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Titel:Charakterisierung metabolisch veränderter Glycolipide in Ustilago maydis & Identifizierung neuer pilzlicher Glycolipide
Autor:Deinzer, Hans-Tobias
Weitere Beteiligte: Bölker, Michael (Prof. Dr.)
Veröffentlicht:2017
URI:https://archiv.ub.uni-marburg.de/diss/z2017/0240
URN: urn:nbn:de:hebis:04-z2017-02408
DOI: https://doi.org/10.17192/z2017.0240
DDC: Biowissenschaften, Biologie
Titel(trans.):Characterization of metabolically engineered glycolipids of Ustilago maydis&Identification of novel fungal glycolipids
Publikationsdatum:2017-05-24
Lizenz:https://creativecommons.org/licenses/by-nc-sa/4.0

Dokument

Schlagwörter:
Biotenside, Glycolipide, fungal secondary metabolites, Glycolipide, glycolipides, pilzliche Sekundärmatabolite, Ustilago zeae

Zusammenfassung:
Sekundärmetabolite sind chemische Stoffe, die von Pilzen, Bakterien und Pflanzen syntheti- viert werden. Sie sind im Gegensatz zu den Primärmetaboliten nicht für das Überleben eines Organismus notwendig, bringen aber Vorteile gegenüber anderen Organismen mit sich. Zu den Sekundärmetaboliten zählen Biotenside, bei denen es sich um mikrobiell hergestellte, chemisch strukturell gleiche Tenside handelt. Sie bestehen aus einem hydrophoben und einem hydrophilen Teil. Dieser amphiphile Aufbau führt zu typischen oberflächenaktiven Eigenschaften. Im Ge- gensatz zu industriell chemisch hergestellten Tenside sind Biotenside unter anderem biologisch abbaubar, ungiftig und hitzestabil. Glycolipide sind eine Gruppe von Biotensiden. Sie bestehen immer aus einem oder mehreren Zuckern, die mit Hydroxyfettsäuren verbunden sind. Im Reich der Pilze gibt es viele verschiedene Glycolipide, zu denen auch die von dem phytopathogenen Basidiomycet phytopathogenen Basidiomycet Ustilago maydis synthetisierte Ustilaginsäure und Mannosylerythritollipide zählen. Diese beiden Glycolipide werden unter Stickstoffmangelbedin- gungen synthetisiert. Die Ustilaginsäure besitzt eine starke Oberflächenaktivität und wirkt anti- biotisch gegen Hefen und Gram-positive Bakterien, wohingegen Mannosylerythritollipide zudem hämolytische Aktivität zeigen. Die Biosynthesewege dieser beiden Glycolipide wurden bereits gut charakterisiert. Bei dem Screening vieler verschiedenster Pilzisolate wurden zwei weitere Basidiomyceten, Ma- calpinomyces eriachnes und Sporisorium scitamineum, gefunden, die ebenfalls in der Lage sind verschiedene Varianten dieser Glycolipide zu synthetisieren. Bei diesen handelt es sich einen um Macalpinomyces eriachnes und Sporisorium scitamineum. Die Glycolipide von M. eriachnes sind oberflächenaktiv und wirken hämolytisch. Die Glycolipide von S. scitamineum hingegen sind auch oberflächenaktiv und wirken antibiotisch gegenüber Hefen. Neben diesen beiden phy- topathogenen Pilzen wurde ein weiterer Pilz gefunden, der in der Lage war einen zuckerhaltigen Sekundärmetabolit zu synthetisieren. Durch Sequenzierung konnte diese schwarze Hefe als Do- thiora cannabinae identifiziert werden. Unter Stickstoffmangelbedingungen ist D. cannabinae in der Lage eine glycolipidartige Substanz zu synthetisieren. Mit Hilfe von Massenspektrometrie und Kernspinresonanzspektroskopie konnte die Substanz als oberflächaktives und hämolytisches Glyco-Oligo-Hydroxy-Alkanoat identifiziert werden. Die Hexadecansäure der Ustilaginsäure liegt in U. maydis in di- und trihydroxylierter Form vor. Die Hydroxylierung wird von den beiden P450 Monooxigenasen Cyp1 und Cyp2 kataly- siert. Die Überexpression von Cyp1 in einem Ustilaginsäure defiziten U. maydis Stamm führt zu einer Monohydroxylierung der Hexadecansäure der MELs. Die Überexpression beider P450 Mo- nooxigenasen führt zur Dihydroxylierung. Diese Modifikation der MELs hatte allerdings keinen Einfluss auf die Eigenschaften der Glycolipide.

Summary:
Secondary metabolites are chemical compounds, that are synthesized by fungi, bacteria and plants. In contrast to primary metabolites, secondary metabolites are not necessary for the sur- vival of an organism, but have advantages over other organisms. The secondary metabolites include microbial biosurfactants, which always show the same composition. They consist of a hy- drophilic and a hydrophobic part. This amphiphilic structure leads to the typical surface-active properties. In contrast to industrially produced surfactants, biosurfactants are biodegradable, non-toxic and heat-stable. Glycolipids are a group of biosurfactants. They consist of one or more sugars, which are decorated with hydroxy fatty acids. In the kindom of fungi many different glyco- lipids can be found, including ustilagic acid and mannosylerythritol lipids, which are synthesized by the phytopathogenic basidiomycete Ustilago maydis. These two glycolipids are synthesized under nitrogen starvation conditions. Ustilagic acid has a strong surface activity and shows an antibiotic effect against yeasts and Gram-positive bacteria. Besides the strong surface activity, the mannosylerythritol lipids also exhibit haemolytic activity. The biosynthesis pathways of these two glycolipids have already been well characterized. During the screening of many different fungal isolates, two further basidiomycetes have been found, named Macalpinomyces eriachnes and Sporisorium scitamineum, which are also able to synthesize different variants of these glycolipids. The glycolipids produced by M. eriachnes are surface-active and have a hemolytic effect. S. scitamineum glycolipids are also surface-active and have an antibiotic effect towards yeasts. Besides these two phytopathogenic fungi, another unidentified fungus was found which is able to produce a sugar containing secondary metabolite. By sequencing, this black yeast was identified as Dothiora cannabinae. Under nitrogen starvation conditions, D. cannabinae is able to synthesize a glycolipid-like substance. By mass spectrometry analysis and nuclear magnetic resonance spectroscopy the substance was identified as glyco-oligo- hydroxy-alkanoate. This substance is surface active and shows haemolytic activity. The hexadecanoic acid of the ustilagic acid is found in U. maydis in di- and trihydroxylated form. The hydroxylations are catalyzed by the two P450 monooxigenases Cyp1 and Cyp2. The overexpression of Cyp1 in a U. maydis strain, which does not synthesize ustilagic acid, results in a monohydroxylation of the hexadecanoic acid of the MELs. The overexpression of both P450 monooxigenases leads to a dihydroxylation of the hexadecanoic acid of the MELs. However, this modification of the MELs had no effect on the properties of this glycolipids.

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