Dokument: Physiologie eines industriellen Produktionsstammes: Proteinsekretion, Regulation und Produktion von Biotensiden in Burkholderia glumae
| Titel: | Physiologie eines industriellen Produktionsstammes: Proteinsekretion, Regulation und Produktion von Biotensiden in Burkholderia glumae | |||||||
| URL für Lesezeichen: | https://docserv.uni-duesseldorf.de/servlets/DocumentServlet?id=16468 | |||||||
| URN (NBN): | urn:nbn:de:hbz:061-20101021-104140-1 | |||||||
| Kollektion: | Dissertationen | |||||||
| Sprache: | Deutsch | |||||||
| Dokumententyp: | Wissenschaftliche Abschlussarbeiten » Dissertation | |||||||
| Medientyp: | Text | |||||||
| Autor: | Knorr, Janina [Autor] | |||||||
| Dateien: |
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| Beitragende: | Prof. Dr. Jaeger, Karl-Erich [Gutachter] Prof. Dr. Ernst, Joachim F. [Gutachter] | |||||||
| Stichwörter: | Burkholderia, Proteinsekretion, Biotenside | |||||||
| Dewey Dezimal-Klassifikation: | 500 Naturwissenschaften und Mathematik » 570 Biowissenschaften; Biologie | |||||||
| Beschreibungen: | Das Gram-negative Bakterium Burkholderia glumae besitzt hohe biotechnologische Relevanz durch die Sekretion einer Lipase (LipA), die bereits industriell Anwendung findet. Im Gegensatz zu dieser Lipase wurde das Bakterium bislang jedoch noch nicht eingehender charakterisiert. Somit konnte das volle biotechnologische Potential dieses vielseitigen Bakteriums noch nicht gezeigt werden. Da das Genom von B. glumae vollständig sequenziert und annotiert wurde, war ein Ziel dieser Arbeit neue biotechnologisch interessante Eigenschaften von Burkholderia glumae zu finden.
Wie andere Spezies der Burkholderia-Gattung besitzt B. glumae zwei Chromosomen. Der GC-Anteil der genomischen DNA beträgt in Chromosom 1 68,36 % bzw. 69,23 % in Chromosom 2. Bei näherer Betrachtung der open reading frames (ORFs) fällt die hohe Anzahl an ORFs auf, die für Transport- oder Sekretionsproteine kodieren. Der Transport der Lipase LipA vom Periplasma in das extrazelluläre Milieu wird über den Typ II Sekretionsmechanismus, auch main terminal branch des Gsp-Systems (general secretory pathway) genannt, postuliert. Dazu besitzt B. glumae ein vollständiges Typ II-Sekretionssystem auf Chromosom 1 und ein zweites rudimentäres System auf Chromosom 2. Durch die Untersuchung zweier Mutanten des Typ II-Systems konnte gezeigt werden, dass nur das Typ II-Sekretionssystem auf Chromosom 1 funktionsfähig ist und nicht durch das unvollständige System auf dem 2. Chromosom komplementiert wird. Die planktonische Lebensweise ist bei Mikroorganismen in natürlichen Habitaten eher selten. Stattdessen werden Biofilme gebildet, die den Bakterien auch in außergewöhnlichen Lebensräumen konstante Lebensbedingungen bieten. Im Rahmen dieser Arbeit konnte gezeigt werden, dass B. glumae Biofilme bildet. Es wurde nachgewiesen, dass die aktive Lipase LipA für die Biofilmbildung von B. glumae essentiell ist. Die Lipase hydrolysiert Öl zu einem Diacylglycerol und einer Oberflächen-aktiven Fettsäure. Diese nutzt B. glumae als Adhäsionsfaktor, wodurch sich das Bakterium z. B. an hydrophobe Oberflächen anlagern kann. Durch diese Erkenntnisse kann man nun die Verunreinigung der Fermenter durch Bildung von Biofilmen bei der Produktion der Lipase vermeiden, indem man andere Kohlenstoffquellen als Olivenöl einsetzt. Biofilme zeigen aber auch biotechnologische Relevanz, da z. B. in sogenannten Biofilmreaktoren die Biofilmbildung genutzt werden kann. Durch die Adhäsion der Mikroorganismen kann die Zelldichte erhöht, die Abführung toxischer Nebenprodukte oder die Abtrennung eines biotechnologischen Endproduktes vereinfacht werden. Biotenside sind durch Mikroorganismen hergestellte Moleküle, die nützliche Eigenschaften besitzen. Ihre einfache Herstellung und der biologische Abbau sind nur zwei der vielen Vorteile gegenüber chemisch hergestellten Komponenten. Rhamnolipide gehören zu den meistverbreiteten Biotensiden. Sie bestehen aus einem (Monorhamnolipid) oder zwei Rhamnosemolekülen (Dirhamnolipid), die mit zwei Fettsäuren verknüpft sind. Die Fettsäuren können in ihrer Kettenlänge variieren. Von einigen humanpathogenen Burkholderia-Spezies ist die Herstellung von Rhamnolipiden bekannt, während die