orientation of the dsDNA fragment was stable in the KW - Ectoine KW - dsDAN monolayer KW - A-DNA conformation PY - 2021 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-533346 SN - 2196-0216 VL - 8 IS - 20 SP - 3844 EP - 3854 PB - Wiley-VCH GmbH AN - OPUS4-53334 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hobmeier, K. A1 - Cantone, M. A1 - Nguyen, Q. A. A1 - Pflüger-Grau, K. A1 - Kremling, A. A1 - Kunte, Hans-Jörg A1 - Pfeiffer, F. A1 - Marin-Sanguino, A. T1 - Adaptation to varying salinity in Halomonas elongata: Much more than ectoine accumulation N2 - The halophilic γ-proteobacterium Halomonas elongata DSM 2581T thrives at salt concentrations well above 10 % NaCl (1.7 M NaCl). A well-known osmoregulatory mechanism is the accumulation of the compatible solute ectoine within the cell in response to osmotic stress. While ectoine accumulation is central to osmoregulation and promotes resistance to high salinity in halophilic bacteria, ectoine has this effect only to a much lesser extent in non-halophiles. We carried out transcriptome analysis of H. elongata grown on two different carbon sources (acetate or glucose), and low (0.17 M NaCl), medium (1 M), and high salinity (2 M) to identify additional mechanisms for adaptation to high saline environments. To avoid a methodological bias, the transcripts were evaluated by applying two methods, DESeq2 and Transcripts Per Million (TPM). The differentially transcribed genes in response to the available carbon sources and salt stress were then compared to the transcriptome profile of Chromohalobacter salexigens, a closely related moderate halophilic bacterium. Transcriptome profiling supports the notion that glucose is degraded via the cytoplasmic Entner-Doudoroff pathway, whereas the Embden-Meyerhoff-Parnas pathway is employed for gluconeogenesis. The machinery of oxidative phosphorylation in H. elongata and C. salexigens differs greatly from that of non-halophilic organisms, and electron flow can occur from quinone to oxygen along four alternative routes. Two of these pathways via cytochrome bo' and cytochrome bd quinol oxidases seem to be upregulated in salt stressed cells. Among the most highly regulated genes in H. elongata and C. salexigens are those encoding chemotaxis and motility proteins, with genes for chemotaxis and flagellar assembly severely downregulated at low salt concentrations. We also compared transcripts at low and high-salt stress (low growth rate) with transcripts at optimal salt concentration and found that the majority of regulated genes were down-regulated in stressed cells, including many genes involved in carbohydrate metabolism, while ribosome synthesis was up-regulated, which is in contrast to what is known from non-halophiles at slow growth. Finally, comparing the acidity of the cytoplasmic proteomes of non-halophiles, extreme halophiles and moderate halophiles suggests adaptation to an increased cytoplasmic ion concentration of H. elongata. Taken together, these results lead us to propose a model for salt tolerance in H. elongata where ion accumulation plays a greater role in salt tolerance than previously assumed. KW - Ectoine KW - Osmoadaptation PY - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-545608 SN - 1664-302X VL - 13 SP - 1 EP - 19 PB - Frontiers Media CY - Lausanne AN - OPUS4-54560 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hobmeier, K. A1 - Oppermann, M. A1 - Stasinski, N. A1 - Kremling, A. A1 - Pflüger-Grau, K. A1 - Kunte, Hans-Jörg A1 - Marin Sanguino, A. T1 - Metabolic engineering of Halomonas elongata: Ectoine secretion is increased by demand and supply driven approaches N2 - The application of naturally-derived biomolecules in everyday products, replacing conventional synthetic manufacturing, is an ever-increasing market. An example of this is the compatible solute ectoine, which is contained in a plethora of treatment formulations for medicinal products and cosmetics. As of today, ectoine is produced in a scale of tons each year by the natural producer Halomonas elongata. In this work, we explore two complementary approaches to obtain genetically improved producer strains for ectoine production. We explore the effect of increased precursor supply (oxaloacetate) on ectoine production, as well as an implementation of increased ectoine demand through the overexpression of a transporter. Both approaches were implemented on an already genetically modified ectoine-excreting strain H. elongata KB2.13 (ΔteaABC ΔdoeA) and both led to new strains with higher ectoine excretion. The supply driven approach led to a 45% increase in ectoine titers in two different strains. This increase was attributed to the removal of phosphoenolpyruvate carboxykinase (PEPCK), which allowed the conversion of 17.9% of the glucose substrate to ectoine. For the demand driven approach, we investigated the potential of the TeaBC transmembrane proteins from the ectoine-specific Tripartite ATP-Independent Periplasmic (TRAP) transporter as export channels to improve ectoine excretion. In the absence of the substrate-binding protein TeaA, an overexpression of both subunits TeaBC facilitated a three-fold increased excretion rate of ectoine. Individually, the large subunit TeaC showed an approximately five times higher extracellular ectoine concentration per dry weight compared to TeaBC shortly after its expression was induced. However, the detrimental effect on growth and ectoine titer at the end of the process hints toward a negative impact of TeaC overexpression on membrane integrity and possibly leads to cell lysis. By using either strategy, the ectoine synthesis and excretion in H. elongata could be boosted drastically. The inherent complementary nature of these approaches point at a coordinated implementation of both as a promising strategy for future projects in Metabolic Engineering. Moreover, a wide variation of intracelllular ectoine levels was observed between the strains, which points at a major disruption of mechanisms responsible for ectoine regulation in strain KB2.13. KW - Osmoadaptation KW - Metabolic engineering PY - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-555644 SN - 1664-302X VL - 13 SP - 1 EP - 13 PB - Frontiers Media CY - Lausanne AN - OPUS4-55564 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schneider, Beate A1 - Pfeiffer, F. A1 - Dyall-Smith, M. A1 - Kunte, Hans-Jörg T1 - Genome Sequence of Cupriavidus campinensis Strain G5, a Member of a Bacterial Consortium Capable of Polyethylene Degradation N2 - Nine different bacterial isolates were recovered from landfills. Each isolate was obtained in pure culture. As a consortium, the bacteria degrade polyethylene. The complete genome sequence of strain G5 was determined by PacBio sequencing. Using the TYGS for taxonomic classification, strain G5 was assigned to the species Cupriavidus campinensis. KW - Polyethylene KW - Cupriavidus campinensis KW - Plastic degradation KW - Genome sequence PY - 2022 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-557897 SP - 1 EP - 2 PB - ASM AN - OPUS4-55789 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Martin-Sanchez, Pedro Maria A1 - Gorbushina, Anna A1 - Kunte, Hans-Jörg A1 - Toepel, Jörg T1 - A novel qPCR protocol for the specific detection and quantification of the fuel-deteriorating fungus Hormoconis resinae N2 - A wide variety of fungi and bacteria are known to contaminate fuels and fuel systems. These microbial contaminants have been linked to fuel system fouling and corrosion. The fungus Hormoconis resinae, a common jet fuel contaminant, is used in this study as a model for developing innovative risk assessment methods. A novel qPCR protocol to detect and quantify H. resinae in, and together with, total fungal contamination of fuel systems is reported. Two primer sets, targeting the markers RPB2 and ITS, were selected for their remarkable specificity and sensitivity. These primers were successfully applied on fungal cultures and diesel samples demonstrating the validity and reliability of the established qPCR protocol. This novel tool allows clarification of the current role of H. resinae in fuel contamination cases, as well as providing a technique to detect fungal outbreaks in fuel systems. This tool can be expanded to other well-known fuel-deteriorating microorganisms. KW - Microbial contamination KW - Real-time quantitative PCR KW - Microbiologically influenced corrosion; KW - Diesel biodeterioration KW - Fouling KW - Indicator PY - 2016 U6 - https://doi.org/10.1080/08927014.2016.1177515 SN - 0892-7014 VL - 32 IS - 6 SP - 635 EP - 644 PB - Taylor & Francis Group CY - Abingdon AN - OPUS4-37337 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Brodehl, Antje A1 - Möller, Anne A1 - Kunte, Hans-Jörg A1 - Koch, Matthias A1 - Maul, Ronald T1 - Biotransformation of the mycotoxin zearalenone by fungi of the genera Rhizopus and Aspergillus N2 - Zearalenone (ZEN) is a nonsteroidal estrogenic mycotoxin biosynthesized by various Fusarium fungi. These fungal species frequently infest grains; therefore, ZEN represents a common contaminant in cereal products. The biotransformation of ZEN differs significantly from species to species, and several metabolites are known to be formed by animals, plants, and microorganisms. The aim of the present study was to investigate the microbial conversion of ZEN by species of the genera Rhizopus and Aspergillus representing relevant fungi for food processing (e.g. fermentation). To monitor the ZEN metabolism, ZEN was added to liquid cultures of the different fungal species. After a period of 3 days, the media were analyzed by HPLC-MS/MS for metabolite formation. Two Aspergillus oryzae strains and all seven Rhizopus species were able to convert ZEN into various metabolites, including ZEN-14-sulfate as well as ZEN-O-14- and ZEN-O-16-glucoside. Microbial transformation of ZEN into the significantly more estrogenic α-zearalenol (α-ZEL) was also observed. Additionally, a novel fungal metabolite, α-ZEL-sulfate, was detected. Semi-quantification of the main metabolites indicates that more than 50% of initial ZEN may be modified. The results show that fungal strains have the potential to convert ZEN into various metabolites leading to a masking of the toxin, for example in fermented food. KW - Microbial conversion KW - Metabolites KW - Fermentation KW - Alpha-zearalenol KW - Conjugation KW - Mycotoxin biotransformation KW - Zearalenone-sulfate PY - 2014 U6 - https://doi.org/10.1111/1574-6968.12586 SN - 0378-1097 SN - 1574-6968 VL - 359 IS - 1 SP - 124 EP - 130 PB - Wiley-Blackwell CY - Malden, Mass., USA AN - OPUS4-31636 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kunte, Hans-Jörg A1 - Schwibbert, Karin A1 - Sanguino, A.M. A1 - Bagyan, I. A1 - Heidrich, Gabriele A1 - Lentzen, G. A1 - Seitz, H. A1 - Rampp, M. A1 - Schuster, S. A1 - Klenk, H.-P. A1 - Pfeiffer, F. A1 - Oesterhelt, D. T1 - A Blueprint of Ectoine Metabolism from the Genome of the Industrial Producer Halomonas elongate DSM 2581T T2 - Jahrestagung der Vereinigung für allgemeine und angewandte Mikrobiologie (VAAM) CY - Hanover, Germany DA - 2010-03-28 PY - 2010 AN - OPUS4-21114 LA - deu AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schweikhard, E.S. A1 - Kuhlmann, S.I. A1 - Kunte, Hans-Jörg A1 - Grammann, K. A1 - Ziegler, C. M. T1 - Structure and function of the universal stress protein TeaD and its role in regulating the ectoine transporter TeaABC of halomonas elongata DSM 2581T N2 - The halophilic bacterium Halomonas elongata takes up the compatible solute ectoine via the osmoregulated TRAP transporter TeaABC. A fourth orf (teaD) is located adjacent to the teaABC locus that encodes a putative universal stress protein (USP). By RT-PCR experiments we proved a cotranscription of teaD along with teaABC. Deletion of teaD resulted in an enhanced uptake for ectoine by the transporter TeaABC and hence a negative activity regulation of TeaABC by TeaD. A transcriptional regulation via DNA binding could be excluded. ATP binding to native TeaD was shown by HPLC, and the crystal structure of TeaD was solved in complex with ATP to a resolution of 1.9 Å by molecular replacement. TeaD forms a dimer–dimer complex with one ATP molecule bound to each monomer, which has a Rossmann-like α/β overall fold. Our results reveal an ATP-dependent oligomerization of TeaD, which might have a functional role in the regulatory mechanism of TeaD. USP-encoding orfs, which are located adjacent to genes encoding for TeaABC homologues, could be identified in several other organisms, and their physiological role in balancing the internal cellular ectoine pool is discussed. PY - 2010 U6 - https://doi.org/10.1021/bi9017522 SN - 0006-2960 SN - 1520-4995 VL - 49 IS - 10 SP - 2194 EP - 2204 PB - ACS Publ. CY - Washington, DC AN - OPUS4-21915 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Kunte, Hans-Jörg A1 - Lentzen, G. A1 - Galinski, E. T1 - Industrial production of the cell protectant ectoine: protection mechanisms, processes, and products N2 - Bacteria, Archaea and Eukarya can adapt to Saline environments by accumulating compatible solutes in order to maintain an osmotic equilibrium. Compatible solutes are of diverse chemical structure (sugars, polyols, amino acid derivatives) and are beneficial for bacterial cells not only as osmoregulatory solutes, but also as protectants of proteins by mitigating detrimental effects of freezing, drying and high temperatures. The aspartate derivative ectoine is a wide spread compatible solute in Bacteria and possesses additional protective properties compared with other compatible solutes, and stabilizes even whole cells against stresses such as UV radiation or cytotoxins. The protective properties of ectoine for proteins can be explained by its strong (kosmotropic) interaction with water and subsequent exclusion from Protein surface, the decrease of the solubility of the peptide backbone and the strengthening of intramolecular hydrogen bonds (secondary structures). The stabilizing and UV-protective properties of ectoine attracted industry, which saw the potential to market ectoine as a novel active component in health care products and cosmetics. In joint efforts of industry and research large-scale fermentation procedures have been developed with the halophilic bacterium Halomonas elongata used as a producer strain. The two key technologies that allow for the annual production of ectoine on a scale of tons are the bacterial milking procedure and the development and application of ectoine-excreting mutants (“leaky” mutant). The details of these two procedures including the strain development and Fermentation processes will be introduced and current and future applications of ectoine will be discussed. KW - Bacterial milking KW - Batch fermentation KW - Continuous culture KW - Ectoine excretion KW - Hofmeister effect KW - Kosmotrope KW - “leaky” mutant KW - Osmophobic effect KW - Protein protection KW - Preferential exclusion PY - 2014 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-376805 SN - 2211-551X VL - 3 IS - 1 SP - 10 EP - 25 PB - Bentham Science AN - OPUS4-37680 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Marinelli, F. A1 - Kuhlmann, S. I. A1 - Grell, E. A1 - Kunte, Hans-Jörg A1 - Ziegler, C. M. A1 - Faraldo-Gómez, J. D. T1 - Evidence for an allosteric mechanism of substrate release from membrane-transporter accessory binding proteins N2 - Numerous membrane importers rely on accessory water-soluble proteins to capture their substrates. These substrate-binding Proteins (SBP) have a strong affinity for their ligands; yet, Substrate release onto the low-affinity membrane transporter must occur for uptake to proceed. It is generally accepted that release is facilitated by the association of SBP and transporter, upon which the SBP adopts a conformation similar to the unliganded state, whose affinity is sufficiently reduced. Despite the appeal of this mechanism, however, direct supporting evidence is lacking. Here, we use experimental and theoretical methods to demonstrate that an allosteric mechanism of enhanced substrate release is indeed plausible. First, we report the atomic-resolution structure of APO TeaA, the SBP of the Na!