Core Facility Hohenheim

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Browsing Core Facility Hohenheim by Subject "Biosurfactants"

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    Genetic code expansion for controlled surfactin production in a high cell-density Bacillus subtilis strain
    (2025) Hermann, Alexander; Hiller, Eric; Hubel, Philipp; Biermann, Lennart; Benatto Perino, Elvio Henrique; Kuipers, Oscar Paul; Hausmann, Rudolf; Lilge, Lars; Hermann, Alexander; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, 70599 Stuttgart, Germany; (A.H.); (E.H.); (L.B.); (E.H.B.P.); (R.H.); Hiller, Eric; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, 70599 Stuttgart, Germany; (A.H.); (E.H.); (L.B.); (E.H.B.P.); (R.H.); Hubel, Philipp; Core Facility Hohenheim, Mass Spectrometry Core Facility, University of Hohenheim, 70599 Stuttgart, Germany;; Biermann, Lennart; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, 70599 Stuttgart, Germany; (A.H.); (E.H.); (L.B.); (E.H.B.P.); (R.H.); Benatto Perino, Elvio Henrique; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, 70599 Stuttgart, Germany; (A.H.); (E.H.); (L.B.); (E.H.B.P.); (R.H.); Kuipers, Oscar Paul; Department of Molecular Genetics, University of Groningen, 9747 AG Groningen, The Netherlands;; Hausmann, Rudolf; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, 70599 Stuttgart, Germany; (A.H.); (E.H.); (L.B.); (E.H.B.P.); (R.H.); Lilge, Lars; Department of Bioprocess Engineering, Institute of Food Science and Biotechnology, University of Hohenheim, 70599 Stuttgart, Germany; (A.H.); (E.H.); (L.B.); (E.H.B.P.); (R.H.); Fouillaud, Mireille
    Background: In biotechnology, B. subtilis is established for heterologous protein production. In addition, the species provides a variety of bioactive metabolites, including the non-ribosomally produced surfactin lipopeptide. However, to control the formation of the target product-forming enzyme, different expression systems could be introduced, including the principle of genetic code expansion by the incorporation of externally supplied non-canonical amino acids. Methods: Integration of an amber stop codon into the srfA operon and additional chromosomal integration of an aminoacyl-tRNA synthetase/tRNA mutant pair from Methanococcus jannaschii enabled site-directed incorporation of the non-canonical amino acid O-methyl-L-tyrosine (OMeY). In different fed-batch bioreactor approaches, OMeY-associated surfactin production was quantified by high-performance thin-layer chromatography (HPTLC). Physiological adaptations of the B. subtilis production strain were analyzed by mass spectrometric proteomics. Results: Using a surfactin-forming B. subtilis production strain, which enables high cell density fermentation processes, the principle of genetic code expansion was introduced. Accordingly, the biosynthesis of the surfactin-forming non-ribosomal peptide synthetase (NRPS) was linked to the addition of the non-canonical amino acid OMeY. In OMeY-associated fed-batch bioreactor fermentation processes, a maximum surfactin titre of 10.8 g/L was achieved. In addition, the effect of surfactin induction was investigated by mass spectrometric proteome analyses. Among other things, adaptations in the B. subtilis motility towards a more sessile state and increased abundances of surfactin precursor-producing enzymes were detected. Conclusions: The principle of genetic code expansion enabled a precise control of the surfactin bioproduction as a representative of bioactive secondary metabolites in B. subtilis . This allowed the establishment of inducer-associated regulation at the post-transcriptional level with simultaneous use of the native promoter system. In this way, inductor-dependent control of the production of the target metabolite-forming enzyme could be achieved.
