Regulatory and metabolic aspects of the phosphate starvation response of Corynebacterium glutamicum [Elektronische Ressource] / vorgelegt von Han Min Woo

Han Min Woo

Sujets

• Biologie

Informations

• Publié par: heinrich-heine-universitat_dusseldorf
• Publié le: 01 janvier 2010
• Nombre de lectures: 18
• Langue: English
• Poids de l'ouvrage: 20 Mo

Extrait

Regulatory and metabolic aspects
of the phosphate starvation response
of Corynebacterium glutamicum

Inaugural-Dissertation



zur Erlangung des Doktorgrades
der Mathematisch-Naturwissenschaftlichen Fakultät
der Heinrich-Heine-Universität Düsseldorf

vorgelegt von
Han Min Woo
aus Ulsan, Südkorea

Jülich, Juni 2010 The thesis in hand has been performed at the Institute of Biotechnology 1,
Forschungszentrum Jülich, from June 2007 until May 2010 under the supervision of Prof. Dr.
M. Bott.






Printed with the permission of the Mathematisch-Naturwissenschaftliche Fakultät of the
Heinrich-Heine-Universität Düsseldorf








Examiner: Prof. Dr. Michael Bott
Institute of Biotechnology 1
Forschungszentrum Jülich

Coexaminer: Prof. Dr. Martina Pohl
Institute of Biotechnology 2
Forschungszentrum Jülich

Oral examination: July 12, 2010
Results described in this dissertation have been submitted or will be submitted for
publication in the following articles:





Woo, H.M., Wessel, M. and Bott, M. (2010) SenX3-RegX3 (CgtS4-CgtR4), an essential two-
component regulatory system of Corynebacterium glutamicum involved in the phosphate
starvation response. To be submitted.


Woo, H.M., Noack, S., Seibold, G, Willbold, S., Eikmanns, B.J., and Bott, M. (2010) A link
between phosphate starvation response and glycogen metabolism in Corynebacterium
glutamicum revealed by metabolomics. Appl. Environ. Microbiol. In press (AEM01375-10
Version 2)






Content I
Content
CONTENT................................................................................................................................I 
1  ABSTRACT..................................................................................................................1 
2  ZUSAMMENFASSUNG ...............................................................................................2 
3  INTRODUCTION ..........................................................................................................3 
3.1  Phosphorus – an essential element for cellular growth and regulation................................ 3 
3.2  Phosphate starvation responses of microorganisms............................................................... 6 
3.2.1  Escherichia coli and the two-component systems PhoBR and CreBC ............................... 6 
3.2.2  Bacillus subtilis and the regulatory systems PhoPR, ResDE and Spo0A............................ 8 
3.2.3  Mycobacterium and the two-component systems PhoPR and SenX3-RegX3 .................. 11 
3.2.4  Streptomyces and the two-component system PhoPR....................................................... 13 
3.3  Phosphate starvation responses of Corynebacterium glutamicum....................................... 14 
3.3.1  The two-component regulatory system PhoSR ................................................................. 15 
3.3.2 latory system SenX3-RegX3..................................................... 16 
3.4  Development of microbial metabolomics .............................................................................. 17 
3.4.1  Sample preparation............................................................................................................ 18 
3.4.2  Metabolome analysis......................................................................................................... 19 
3.5  Aims of this work..................................................................................................................... 20 
4  RESULTS...................................................................................................................22 
4.1  SenX3-RegX3 (CgtS4-CgtR4), an essential two-component regulatory system of C.
glutamicum involved in the phosphate starvation response................................................ 24 
4.2  A link between phosphate starvation and glycogen metabolism in C. glutamicum revealed
by metabolomics ..................................................................................................................... 56 
5  DISCUSSION .............................................................................................................88 
5.1  Genetic regulation of P starvation responses in C. glutamicum.......................................... 88 i
5.1.1  Regulation of P -dependent genes by PhoRS and SenX3-RegX3 ..................................... 89 i II Content
5.1.2  The essentiality of the regX3 gene .................................................................................... 92 
5.1.3  Regulatory model of the phosphate starvation response of C. glutamicum ...................... 96 
5.2  Cross-regulation of carbon metabolism and the P starvation response ............................ 96 i
5.2.1  Glycogen metabolism of C. glutamicum under P limitation............................................. 98 i
5.2.2  Glucose uptake under P limitation in C. glutamicum ..................................................... 100 i
6  REFERENCES .........................................................................................................102 
7  APPENDIX ...............................................................................................................111 
7.1  Influence of the phosphate limitation on L-lysine producer, C. glutamicum DM1945.... 111 

Abbreviations III
Abbreviations
ATC Anhdyrotetracycline
ATCC American Type Culture Collection
ATP/ADP Adenosine triphosphate/Adenosine diphosphate
BHI(S) Brain Heart Infusion (+ Sorbitol)
RCm Chloramphenicol resistance
DTT Dithiothreitol
EMSA Electrophoretic gel mobility shift assay
et al. et alii
GC-Tof-MS Gas chromatography time-of-flight mass spectrometry
IPTG Isopropyl-thio--D-galactopyranoside
R Kan Kanamycin resistance
LB Lysis Broth
MSTFA N-methyl-N-(trimethylsilyl)trifluoroacetamide
OD Optical density at 600 nm 600
P Phosphorus
PAGE Polyacrylamide gel electrophoresis
P Inorganic phosphate i
31P-NMR Phosphorus nuclear magnetic resonance
RBS Ribosome binding site
SDS Sodiumdodecylsulfate
TBE Tris base - Boric acid - EDTA
TCA Trichloric acid
TCS Two-component regulatory system
TE Tris base – EDTA
TEMED N,N,N,N,-Tetramethylethylenediamine
TNI Tris base – NaCl - Imidazol
v/v Volume per volume
w/v Weight per volume

Further abbreviations not included in this section are according to international standards, as
for example listed in the author guidelines of the FEBS Journal.