-coupled ectoine TRAP transporter TeaBC from Halomonas elongata DSM2581T, and compare it with the substrate-bound structure previously reported. Conformational freeenergy landscape calculations based upon molecular Dynamics simulations are then used to dissect the mechanism that couples ectoine binding to structural change in TeaA. These insights allow us to design a triple mutation that biases TeaA toward apo-like conformations without directly perturbing the binding cleft, thus mimicking the influence of the membrane transporter. Calorimetric measurements demonstrate that the ectoine affinity of the conformationally biased triple mutant is 100-fold weaker than that of the wild type. By contrast, a control mutant predicted to be conformationally unbiased displays wild-type affinity. This work thus demonstrates that substrate release from SBPs onto their Membrane transporters can be facilitated by the latter through a mechanism of allosteric modulation of the former. KW - Binding thermodynamics KW - Periplasmic binding protein KW - Secondary transporter KW - ABC transporter KW - Replica-exchange metadynamics PY - 2011 U6 - https://doi.org/10.1073/pnas.1112534108 SN - 0027-8424 SN - 1091-6490 VL - 108 IS - 49 SP - E1285 EP - E1292 AN - OPUS4-37681 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schweikhard, E. S. A1 - Kuhlmann, S. I. A1 - Kunte, Hans-Jörg A1 - Grammann, K. A1 - Ziegler, C. M. T1 - Structure and function of the universal stress protein TeaD and its role in regulating the ectoine transporter TeaABC of halomonas elongata DSM 2581T N2 - The halophilic bacterium Halomonas elongata takes up the compatible solute ectoine via the osmoregulated TRAP transporter TeaABC. A fourth orf (teaD) is located adjacent to the teaABC locus that encodes a putative universal stress protein (USP). By RT-PCR experiments we proved a cotranscription of teaD along with teaABC. Deletion of teaD resulted in an enhanced uptake for ectoine by the Transporter TeaABC and hence a negative activity regulation of TeaABC by TeaD. Atranscriptional regulation viaDNA binding could be excluded. ATP binding to native TeaD was shown by HPLC, and the Crystal structure of TeaD was solved in complex with ATP to a resolution of 1.9 A ° by molecular replacement. TeaD forms a dimer-dimer complex with one ATP molecule bound to each monomer, which has a Rossmann-like R/β overall fold. Our results reveal an ATP-dependent oligomerization of TeaD, which might have a functional role in the regulatory mechanism of TeaD. USP-encoding orfs, which are located adjacent to genes Encoding for TeaABC homologues, could be identified in several other organisms, and their physiological role in balancing the internal cellular ectoine pool is discussed. KW - Universal stress protein KW - Ectoine KW - Transporter TeaABC PY - 2010 U6 - https://doi.org/10.1021/bi9017522 SN - 1520-4995 SN - 0006-2960 VL - 49 IS - 10 SP - 2194 EP - 2204 AN - OPUS4-37682 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Kuhne, Maren A1 - Dippong, Martin A1 - Flemig, Sabine A1 - Hoffmann, Katrin A1 - Petsch, K. A1 - Schenk, J.A. A1 - Kunte, Hans-Jörg A1 - Schneider, Rudolf T1 - Comparative characterization of mAb producing hapten-specific hybridoma cells by flow cytometric analysis and ELISA N2 - A novel method that optimizes the screening for antibody-secreting hapten-specific hybridoma cells by using flow cytometry is described. Cell clones specific for five different haptens were analyzed. We selectively double stained and analyzed fixed hybridoma cells with fluorophore-labeled haptens to demonstrate the target-selectivity, and with a fluorophore-labeled anti-mouse IgG antibody to characterize the level of surface expression of membrane-bound IgGs. ELISA measurements with the supernatants of the individual hybridoma clones revealed that antibodies from those cells, which showed the highest fluorescence intensities in the flow cytometric analysis, also displayed the highest affinities for the target antigens. The fluorescence intensity of antibody-producing cells corresponded well with the produced antibodies' affinities toward their respective antigens. Immunohistochemical staining verified the successful double labeling of the cells. Our method makes it possible to perform a high-throughput screening for hybridoma cells, which have both an adequate IgG production rate and a high target affinity. KW - Immunization KW - Hapten KW - Monoclonal antibodies KW - Hybridoma KW - Flow cytometry KW - ELISA KW - Estradiol KW - Estrone KW - Digoxigenin KW - Zearalenone KW - Aflatoxin KW - CLSM PY - 2014 U6 - https://doi.org/10.1016/j.jim.2014.07.004 SN - 0022-1759 SN - 1872-7905 VL - 413 SP - 45 EP - 56 PB - Elsevier B.V. CY - Amsterdam AN - OPUS4-32322 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hahn, Marc Benjamin A1 - Meyer, Susann A1 - Schröter, Maria-Astrid A1 - Seitz, H. A1 - Kunte, Hans-Jörg A1 - Solomun, Tihomir A1 - Sturm, Heinz T1 - Direct electron irradiation of DNA in fully aqueous environment. Damage determination in combination with Monte Carlo simulations N2 - We report on a study in which plasmid DNA in water was irradiated with 30 keV electrons generated by a scanning electron microscope and passed through a 100 nm thick Si3N4 membrane. The corresponding Monte Carlo simulations suggest that the kinetic energy spectrum of the electrons throughout the water is dominated by low energy electrons (<100 eV). The DNA radiation damage, single-strand breaks (SSB) and double-strand breaks (DSB), was determined by electrophoresis. The median lethal dose of D1/2 = 1.7 ± 0.3 Gy was found to be much smaller compared to partially or fully hydrated DNA irradiated under vacuum conditions. The ratio of DSB to SSB was found to be (1:12) as compared to 1:88) found for hydrated DNA. Our method enables quantitative measurements of radiation damage to biomolecules (DNA, proteins) in solutions under varying conditions (pH, salinity, cosolutes) for an electron energy range which is difficult to probe by standard methods. KW - Plasmid DNA in water KW - Monte Carlo simulation KW - Low energy electrons KW - DNA radiation damage KW - Single-strand break (SSB) KW - Double-strand break (DSB) KW - Lethal dose KW - Radiation damage to biomolecules KW - Solutions (pH, salinity, cosolutes) PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-386981 SN - 1463-9076 SN - 1463-9084 VL - 19 IS - 3 SP - 1798 EP - 1805 PB - Royal Society of Chemistry AN - OPUS4-38698 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Moghaddam, J. A. A1 - Boehringer, N. A1 - Burdziak, A. A1 - Kunte, Hans-Jörg A1 - Galinski, E. A. A1 - Schäberle, T. F. T1 - Different strategies of osmoadaptation in the closely related marine myxobacteria Enhygromyxa salina SWB007 and Plesiocystis pacifica SIR-1 N2 - Only a few myxobacteria are known to date that are classified as marine, owing to their salt dependency. In this study, the salt tolerance mechanism of these bacteria was investigated. To this end, a growth medium was designed in which the mutated Escherichia coli strain BKA13 served as sole food source for the predatory, heterotrophic myxobacteria. This enabled measurement of the osmolytes without any background and revealed that the closely related strains Enhygromyxa salina SWB007 and Plesiocystis pacifica SIR-1 developed different strategies to handle salt stress. Ple. pacifica SIR-1, which was grown between 1 and 4% NaCl, relies solely on the accumulation of amino acids, while Enh. salina SWB007, which was grown between 0.5 and 3% NaCl, employs, besides betaine, hydroxyectoine as the major compatible solute. In accordance with this analysis, only in the latter strain was a locus identified that codes for genes corresponding to the biosynthesis of betaine, ectoine and hydroxyectoine. KW - Osmoregulation KW - Myxobacteria KW - Betaine KW - Ectoine PY - 2016 U6 - https://doi.org/10.1099/mic.0.000250 VL - 162 IS - 4 SP - 651 EP - 661 AN - OPUS4-39115 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Kindzierski, V. A1 - Raschke, Silvia A1 - Knabe, Nicole A1 - Siedler, F. A1 - Scheffer, B. A1 - Pflüger-Grau, K. A1 - Pfeiffer, F. A1 - Oesterhelt, D. A1 - Marin-Sanguino, A. A1 - Kunte, Hans-Jörg T1 - Osmoregulation in the halophilic bacterium halomonas elongata: A case study for integrative systems biology N2 - Halophilic bacteria use a variety of osmoregulatory methods, such as the accumulation of one or more compatible solutes. The wide diversity of compounds that can act as compatible solute complicates the task of understanding the different strategies that halophilic bacteria use to cope with salt. This is specially challenging when attempting to go beyond the pathway that produces a certain compatible