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    Structure elucidation and characterization of novel glycolipid biosurfactant produced by Rouxiella badensis DSM 100043T
    (2025) Harahap, Andre Fahriz Perdana; Conrad, Jürgen; Wolf, Mario; Pfannstiel, Jens; Klaiber, Iris; Grether, Jakob; Hiller, Eric; Vahidinasab, Maliheh; Salminen, Hanna; Treinen, Chantal; Perino, Elvio Henrique Benatto; Hausmann, Rudolf; Harahap, Andre Fahriz Perdana; Department of Bioprocess Engineering (150k), Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599 Stuttgart, Germany; (A.F.P.H.); (J.G.); (E.H.); (M.V.); (E.H.B.P.); Conrad, Jürgen; Department of Organic Chemistry (130b), Institute of Chemistry, University of Hohenheim, Garbenstr. 30, 70599 Stuttgart, Germany; (J.C.); (M.W.); Wolf, Mario; Department of Organic Chemistry (130b), Institute of Chemistry, University of Hohenheim, Garbenstr. 30, 70599 Stuttgart, Germany; (J.C.); (M.W.); Pfannstiel, Jens; Mass Spectrometry Unit, Core Facility Hohenheim, University of Hohenheim, Ottilie-Zeller-Weg 2, 70599 Stuttgart, Germany; (J.P.); (I.K.); Klaiber, Iris; Mass Spectrometry Unit, Core Facility Hohenheim, University of Hohenheim, Ottilie-Zeller-Weg 2, 70599 Stuttgart, Germany; (J.P.); (I.K.); Grether, Jakob; Department of Bioprocess Engineering (150k), Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599 Stuttgart, Germany; (A.F.P.H.); (J.G.); (E.H.); (M.V.); (E.H.B.P.); Hiller, Eric; Department of Bioprocess Engineering (150k), Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599 Stuttgart, Germany; (A.F.P.H.); (J.G.); (E.H.); (M.V.); (E.H.B.P.); Vahidinasab, Maliheh; Department of Bioprocess Engineering (150k), Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599 Stuttgart, Germany; (A.F.P.H.); (J.G.); (E.H.); (M.V.); (E.H.B.P.); Salminen, Hanna; Department of Food Material Science (150g), Institute of Food Science and Biotechnology, University of Hohenheim, Garbenstr. 21/25, 70599 Stuttgart, Germany;; Treinen, Chantal; Cellular Agriculture, TUM School of Life Sciences, Technical University of Munich, 85354 Freising, Germany;; Perino, Elvio Henrique Benatto; Department of Bioprocess Engineering (150k), Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599 Stuttgart, Germany; (A.F.P.H.); (J.G.); (E.H.); (M.V.); (E.H.B.P.); Hausmann, Rudolf; Department of Bioprocess Engineering (150k), Institute of Food Science and Biotechnology, University of Hohenheim, Fruwirthstr. 12, 70599 Stuttgart, Germany; (A.F.P.H.); (J.G.); (E.H.); (M.V.); (E.H.B.P.); Serianni, Anthony S.
    Microbial biosurfactants have become increasingly attractive as promising ingredients for environmentally friendly products. The reasons for this are their generally good performance and biodegradability, low toxicity, production from renewable raw materials, and benefits for the environment perceived by consumers. In this study, we investigated the chemical structure and properties of a novel glycolipid from a new biosurfactant-producing strain, Rouxiella badensis DSM 100043 T . Bioreactor cultivation was performed at 30 °C and pH 7.0 for 28 h using 15 g/L glycerol as a carbon source. The glycolipid was successfully purified from the ethyl acetate extract of the supernatant using medium pressure liquid chromatography (MPLC). The structure of the glycolipid was determined by one- and two-dimensional ( 1 H and 13 C) nuclear magnetic resonance (NMR) and confirmed by liquid chromatography electrospray ionization mass spectrometry (LC-ESI/MS). NMR analysis revealed the hydrophilic moiety as a glucose molecule and the hydrophobic moieties as 3-hydroxy-5-dodecenoic acid and 3-hydroxydecanoic acid, which are linked with the glucose by ester bonds at the C2 and C3 positions. Surface tension measurement with tensiometry indicated that the glucose–lipid could reduce the surface tension of water from 72.05 mN/m to 24.59 mN/m at 25 °C with a very low critical micelle concentration (CMC) of 5.69 mg/L. Moreover, the glucose–lipid demonstrated very good stability in maintaining emulsification activity at pH 2–8, a temperature of up to 100 °C, and a NaCl concentration of up to 15%. These results show that R. badensis DSM 100043 T produced a novel glycolipid biosurfactant with excellent surface-active properties, making it promising for further research or industrial applications.