Herstellung von Rhamnolipiden in B. glumae noch nicht beschrieben wurde. Innerhalb dieser Arbeit konnte gezeigt werden, dass B. glumae rhl-homologe Gene besitzt und Dirhamnolipide produziert. Das dominierende Dirhamnolipid besitzt zwei C14-Ketten. Nähere Untersuchungen der Rhamnolipide zeigten, dass diese eine bakterizide Wirkung gegen Gram-positive Bakterien besitzen, die bisher nur bei den Rhamnolipiden des humanpathogenen Bakteriums P. aeruginosa beschrieben wurde. Aus biotechnologischer Sicht und aufgrund des S1-Sicherheitsstatus können die Rhamnolipide von B. glumae als Alternative zu den Rhamnolipiden von P. aeruginosa dienen.The Gram-negative bacterium Burkholderia glumae has biotechnological relevance because it secrets of a lipase (LipA) which has industrial applications. Therefore, the lipase was already characterized in contrary to the bacterium. The full biotechnological potential of B. glumae was not shown until now. Because the complete genome was sequenced and annotated, the aim of the work was to investigate new biotechnological characteristics of B. glumae should. Like other species of the genus Burkholderia, B. glumae has two chromosomes. The GC-part of genomic DNA is 68,36 % in chromosome 1 and 69,23 % in chromosome 2. Analysis of open reading frames (orfs) showed a high quantity of orfs coding for transport or secretion proteins. The transport of the lipase LipA from periplasm to the extracellular media is postulated to occur via the type II secretion mechanism of the Gsp-System (general secretory pathway). B. glumae has a complete typ II secretion system encoded on chromosome 1 and an incomplete system on chromosome 2. After construction of two gsp-mutants, it was shown that only the system of chromosome 1 is functional and it cannot be complemented by the second system on chromosome 2. The planktonic life of microorganisms is really unusual in natural life. Microorganisms prefer to live in biofilms which offer a constant environment to organisms also in extremely habitats. Within this work, it was shown that B. glumae is able to form biofilms. The ability to form biofilms is dependent of the active lipase LipA. The lipase hydrolyses oil to a yield of fatty acids and a diacylglycerol. Fatty acids are surface-active compounds. B. glumae uses the produced fatty acids as an adhesion factor which allows the cells to attach to hydrophobic surfaces. This information is valuable to avoid contaminating biofilms at the production of lipase by using another carbon source than olive oil. Biofilms also have biotechnological relevance e. g. in biofilm reactors. A biofilm reactor has the advantage that the cell density can be increased or toxic components and end products can be isolated in an easy way. Biotensides are surface-active molecules which are produced by microorganisms. The inartificial synthesis and the lightened biodegradation of pesticides are only two of the many advantages of biotensides compared to chemical compounds. Rhamnolipids belongs to common biotensides. They consist of one (mono-rhamnolipid) or two rhamnose molecules (di-Rhamnolipid), which are linked to two fatty acids. The fatty acids can vary in length. Some humanpathogenic Burkholderia-species are able to produce rhamnolipids, whereas the synthesis of rhamnolipids by B. glumae was not described yet. It was shown that B. glumae has rhl-homologous genes and it produces a mixture of di-Rhamnolipids. The significant rhamnolipid has two fatty acids with C14-chains. Analysis of characteristics of rhamnolipids showed that the rhamnolipids exhibit an anti-microbial effect on Gram-positive bacteria. This effect was described for rhamnolipids of the humanpathogenic bacterium P. aeruginosa. Because of the S1-security level of B. glumae, the rhamnolipids of B. glumae can be used as an alternative to rhamnolipids of P. aeruginosa. | |||||||
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| Lizenz: |  Urheberrechtsschutz | |||||||
| Fachbereich / Einrichtung: | Mathematisch- Naturwissenschaftliche Fakultät | |||||||
| Dokument erstellt am: | 21.10.2010 | |||||||
| Dateien geändert am: | 13.10.2010 | |||||||
| Promotionsantrag am: | 16.04.0010 | |||||||
| Datum der Promotion: | 23.06.0010 |
