Electrostatic and non-electrostatic interactions of positively charged polypeptides with negatively charged lipid membranes [Elektronische Ressource] / von Christian Schwieger

Christian Schwieger

Sujets

• Biologie

Informations

• Publié par: martin-luther-universitat_halle-wittenberg
• Publié le: 01 janvier 2008
• Nombre de lectures: 19
• Langue: English
• Poids de l'ouvrage: 4 Mo

Extrait

Electrostatic and Non-Electrostatic Interactions
of Positively Charged Polypeptides
with Negatively Charged Lipid Membranes


Dissertation

zur Erlangung des akademischen Grades
Doctor rerum naturalium (Dr. rer. nat.)


vorgelegt der

Naturwissenschaftlichen Fakultät II (Chemie und Physik)
der Martin-Luther-Universität Halle-Wittenberg


von Herrn Diplomchemiker Christian Schwieger
geb. am 09. April 1976 in Halle (Saale)




Gutachter:
1. Prof. Dr. rer. nat. Alfred Blume
2. Prof. Dr. rer. nat. Andreas Janshoff

Tag der Verteidigung: 02. Feb. 2008
urn:nbn:de:gbv:3-000013374
[http://nbn-resolving.de/urn/resolver.pl?urn=nbn%3Ade%3Agbv%3A3-000013374]



































to my parents I
Contents

1 INTRODUCTION 1
1.1 Biological and model membranes............................................................................ 1
1.2 Lipid polymorphism ................................................................................................. 2
1.3 Motivation and aim of this work ............................................................................. 3
2 THE MODEL SYSTEM 5
2.1 The lipids ................................................................................................................... 5
2.2 The peptides............................................................................................................... 7
2.3 Membrane electrostatics ........................................................................................ 10
3 INTERACTION OF POLYLYSINE WITH PG CONTAINING MEMBRANES 13
3.1 Introduction............................................................................................................. 13
3.2 Differential scanning calorimetry.......................................................................... 14
3.2.1 Influence of PLL on the phase behaviour of pure DPPG membranes .......... 14
3.2.2 phase behaviour and the miscibility of mixed
DPPG/DPPC and DPPG/DMPC membranes ............................................... 16
3.3 Infrared spectroscopy............................................................................................. 20
3.3.1 Pure DPPG membranes................................................................................. 21
3.3.2 PLL complexes with mixed DPPG/DPPC membranes................................. 27
3.4 Isothermal titration calorimetry............................................................................ 29
3.4.1 Phase dependent binding............................................................................... 30
3.4.2 Chain length dependent binding ................................................................... 33
3.4.3 Temperature dependent binding 35
3.5 Monolayer experiments.......................................................................................... 38
3.6 X-ray diffraction ..................................................................................................... 40
3.6.1 Polymorphism and structure of DPPG in pure water.................................... 40
3.6.2 Polymo and structure of DPPG in NaCl solution .............................. 43
3.6.3 Influence of poly(L-lysine) on the bilayer structure ..................................... 44
3.7 Summary ................................................................................................................. 48 II CONTENTS
4 INTERACTION OF POLYARGININE WITH PG CONTAINING
MEMBRANES 51
4.1 Introduction............................................................................................................. 51
4.2 Differential scanning calorimetry.......................................................................... 52
4.2.1 Influence of PLA on the phase behaviour of DPPG membranes.................. 52
4.2.2 Influence of PLA binding on the miscibility of DMPC/DPPG
membranes .................................................................................................... 57
4.3 Isothermal titration calorimetry............................................................................ 61
4.4 Monolayer experiments.......................................................................................... 66
4.5 Infrared spectroscopy............................................................................................. 70
4.5.1 Complexes with pure DPPG ......................................................................... 70
4.5.2 Complexes with mixed DMPC/DPPG membranes ...................................... 83
4.6 Fluorescence experiments ...................................................................................... 86
4.7 Summary.................................................................................................................. 89
4.8 Comparison of PLL and PLA binding to PG membranes .................................. 92
5 INTERACTION OF POLYLYSINES AND POLYARGININES WITH PA
CONTAINING MEMBRANES 97
5.1 Thermotropic phase behaviour ............................................................................. 97
5.1.1 Polylysine / PA complexes ........................................................................... 97
5.1.2 Polyarginine / PA complexes........................................................................ 99
5.2 Domain formation in mixed DMPC/DMPA membranes .................................. 102
5.2.1 Influence of PLL binding............................................................................ 102
5.2.2 Influence of PLA binding 103
5.3 Binding enthalpies................................................................................................. 106
5.3.1 PLL binding to PA membranes................................................................... 106
5.3.2 PLA binding branes .................................................................. 108
5.4 The methylene stretching vibrations 109
5.4.1 Polylysine / PA complexes ......................................................................... 109
5.4.2 Polyarginine / PA complexes...................................................................... 110
5.5 Influence of polypeptide binding on the interfacial hydration ......................... 111
5.5.1 PLL binding ................................................................................................ 111
5.5.2 PLA binding 113
5.6 Secondary structure of the polypeptides............................................................. 114
5.6.1 PLL ............................................................................................................. 114
5.6.2 PLA 116 CONTENTS III
5.7 Polypeptide adsorption to DMPA monolayers................................................... 117
5.7.1 PLL adsorption ........................................................................................... 117
5.7.2 PLA adsorption 119
5.8 Summary and comparison of polypeptide binding to PG and PA
membranes ............................................................................................................ 121
6 SUMMARY 123
7 CONCLUSIONS 127
8 ZUSAMMENFASSUNG 128
9 APPENDIX 131
9.1 Materials................................................................................................................ 131
9.1.1 Polylysines.................................................................................................. 131
9.1.2 Polyarginines .............................................................................................. 131
9.1.3 Lipids .......................................................................................................... 131
9.1.4 Others 131
9.2 Experimental ......................................................................................................... 132
9.2.1 Vesicle preparation ..................................................................................... 132
9.2.2 Differential scanning calorimetry ............................................................... 132
9.2.3 Infrared spectroscopy.................................................................................. 132
9.2.4 Isothermal titration calorimetry .................................................................. 133
9.2.5 Monolayer adsorption experiments............................................................. 133
9.2.6 yer pressure/area isotherms ............................................................ 133
9.2.7 X-ray diffraction ......................................................................................... 134
9.2.8 Fluorescence experiments........................................................................... 134
9.3 Summarizing Tables............................................................................................. 136
10 REFERENCES 139
11 ACKNOWLEDGMENTS 151
12 PUBLICATIONS 153
13 CURRICULUM VITAE 155

V
Abbreviations

DPPG 1,2-dipalmitoyl-sn-glycero-3-phosphoglycerol
DMPG 1,2-myristoyl-sn-glycero-3-phosphoglycerol
POPG 1-palmitoyl-2-myristoyl-sn-glycero-3-phosphoglycerol
DPPC 1,2-dipalmitoyl-sn-glycero-