solute towards an understanding of how the metabolic network as a whole addresses the problem. Metabolic reconstruction based on genomic data together with Flux Balance Analysis (FBA) is a promising tool to gain insight into this problem. However, as more of these reconstructions become available, it becomes clear that processes predicted by genome annotation may not reflect the processes that are active in vivo. As a case in point, E. coli is unable to grow aerobically on citrate in spite of having all the necessary genes to do it. It has also been shown that the realization of this genetic potential into an actual capability to metabolize citrate is an extremely unlikely event under normal evolutionary conditions. Moreover, many marine bacteria seem to have the same pathways to metabolize glucose but each species uses a different one. In this work, a metabolic network inferred from genomic annotation of the halophilic bacterium Halomonas elongata and proteomic profiling experiments are used as a starting point to motivate targeted experiments in order to find out some of the defining features of the osmoregulatory strategies of this bacterium. This new information is then used to refine the network in order to describe the actual capabilities of H. elongata, rather than its genetic potential. KW - Halomonas elongata KW - Systems biology KW - Flux balance analysis KW - Proteomic analysis PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-391172 SN - 1932-6203 VL - 12 IS - 1 SP - Article e0168818, 1 EP - 22 AN - OPUS4-39117 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Borzekowski, Antje A1 - Drewitz, T. A1 - Keller, Julia A1 - Pfeifer, Dietmar A1 - Kunte, Hans-Jörg A1 - Koch, Matthias A1 - Rohn, S. A1 - Maul, R. T1 - Biosynthesis and characterization of zearalenone-14-sulfate, zearalenone-14-glucoside and zearalenone-16-glucoside using common fungal strains N2 - Zearalenone (ZEN) and its phase II sulfate and glucoside metabolites have been detected in food and feed commodities. After consumption, the conjugates can be hydrolyzed by the human intestinal microbiota leading to liberation of ZEN that implies an underestimation of the true ZEN exposure. To include ZEN conjugates in routine analysis, reliable standards are needed, which are currently not available. Thus, the aim of the present study was to develop a facilitated biosynthesis of ZEN-14-sulfate, ZEN-14-glucoside and ZEN-16-glucoside. A metabolite screening was conducted by adding ZEN to liquid fungi cultures of known ZEN conjugating Aspergillus and Rhizopus strains. Cultivation conditions and ZEN incubation time were varied. All media samples were analyzed for metabolite formation by HPLC-MS/MS. In addition, a consecutive biosynthesis was developed by using Fusarium graminearum for ZEN biosynthesis with subsequent conjugation of the toxin by utilizing Aspergillus and Rhizopus species. ZEN-14-sulfate (yield: 49%) is exclusively formed by Aspergillus oryzae. ZEN-14-glucoside (yield: 67%) and ZEN-16-glucoside (yield: 39%) are formed by Rhizopus oryzae and Rhizopus oligosporus, respectively. Purities of ≥73% ZEN-14-sulfate, ≥82% ZEN-14-glucoside and ≥50% ZEN-16-glucoside were obtained by 1H-NMR. In total, under optimized cultivation conditions, fungi can be easily utilized for a targeted and regioselective synthesis of ZEN conjugates. KW - Mycotoxin KW - Zearalenone KW - Conjugate KW - Biosynthesis KW - Fusarium KW - Aspergillus KW - Rhizopus PY - 2018 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-444246 SN - 2072-6651 VL - 10 IS - 3 SP - Article 104, 1 EP - 15 PB - MDPI CY - Basel AN - OPUS4-44424 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - He, Shulin A1 - Johnston, P. R. A1 - Kuropka, B. A1 - Lokatis, S. A1 - Weise, C. A1 - Plarre, Rüdiger A1 - Kunte, Hans-Jörg A1 - McMahon, Dino Peter T1 - Termite soldiers contribute to social immunity by synthesizing potent oral secretions N2 - The importance of soldiers to termite Society defence has long been recognized, but the contribution of soldiers to other societal functions, such as colony immunity, is less well understood. We explore this issue by examining the role of soldiers in protecting nestmates against pathogen infection. Even though they are unable to engage in grooming behaviour, we find that the presence of soldiers of the Darwin termite, Mastotermes darwiniensis, significantly improves the survival of nestmates following entomopathogenic infection. We also show that the copious exocrine oral secretions produced by Darwin termite soldiers contain a high concentration of Proteins involved in digestion, chemical biosynthesis, and immunity. The oral secretions produced by soldiers are sufficient to protect nestmates against infection, and they have potent inhibitory activity against a broad spectrum of microbes. Our findings support the view that soldiers may play an important role in colony immunity, and broaden our understanding of the possible function of soldiers during the origin of soldier-first societies. KW - External KW - Social KW - Immunity KW - Soldier KW - Antimicrobial KW - Proteome PY - 2018 U6 - https://doi.org/10.1111/imb.12499 SN - 1365-2583 SN - 0962-1075 VL - 27 IS - 5 SP - 564 EP - 576 PB - Wiley-Blackwell CY - Oxford AN - OPUS4-45726 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hobmeier, K. A1 - Goëss, M. C. A1 - Sehr, C. A1 - Schwaminger, S. A1 - Berensmeier, S. A1 - Kremling, A. A1 - Kunte, Hans-Jörg A1 - Pflüger-Grau, K. A1 - Marin-Sanguino, A. T1 - Anaplerotic Pathways in Halomonas elongata: The Role of the Sodium Gradient N2 - Salt tolerance in the γ-proteobacterium Halomonas elongata is linked to its ability to produce the compatible solute ectoine. The metabolism of ectoine production is of great interest since it can shed light on the biochemical basis of halotolerance as well as pave the way for the improvement of the biotechnological production of such compatible solute. Ectoine belongs to the biosynthetic family of aspartate-derived amino-acids. Aspartate is formed from oxaloacetate, thereby connecting ectoine production to the anaplerotic reactions that refill carbon into the tricarboxylic acid cycle (TCA cycle). This places a high demand on these reactions and creates the need to regulate them not only in response to growth but also in response to extracellular salt concentration. In this work, we combine modeling and experiments to analyze how these different needs shape the anaplerotic reactions in H. elongata. First, the stoichiometric and thermodynamic factors that condition the flux distributions are analyzed, then the optimal patterns of operation for oxaloacetate production are calculated. Finally, the phenotype of two deletion mutants lacking potentially relevant anaplerotic enzymes: phosphoenolpyruvate carboxylase (Ppc) and oxaloacetate decarboxylase (Oad) are experimentally characterized. The results show that the anaplerotic reactions in H. elongata are indeed subject to evolutionary pressures that differ from those faced by other gram-negative bacteria. Ectoine producing halophiles must meet a higher metabolic demand for oxaloacetate and the reliance of many marine bacteria on the Entner-Doudoroff pathway compromises the anaplerotic efficiency of Ppc, which is usually one of the main enzymes fulfilling this role. The anaplerotic flux in H. elongata is contributed not only by Ppc but also by Oad, an enzyme that has not yet been shown to play this role in vivo. Ppc is necessary for H. elongata to grow normally at low salt concentrations but it is not required to achieve near maximal growth rates as long as there is a steep sodium gradient. On the other hand, the lack of Oad presents serious difficulties to grow at high salt concentrations. This points to a shared role of these two enzymes in guaranteeing the supply of oxaloacetate for biosynthetic reactions. KW - Metabolic flux analysis KW - Halophilic bacteria KW - Halomonas elongata KW - Metabolic modeling PY - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-513124 VL - 11 SP - 561800 AN - OPUS4-51312 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Borzekowski, Antje A1 - Anggriawan, R. A1 - Auliyati, M. A1 - Kunte, Hans-Jörg A1 - Koch, Matthias A1 - Rohn, S. A1 - Karlovsky, P. A1 - Maul, Ronald T1 - Formation of Zearalenone Metabolites