Molecular investigations of peptidoglycan binding proteins in Listeria monocytogenes [Elektronische Ressource] / vorgelegt von Silke Machata

Sujets

• Biologie

Informations

• Publié par: justus-liebig-universitat_giessen
• Publié le: 01 janvier 2008
• Nombre de lectures: 40
• Poids de l'ouvrage: 2 Mo

Extrait

Aus dem Institut für Medizinische Mikrobiologie







Molecular investigations of peptidoglycan-
binding proteins in Listeria monocytogenes






Inauguraldissertation
zur Erlangung des Doktorgrades des Naturwissenschaftlichen
Fachbereichs
der Justus-Liebig-Universität Gießen



Vorgelegt von Silke Machata
aus Frankfurt, Deutschland



Gießen, 2008








Gutachter: Prof. Dr. Trinad Chakraborty


Gutachter: Prof. Dr. Alfred Pingoud















Tag der mündlichen Prüfung: 24. 06. 2008Publikationsliste:
Teilergebnisse aus dieser Arbeit wurden mit in den folgenden Beiträgen vorab veröffentlicht:

Publikationen:
Machata, S., T. Hain, M. Rohde, T. Chakraborty. 2005. Simultaneous deficiency of both
MurA and p60 proteins generates a rough phenotype in Listeria monocytogenes. J Bacteriol
187(24):8385-94

Chatterjee, S.S., H. Hossain, S. Otten, C. Kuenne, K. Kuchmina, S. Machata, E. Domann, T.
Chakraborty, T. Hain. 2006. Intracellular gene expression profile of Listeria monocytogenes.
Infect Immun 74(2):1323-38

Hain T., H. Hossain, S.S. Chatterjee, S. Machata, U. Volk, S. Wagner,. B. Brors, S. Haas,
C.T. Kuenne, A. Billion, S. Otten, J. Pane-Farre, S. Engelmann, T. Chakraborty. 2008
Temporal transcriptomic analysis of the Listeria monocytogenes EGD-e sigmaB regulon.
BMC Microbiol 28;8:20

Machata, S., S. Tchatalbachev, W. Mohamed, L. Jänsch, T. Hain, T. Chakraborty.
Lipoproteins of Listeria monocytogenes are critical for virulence and TLR2-mediated
immune activation. Accepted at Journal of Immunology.


Tagungsbeiträge:
Machata, S., T. Hain, M. Rohde, T. Chakraborty. (Poster) Simultaneous deficiency of both
MurA and p60 proteins generates a rough phenotype in Listeria monocytogenes. DGHM,
Göttingen, Deutschland (2005)

Machata, S., T. Hain, T. Chakraborty. (Poster) Characterization of mutants of putative
murein hydrolases in Listeria monocytogenes. Meeting of the Network of Excellence (NoE)
EuroPathoGenomics, Celakovic, Tschechien (2007) TABLE OF CONTENTS I
TABLE OF CONTENTS....................................................................................................................................................... I
LIST OF ABBREVIATIONS...............................................................................................................................................V
1. INTRODUCTION ......................................................................................................................................................1
1.1. CHARACTERISTICS OF LISTERIA........................................................................................................................1
1.2. VIRULENCE OF LISTERIA MONOCYTOGENES ......................................................................................................1
1.3. PROPERTIES AND STRUCTURE OF THE BACTERIAL CELL WALL........................................................................8
1.4. CELL WALL HYDROLASES ............................................................................................................................. 10
1.5. DETERMINATION AND MAINTENANCE OF CELL SHAPE ................................................................................. 11
1.6. LIPOPROTEINS IN L. MONOCYTOGENES .......................................................................................................... 13
1.7. HOST DEFENSE AGAINST MICROBES............................................................................................................. 14
1.7.1. Pattern recognition receptors (PRR) ........................................................................................................ 15
1.7.2. Antimicrobial peptides ............................................................................................................................. 17
1.8. AIMS OF THIS WORK...................................................................................................................................... 18
2. MATERIALS AND METHODS ............................................................................................................................ 20
2.1. MICE.............................................................................................................................................................. 20
2.2. CELL CULTURE.............................................................................................................................................. 20
2.3. ANTIBODIES .................................................................................................................................................. 21
2.4. BACTERIAL STRAINS ..................................................................................................................................... 21
2.5. PLASMIDS AND OLIGONUCLEOTIDES ............................................................................................................ 22
2.6. GROWTH MEDIA ............................................................................................................................................ 25
2.7. ANTIBIOTICS ................................................................................................................................................. 26
2.8. SOLUTIONS, BUFFERS AND STANDARDS........................................................................................................ 26
2.9. EQUIPMENT ................................................................................................................................................... 28
2.10. BACTERIAL CULTURES .................................................................................................................................. 29
2.10.1. Cultivation of strains ................................................................................................................................ 29
2.10.2. Measurement of bacterial growth............................................................................................................. 29
2.10.3. Autolysis assay ......................................................................................................................................... 29
2.10.4. Antibiogram.............................................................................................................................................. 30
2.10.5. Swarming motility assay .......................................................................................................................... 30
2.11.6. Biofilm assay ............................................................................................................................................ 30
2.11. DNA AND GENETIC MANIPULATIONS ........................................................................................................... 30
2.11.1. Chromosomal DNA isolation................................................................................................................... 30
2.11.2. Plasmid isolation and purification............................................................................................................ 31
2.11.3. Agarose gel electrophoresis ..................................................................................................................... 31
2.11.4. Enzymatic modification of DNA ............................................................................................................. 31
2.11.5. Cell transformation................................................................................................................................... 32
2.11.6. Polymerase chain reaction (PCR) ............................................................................................................ 33
2.11.7. Generation of deletion mutants ................................................................................................................ 34
2.12. MICROSCOPY................................................................................................................................................. 34
2.13. RNA.............................................................................................................................................................. 35 TABLE OF CONTENTS II
2.13.1. RNA isolation and purification ................................................................................................................ 35
2.13.2. Quantitative PCR analysis........................................................................................................................ 35
2.14. CELL CULTURE AND VIRULENCE STUDIES..................................................................................................... 36
2.14.1. Infection assay .......................................................................................................................................... 36
2.14.2. Luciferase reporter assay.......................................................................................................................... 36
2.15. PROTEINS ..............................................