in Tempeh Fermentation N2 - Tempeh is a common food in Indonesia, produced by fungal fermentation of soybeans using Rhizopus sp., as well as Aspergillus oryzae, for inoculation. Analogously, for economic reasons, mixtures of maize and soybeans are used for the production of so-called tempeh-like products. For maize, a contamination with the mycoestrogen zearalenone (ZEN) has been frequently reported. ZEN is a mycotoxin which is known to be metabolized by Rhizopus and Aspergillus species. Consequently, this study focused on the ZEN transformation during tempeh fermentation. Five fungal strains of the genera Rhizopus and Aspergillus, isolated from fresh Indonesian tempeh and authentic Indonesian inocula, were utilized for tempeh manufacturing from a maize/soybean mixture (30:70) at laboratory-scale. Furthermore, comparable tempeh-like products obtained from Indonesian markets were analyzed. Results from the HPLC-MS/MS analyses show that ZEN is intensely transformed into its metabolites alpha-zearalenol (alpha-ZEL), ZEN-14-sulfate, alpha-ZEL-sulfate, ZEN-14-glucoside, and ZEN-16-glucoside in tempeh production. alpha-ZEL, being significantly more toxic than ZEN, was the main metabolite in most of the Rhizopus incubations, while in Aspergillus oryzae fermentations ZEN-14-sulfate was predominantly formed. Additionally, two of the 14 authentic samples were contaminated with ZEN, alpha-ZEL and ZEN-14-sulfate, and in two further samples, ZEN and alpha-ZEL, were determined. Consequently, tempeh fermentation of ZEN-contaminated maize/soybean mixture may lead to toxification of the food item by formation of the reductive ZEN metabolite, alpha-ZEL, under model as well as authentic conditions. KW - Modified mycotoxins KW - Zearalenone sulfate KW - a-zearalenol KW - Food fermentation KW - Rhizopus and Aspergillus oryzae PY - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-491126 VL - 24 IS - 15 SP - 2697 PB - MDPI AN - OPUS4-49112 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Wittmar, J. A1 - Meyer, Susann A1 - Sieling, T. A1 - Kunte, Hans-Jörg A1 - Smiatek, Jens A1 - Brand, Izabella T1 - What Does Ectoine Do to DNA? A Molecular-Scale Picture of Compatible Solute−Biopolymer Interactions N2 - Compatible solutes are accumulated in the cytoplasm of halophilic microorganisms. These molecules enable their survival in a high salinity environment. Ectoine is such a compatible solute. It is a zwitterionic molecule which strongly interacts with surrounding water molecules and changes the dynamics of the local hydration shell. Ectoine interacts with biomolecules such as lipids, proteins and DNA. The molecular interaction between ectoine and biomolecules in particular the interaction between ectoine and DNA is far from being understood. In this paper we describe molecular aspects of the interaction between ectoine and double stranded DNA(dsDNA). Two 20 base pairs long dsDNA fragments were immobilized on a Gold surface via a thiol-tether. The interaction between the dsDNA monolayers with diluted and concentrated ectoine solutions was examined by means of X-ray photoelectron and polarization modulation infrared reflection absorption spectroscopies (PM IRRAS). Experimental results indicate that the ability of ectoine to bind water reduces the strength of hydrogen bonds formed to the ribose-phosphate backbone in the dsDNA. In diluted (0.1 M) ectoine solution, DNA interacts predominantly with water molecules. The sugar-phosphate backbone is involved in the formation of strong hydrogen bonds to water, which with elapsing time leads to a reorientation of the planes of nucleic acid bases. This reorientation destabilizes the hydrogen bonds strength between the bases and leads to a partial dehybridizaiton of the dsDNA. In concentrated ectoine solution (2.5 M), almost all water molecules interact with ectoine. Under this condition ectoine is able to interact directly with DNA. Density functional theory (DFT) calculations demonstrate that the direct interaction involves the nitrogen atoms in ectoine and phosphate groups in the DNA molecule. The results of the quantum chemical calculations Show that rearrangements in the ribose-phosphate backbone, caused by a direct interaction with ectoine, facilitates contacts between O atom in the phosphate group and H atoms in a nucleic acid base. In the PM IRRA spectra, an increase in the number of the IR absorption modes in the base pair frequency region proves that the hydrogen bonds between bases become weaker. Thus, a sequence of reorientations caused by interaction with ectoine leads to a breakdown of hydrogen bonds between bases in the double helix. KW - Compatible solute KW - Ectoine KW - DNA KW - Self-assembled monolayer KW - IR spectroscopy KW - XPS PY - 2020 U6 - https://doi.org/10.1021/acs.jpcb.0c05273 VL - 124 IS - 37 SP - 7999 EP - 8011 PB - ACS Publicatios AN - OPUS4-51182 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Moghaddam, J. A. A1 - Böhringer, N. A1 - Burdziak, A. A1 - Kunte, Hans-Jörg A1 - Galinski, E. A. A1 - Schäberle, T. F. T1 - Different strategies of osmoadaptation in the closely related marine myxobacteria Enhygromyxa salina SWB007 and Plesiocystis pacifica SIR-1 N2 - Only a few myxobacteria are known to date that are classified as marine, owing to their salt dependency. In this study, the salt tolerance mechanism of these bacteria was investigated. To this end, a growth medium was designed in which the mutated Escherichia coli strain BKA13 served as sole food source for the predatory, heterotrophic myxobacteria. This enabled measurement of the osmolytes without any background and revealed that the closely related strains Enhygromyxa salina SWB007 and Plesiocystis pacifica SIR-1 developed different strategies to handle salt stress. Ple. pacifica SIR-1, which was grown between 1 and 4% NaCl, relies solely on the accumulation of amino acids, while Enh. salina SWB007, which was grown between 0.5 and 3% NaCl, employs, besides betaine, hydroxyectoine as the major compatible solute. In accordance with this analysis, only in the latter strain was a locus identified that codes for genes corresponding to the biosynthesis of betaine, ectoine and hydroxyectoine. KW - Osmoadaptation PY - 2016 U6 - https://doi.org/10.1099/mic.0.000250 VL - 162 SP - 651 EP - 661 PB - SOC General Microbiology CY - Berks, UK AN - OPUS4-35782 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hahn, Marc Benjamin A1 - Solomun, Tihomir A1 - Wellhausen, Robert A1 - Herrmann, S A1 - Seitz, H A1 - Meyer, Susann A1 - Kunte, Hans-Jörg A1 - Zeman, J. A1 - Uhlig, F A1 - Smiatek, J A1 - Sturm, Heinz T1 - Influence of the Compatible Solute Ectoine on the Local Water Structure: Implications for the Binding of the Protein G5P to DNA N2 - Microorganisms accumulate molar concentrations of compatible solutes like ectoine to prevent proteins from denaturation. Direct structural or spectroscopic information on the mechanism and about the hydration shell around ectoine are scarce. We combined surface plasmon resonance (SPR), confocal Raman spectroscopy, molecular dynamics simulations, and density functional theory (DFT) calculations to study the local hydration shell around ectoine and its influence on the binding of a gene-S-protein (G5P) to a single-stranded DNA (dT(25)). Due to the very high hygroscopicity of ectoine, it was possible to analyze the highly stable hydration shell by confocal Raman spectroscopy. Corresponding molecular dynamics simulation results revealed a significant change of the water dielectric constant in the presence of a high molar ectoine concentration as compared to pure water. The SPR data showed that the amount of protein bound to DNA decreases in the presence of ectoine, and hence, the protein-DNA dissociation constant increases in a concentration-dependent manner. Concomitantly, the Raman spectra in terms of the amide I region revealed large changes in the protein secondary structure. Our results indicate that ectoine strongly affects the molecular recognition between the protein and the oligonudeotide, which has important consequences for osmotic regulation mechanisms. KW - Aqueous solution KW - Biological structure KW - Raman spectroscopy KW - Organic osmolytes KW - High throughput KW - Gene-5 protein KW - Amino acid KW - Water structure PY - 2015 U6 - https://doi.org/10.1021/acs.jpcb.5b09506 SN - 1520-6106 SN - 1089-5647 SN - 1520-5207 VL - 119 IS - 49 SP - 15212 EP - 15220 AN - OPUS4-35800 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Zhang, Q. A1 - Zheng, M. A1 - Huang, Y. A1 - Kunte, Hans-Jörg A1 - Wang, X. A1 - Liu, Y. A1 - Zheng, C. T1 - Long term corrosion estimation of carbon steel, titanium and its alloy in backfill material of compacted bentonite for nuclear waste repository N2 - The container of high-level radioactive waste (HLRW) being in deep geological disposal, the backfill material is needed to serve as the second defense for HLRW and the highly compacted bentonite is generally selected. As the time goes, the underground water will infiltrate the backfill, causing the corrosion of materials for the building of containers in the formed electrolyte. Carbon steel, titanium and its alloy are the potential candidate materials for the fabrication of HLRW containers. The current investigation aims at assessing the safety of HLRW container in deep geological disposal for hundreds of thousands of years and facilitating the material selection for future Container fabrication by estimating their corrosion behavior in compacted bentonite with a series of moisture content at different temperatures through electrochemical methods including open circuit potential (OCP), electrochemical impedance spectroscopy (EIS) and potentiodynamic polarization curve (PC) measurements. The corrosion rates were estimated for a carbon steel, a pure titanium and a titanium alloy in compacted Gaomiaozi Bentonite infiltrated with simulated underground water in Beishan area of China over an expected disposal period up to 106 years respectively, showing that titanium and its alloy are more reliable materials for building HLRW containers than carbon steel. KW - Issues KW - Disposal KW - Performance KW - Moisture KW - Lifetime KW - Water KW - Model PY - 2019 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-492413 SN - 2045-2322 VL - 9 SP - 3195 PB - Nature Publishing Group AN - OPUS4-49241 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Martynenko, Irina V. A1 - Kusic, Dragana A1 - Weigert, Florian A1 - Stafford, S. A1 - Donnelly, F. C. A1 - Evstigneev, R. A1 - Gromova, Y. A1 - Baranov, A. V. A1 - Rühl, Bastian A1 - Kunte, Hans-Jörg A1 - Gun'ko, Y. K. A1 - Resch-Genger, Ute T1 - Magneto-fluorescent microbeads for bacteria detection constructed from superparamagnetic Fe3O4 nanoparticles and AIS/ZnS quantum dots N2 - The efficient and sensitive detection of pathogenic microorganisms in aqueous environments, such as water used in medical applications, drinking water, and cooling water of industrial plants, requires simple and fast methods suitable for multiplexed detection such as flow cytometry (FCM) with optically encoded carrier beads. For this purpose, we combine fluorescent Cd-free Ag−In−S ternary quantum dots (t-QDs) with fluorescence Lifetimes (LTs) of several hundred nanoseconds and superparamagnetic Fe3O4 nanoparticles (SPIONs) with mesoporous CaCO3 microbeads to a magneto-fluorescent bead platform that can be surface-functionalized with bioligands, such as antibodies. This inorganic bead platform enables immuno-magnetic separation, target enrichment, and target quantification with optical readout. The beads can be detected with steady-state and time-resolved fluorescence microscopy and flow cytometry (FCM). Moreover, they are suited for readout by time gated emission. In the following, the preparation of these magneto-fluorescent CaCO3 beads, their spectroscopic and analytic characterization, and their conjugation with bacteria-specific antibodies are presented as well as proof-of-concept measurements with Legionella pneumophila including cell cultivation and plating experiments for bacteria quantification. Additionally, the possibility to discriminate between the long-lived emission of the LT-encoded capture and carrier CaCO3 beads and the short-lived Emission of the dye-stained bacteria with time-resolved fluorescence techniques and single wavelength excitation is demonstrated. KW - Fluorescence KW - method KW - lifetime KW - quantum yield KW - particle KW - magnetic nanoparticle KW - immunoseparation KW - flow cytometry KW - fluorescence microscopy KW - nanoparticle KW - quantum dot KW - AIS QD KW - fluorescence KW - bacteria detection KW - bacteria KW - antibody KW - Legionella KW - screening tes KW - FLIM PY - 2019 U6 - https://doi.org/10.1021/acs.analchem.9b01812 SN - 0003-2700 SN - 1520-6882 VL - 91 SP - 12661 EP - 12669 PB - American Chemical Society CY - Washington, DC AN - OPUS4-50117 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Vandrich, Jasmina A1 - Pfeiffer, F. A1 - Alfaro Espinoza, Gabriela A1 - Kunte, Hans-Jörg T1 - Contribution of mechanosensitive channels to osmoadaptation and ectoine excretion in Halomonas elongata N2 - For osmoadaptation the halophilic bacterium Halomonas elongata synthesizes as its main compatible solute the aspartate derivative ectoine. H. elongata does not rely entirely on synthesis but can accumulate ectoine by uptake from the surrounding environment with the help of the osmoregulated transporter TeaABC. Disruption of the TeaABC-mediated ectoine uptake creates a strain that is constantly losing ectoine to the medium. However, the efflux mechanism of ectoine in H. elongata is not yet understood. H. elongata possesses four genes encoding mechanosensitive channels all of which belong to the small conductance type (MscS). Analysis by qRT-PCR revealed a reduction in transcription of the mscS genes with increasing salinity. The response of H. elongata to hypo- and hyperosmotic shock never resulted in up-regulation but rather in downregulation of mscS transcription. Deletion of all four mscS genes created a mutant that was unable to cope with hypoosmotic shock. However, the knockout mutant grew significantly faster than the wildtype at high salinity of 2 M NaCl, and most importantly, still exported 80% of the ectoine compared to the wildtype. We thus conclude that a yet unknown system, which is independent of mechanosensitive channels, is the major export route for ectoine in H. elongata. KW - Halomonas elongata KW - Ectoine KW - Osmotic shock KW - Solute excretion KW - Osmoadaptation KW - Mechanosensitive channel KW - MscS PY - 2020 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-507229 VL - 24 SP - 421 EP - 432 PB - Springer AN - OPUS4-50722 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kunte, Hans-Jörg T1 - Effect of Ectoine on DNA: Mechanisms of Interaction and Protection N2 - The compatible solute ectoine is a versatile protectant synthesized by many prokaryotes. It is used as an osmolyte helping microorganisms to maintain an osmotic equilibrium. In addition, ectoine acts as a stabilizer and protects proteins, membranes and whole cells against detrimental effects such as freezing and thawing, drying and high temperatures. Its protective effect is explained by the preferential exclusion model, which postulates that ectoine does not directly interact with biomolecules but is excluded from their surface. Interestingly, details on the interaction of ectoine with DNA are still unknown. Therefore, we studied the influence of ectoine on DNA and the mechanisms by which ectoine protects DNA against ionizing radiation. To emulate biological conditions, we used a sample holder comprising a silicon chip with a Si3N4 membrane, which allows for electron irradiation of DNA in aqueous solution. Analysis by atomic force microscopy revealed that without ectoine, DNA was damaged by irradiation with a dose of 1,7 +/-0,3 Gy. With ectoine, DNA remained undamaged, even after irradiation with 15 Gy. Simulations with dsDNA and ectoine in water revealed a preferential binding of the zwitterionic ectoine to the negatively charged DNA. According to the simulations, binding of ectoine will destabilize dsDNA. Destabilizing is probably caused by the transition of B-DNA to A-DNA and will reduce the DNA melting temperature, which was experimentally proven. The preferential binding provides a stable ectoine shell around DNA, which allows ectoine to reduce OH-radicals and electrons near the DNA and thereby mitigating the damaging effect of ionizing radiation. T2 - Halophiles 2019 CY - Cluj-Napoca, Romania DA - 24.06.2019 KW - Ectoine KW - Ionizing radiation KW - Preferential binding PY - 2019 AN - OPUS4-48618 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CHAP A1 - Kunte, Hans-Jörg A1 - Schwarz, T. A1 - Galinski, E. A. ED - Lee, N. M. T1 - The compatible solute ectoine: protection mechanisms, strain development, and industrial production N2 - Bacteria, Archaea, and Eukarya can adapt to saline environments by accumulating compatible solutes in order to maintain an osmotic equilibrium. Compatible solutes are of diverse chemical structure (sugars, polyols, amino acid derivatives) and are beneficial for bacterial cells not only as osmoregulatory solutes but also as protectants of proteins by mitigating detrimental effects of freezing, drying, and high temperatures. The aspartate derivative ectoine is a widespread compatible solute in Bacteria and possesses additional protective properties compared with other compatible solutes and stabilizes even whole cells against stresses such as ultraviolet radiation or cytotoxins. Here, it is our intention to go beyond a simple description of effects, but to depict the molecular interaction of ectoine with biomolecules, such as proteins, membranes, and DNA and explain the underlying principles. The stabilizing properties of ectoine attracted industry, which saw the potential to market ectoine as a novel active component in health care products and cosmetics. In joint efforts of industry and research, a large-scale fermentation procedure has been developed with the halophilic bacterium Halomonas elongata used as a producer strain. The development and application of ectoine-excreting mutants from H. elongata (“leaky” mutants) allow for the annual production of ectoine on a scale of tons. The details of the strain development and fermentation processes will be introduced. KW - Ectoine KW - Biotechnology KW - Compatible solute KW - Preferential exclusion KW - Osmophobic effect PY - 2020 SN - 978-3-11-042773-8 SP - 121 EP - 136 PB - De Gruyter AN - OPUS4-51472 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Xu, Yong A1 - Huang, Yanliang A1 - Yang, D. A1 - Kunte, Hans-Jörg A1 - De Marco, R. A1 - Wang, X. T1 - Investigation of the calcareous deposits formation controlled by interfacial pH and its effect on the hydrogen entry into AISI 4135 steel in seawater N2 - The influence of interfacial pH between AISI 4135 steel and seawater under different polarization potentials on the formation of calcareous deposits has been studied. An interfacial pH of 9.61 at −0.9 V vs. SCE using state of the art iridium oxide microelectrode was found to be the critical pH for the precipitation of magnesium hydroxide. Calcareous deposits with a double-layer structure comprising an inner-brucite layer and an outer-aragonite layer were found to form at potentials between −1.0 V and −1.2 V vs. SCE. Furthermore, the facilitation of hydrogen permeation into steel induced by the formation of calcareous deposits was verified using the Devanathan-Stachurski electrochemical test. The mechanism of calcareous deposits facilitates hydrogen permeation into steel is related to its inhibition on hydrogen recombination and escape processes. KW - Low alloy steel KW - Cathodic protection KW - Calcareous deposits KW - Hydrogen permeation PY - 2020 U6 - https://doi.org/10.1016/j.ijhydene.2020.11.040 SN - 0360-3199 SP - 1 EP - 18 PB - Elsevier Ltd. AN - OPUS4-51730 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - An, Biwen Annie A1 - Kunte, Hans-Jörg A1 - Koerdt, Andrea ED - Eibergen, N. ED - Poulassichidis, T. T1 - Microbiologically Influenced Corrosion (MIC) by Halophilic (Salt-Loving) Nitrate and Sulfate-Reducing Microorganisms N2 - The survey of Canadian shale sites showed a dominance of halophilic microorganisms, including Halomonas (HA). Nitrate-amended incubations of the field samples under high salinity (14.6% NaCl), revealed a dominance of HA (>72%) and an accumulation of nitrite. Nitrite accumulation directly inhibited the growth of SRB, thereby decreasing their souring and corrosion risks. However, accumulated nitrite may also contribute to iron corrosion, which will be tested by using different concentrations of nitrate as an electron acceptor to HA. Different salinities are further tested on HA strains supplemented with iron coupons to determine their effects on iron corrosion rates. HA incubated with separate cultures of corrosive methanogen and SRB were tested to determine whether a positive or adverse effect will occur between them. Lastly, analyses of iron coupons will be conducted using TOF-SIMS, FIB-SEM and EDS for corrosion product characterization T2 - Corrosion 2021 CY - Online Meeting DA - 19.04.2021 KW - MIC KW - Bacteria KW - Halophile KW - Corrosion KW - Environmental condition KW - Korrosion KW - High salinity PY - 2021 UR - https://my.nace.org/PaperTrail/Authors/Submission.aspx?id=2914f145-7f8f-ea11-813a-005056a95a7c SP - Paper C2021-16284, 1 AN - OPUS4-52479 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hahn, Marc Benjamin A1 - Meyer, Susann A1 - Kunte, Hans-Jörg A1 - Solomun, Tihomir A1 - Sturm, Heinz T1 - Measurements and simulations of microscopic damage to DNA in water by 30 keV electrons: A general approach applicable to other radiation sources and biological targets N2 - The determination of the microscopic dose-damage relationship for DNA in an aqueous environment is of a fundamental interest for dosimetry and applications in radiation therapy and protection. We combine geant4 particle-scattering simulations in water with calculations concerning the movement of biomolecules to obtain the energy deposit in the biologically relevant nanoscopic volume. We juxtaposition these results to the experimentally determined damage to obtain the dose-damage relationship at a molecular level. This approach is tested for an experimentally challenging system concerning the direct irradiation of plasmid DNA (pUC19) in water with electrons as primary particles. Here a microscopic target model for the plasmid DNA based on the relation of lineal energy and radiation quality is used to calculate the effective target volume. It was found that on average fewer than two ionizations within a 7.5-nm radius around the sugar-phosphate backbone are sufficient to cause a single strand break, with a corresponding median lethal energy deposit being E1/2=6±4 eV. The presented method is applicable for ionizing radiation (e.g., γ rays, x rays, and electrons) and a variety of targets, such as DNA, proteins, or cells. KW - DNA KW - Radiation damage KW - Dosimetry KW - DNA radiation damage KW - Single-strand break (SSB) KW - Double-strand break (DSB) KW - Low energy electrons KW - Monte Carlo simulation KW - Radiation damage to biomolecules KW - Plasmid DNA in water KW - Lethal dose KW - Solutions (pH, salinity, cosolutes) KW - Geant4 KW - Microdosimetry PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-404244 SN - 2470-0045 SN - 2470-0053 VL - 95 IS - 5 SP - 052419-1 EP - 052419-8 PB - American Physical Society CY - USA AN - OPUS4-40424 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Pfeiffer, F. A1 - Bagyan, I. A1 - Alfaro Espinoza, Gabriela A1 - Zamora-Lagos, M.-A. A1 - Habermann, B. A1 - Marin-Sanguino, A. A1 - Oesterhelt, D. A1 - Kunte, Hans-Jörg T1 - Revision and reannotation of the Halomonas elongata DSM 2581(T) genome N2 - The genome of the Halomonas elongata type strain DSM 2581, an industrial producer, was reevaluated using the Illumina HiSeq2500 technology. To resolve duplication-associated ambiguities, PCR products were generated and sequenced. Outside of duplications, 72 sequence corrections were required, of which 24 were point mutations and 48 were indels of one or few bases. Most of these were associated with polynucleotide stretches (poly-T stretch overestimated in 19 cases, poly-C underestimated in 15 cases). These problems may be attributed to using 454 technology for original Genome sequencing. On average, the original genome sequence had only one error in 56 kb. There were 23 frameshift error corrections in the 29 protein-coding genes affected by sequence revision. The genome has been subjected to major reannotation in order to substantially increase the annotation quality. KW - Frameshift KW - Genome annotation KW - Genome sequencing KW - Halomonas elongata KW - Halophilic bacteria KW - Sequence revision PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-403292 SN - 2045-8827 VL - 6 IS - 4 SP - Article e465, 1 EP - 6 PB - Wiley AN - OPUS4-40329 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Meyer, Susann A1 - Schröter, Maria-Astrid A1 - Hahn, Marc Benjamin A1 - Solumon, Tihomir A1 - Strum, Heinz A1 - Kunte, Hans-Jörg T1 - Ectoine can enhance structural changes in DNA in vitro N2 - Strand breaks and conformational changes of DNA have consequences for the physiological role of DNA. The natural protecting molecule ectoine is beneficial to entire bacterial cells and biomolecules such as proteins by mitigating detrimental effects of environmental stresses. It was postulated that ectoine-like molecules bind to negatively charged spheres that mimic DNA surfaces. We investigated the effect of ectoine on DNA and whether ectoine is able to protect DNA from damages caused by ultraviolet radiation (UV-A). In order to determine different isoforms of DNA, agarose gel electrophoresis and atomic force microscopy experiments were carried out with plasmid pUC19 DNA. Our quantitative results revealed that a prolonged incubation of DNA with ectoine leads to an increase in transitions from supercoiled (undamaged) to open circular (single-strand break) conformation at pH 6.6. The effect is pH dependent and no significant changes were observed at physiological pH of 7.5. After UV-A irradiation in ectoine solution, changes in DNA conformation were even more pronounced and this effect was pH dependent. We hypothesize that ectoine is attracted to the negatively charge surface of DNA at lower pH and therefore fails to act as a stabilizing agent for DNA in our in vitro experiments. KW - Ectoine KW - DNA KW - UV radiation PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-413139 VL - 7 IS - 1 SP - Article 7170, 1 EP - 10 AN - OPUS4-41313 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Hahn, Marc Benjamin A1 - Susann, Meyer A1 - Schröter, Maria-Astrid A1 - Kunte, Hans-Jörg A1 - Solomun, Tihomir A1 - Sturm, Heinz T1 - DNA protection by ectoine from ionizing radiation: molecular mechanisms N2 - Ectoine, a compatible solute and osmolyte, is known to be an effective protectant of biomolecules and whole cells against heating, freezing and extreme salinity. Protection of cells (human keratinocytes) by ectoine against ultraviolet radiation has also been reported by various authors, although the underlying mechanism is not yet understood. We present the first electron irradiation of DNA in a fully aqueous environment in the presence of ectoine and at high salt concentrations. The results demonstrate effective protection of DNA by ectoine against the induction of single-strand breaks by ionizing radiation. The effect is explained by an increase in low-energy electron scattering at the enhanced free-vibrational density of states of water due to ectoine, as well as the use of ectoine as an ˙OH-radical scavenger. This was demonstrated by Raman spectroscopy and electron paramagnetic resonance (EPR). KW - Ectoine KW - DNA KW - Radiation protection KW - Ionizing radiation KW - Compatible solute KW - Biomolecules KW - Sodium chloride KW - Aqueous solution KW - Hydroxyectoine KW - Raman spectroscopy KW - Electron irradiation KW - Cancer KW - Radical scavenger KW - Low energy electrons KW - Hydroxyl radical KW - OH-radical KW - Ectoin KW - UV radiation KW - Sun KW - Salt KW - Radiation therapy PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-419332 UR - http://pubs.rsc.org/en/content/articlehtml/2017/cp/c7cp02860a SN - 1463-9076 VL - 19 IS - 37 SP - 25717 EP - 25722 PB - Royal Society of Chemistry CY - United Kingdom AN - OPUS4-41933 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schröter, Maria-Astrid A1 - Meyer, Susann A1 - Hahn, Marc Benjamin A1 - Solomun, Tihomir A1 - Sturm, Heinz A1 - Kunte, Hans-Jörg T1 - Ectoine protects DNA from damage by ionizing radiation N2 - Ectoine plays an important role in protecting biomolecules and entire cells against environmental stressors such as salinity, freezing, drying and high temperatures. Recent studies revealed that ectoine also provides effective protection for human skin cells from damage caused by UV-A radiation. These protective properties make ectoine a valuable compound and it is applied as an active ingredient in numerous pharmaceutical devices and cosmetics. Interestingly, the underlying mechanism resulting in protecting cells from radiation is not yet fully understood. Here we present a study on ectoine and its protective influence on DNA during electron irradiation. Applying gel electrophoresis and atomic force microscopy, we demonstrate for the first time that ectoine prevents DNA strand breaks caused by ionizing electron radiation. The results presented here point to future applications of ectoine for instance in cancer radiation therapy. KW - Plasmid DNA pUC19 KW - Electron irradiation 30 [kV] KW - Effective irradiation dose 0.2-16 [Gy] KW - Gel electrophoresis KW - AFM intermittent contact KW - Radioprotector ectoine KW - Compatible solute PY - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-428287 SN - 2045-2322 VL - 7 IS - 1 SP - 15272, 1 EP - 15272, 7 PB - Nature AN - OPUS4-42828 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Huang, A1 - Zheng, A1 - Zhang, A1 - Lu, A1 - Wang, A1 - Kunte, Hans-Jörg A1 - Sand, W. T1 - Long Term Temperature and Humidity Evolution Forecast in Near Field of Nuclear Waste Container N2 - The paper aims to simulate the corrosion environment of nuclear waste containers at different geological disposal periods to obtain temperature and humidity information at the interface between the container and the surrounding environment. The simulated data and calculation results of long term temperature evolution at the surface of nuclear waste (HLW) containers from some typical nuclear countries about the safety disposition were reviewed. Combining different burial patterns, this paper speculated the long term temperature evolution rule for China. According to the study about saturation variation of buffer/backfill material at home and abroad, the humidity evolution of bentonite at the surface of HLW containers was specu- lated. The study showed that the temperature of the container surface increased rapidly at the beginning, and gradually decreased after the climax. For safety reasons, the maximum temperature was designed below 100°C. The saturation of bentonite was af- fected by the mutual influences of the heat released by nuclear waste decay and the infiltration of groundwater. It was dominated by the released heat in the early stage, and later was influenced greatly by the infiltration of groundwater. It is generally believed that the water content at the surface of the container will increase obviously in about 3 years, and will be saturated in about 10 years. The prediction of long-term temperature and humidity evolution will lay a foundation for study of corrosion evolution of nuclear waste containers in China. KW - Corrosion evolution KW - Nuclear waste KW - Disposal repository KW - Temperature and humidity PY - 2018 SN - 1672-9242 VL - 15 IS - 10 SP - 109 EP - 113 PB - Zhongguo Bingqi Gongye di-wujiu Yanjiusuo CY - Chongqing AN - OPUS4-46953 LA - zho AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Huang, Y.-L. A1 - Zheng, A1 - Zhang, A1 - Lu, A1 - Wang, A1 - Kunte, Hans-Jörg A1 - Sand, W. T1 - Forecast on Long Term Chemical Environment Evolution on Surface of Nuclear Waste Container N2 - Surface environment changes of nuclear waste container, mainly including oxygen contents, buffer pore water components and pH under the conditions of “borehole” type and “In-floor” type with bentonite buffer/backfill and concrete buffer were summarized. This summarization provides a basic corrosion environment reference for the corrosion evolution re- search of high-level radioactive waste disposal repository in our country. KW - Corrosion evolution KW - Nuclear waste KW - Disposal repository KW - Corrosion environment PY - 2018 U6 - https://doi.org/10.7643/ issn.1672-9242.2018.10.017 SN - 1672-9242 VL - 15 IS - 10 SP - 103 EP - 108 PB - Zhongguo Bingqi Gongye di-wujiu Yanjiusuo CY - Chongqing AN - OPUS4-46952 LA - zho AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -