Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897):
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
Berlin
Mensch und Buch Verl.
2011
|
| Schriftenreihe: | Biological science
|
| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | 128 S. Ill., graph. Darst., Kt. |
| ISBN: | 9783863870553 |
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| 245 | 1 | 0 | |a Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) |c vorgelegt von Christof Grüring |
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Datensatz im Suchindex
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| adam_text | IMAGE 1
TABLE OF CONTENTS
I TABLE OF CONTENTS
SUMMARY 1
ZUSAMMENFASSUNG 3
CHAPTER 1: INTRODUCTION 6
1.1 MALARIA 6
1.1.1 PREVALENCE 6
1.1.2 THE ETIOLOGIC AGENT OF MALARIA 7
1.1.3 PATHOPHYSIOLOGY 7
1.1.4 DRUGS AND VACCINES 8
1.1.5 THE PLASMODIUM LIFE CYCLE 10
1.1.6 THE ASEXUAL LIFE CYCLE 11
1.2 ASEXUAL BLOOD STAGE BIOLOGY 13
1.2.1 INVASION 13
1.2.2 ACTIN AND ITS ROLE IN BLOOD STAGE BIOLOGY 15
1.2.3 THE APICOPLAST 16
1.2.4 THE FOOD VACUOLE 17
1.3 HOST CELL MODIFICATIONS 18
1.3.1 NEW PERMEATION PATHWAYS 19
1.3.2 MODIFICATION OF THE HOST CELL SURFACE 20
1.3.3 MAURER S CLEFTS 21
1.4 PROTEIN TRAFFICKING 22
1.4.1 APICOPLAST TARGETING 22
1.4.2 PROTEIN EXPORT TO THE HOST CELL 23
1.4.3 PNEPS 25
1.5 AIM OF THIS STUDY 26
CHAPTER 2: MATERIAL AND METHODS 27
2.1 MATERIALS 27
2.1.1 TECHNICAL AND MECHANICAL DEVICES 27
2.1.2 LABWARE AND DISPOSABLES 28
2.1.3 CHEMICALS AND BIOLOGICAL REAGENTS .29
2.1.4 KITS AND STANDARS 31
2.1.5 STOCK SOLUTIONS, BUFFERS AND MEDIA FOR P. FALCIPARUM CULTURE 31
2.1.6 STOCK SOLUTIONS, BUFFERS AND MEDIA FOR . CO//CULTURE 32
2.1.7 BUFFERS FOR COMPETENT E. CO//CELLS 33
2.1.8 BUFFERS AND SOLUTIONS FOR MOLECULAR BIOLOGY WORK 33
2.1.9 BUFFERS AND SOLUTIONS FOR BIOCHEMICAL WORK 33
2.1.10 BUFFERS FOR PROTEIN TRANSFER (WESTERN BLOT) 34
2.1.11 BACTERIAL AND PLASMODIUM STRAINS 34
2.1.12 ENZYMES FOR DNA AMPLIFICATION AND LIGATION 34
2.1.13 RESTRICTION ENZYMES 34
2.1.14 PRIMARY ANTIBODIES 35
2.1.15 SECONDARY ANTIBODIES 35
2.2 CELL BIOLOGICAL AND MOLECULAR PROCEDURES FOR P. FALCIPARUM 35 2.2.1
P. FALCIPARUM CULTURE 35
2.2.2 GIEMSA SMEARS 36
BIBLIOGRAFISCHE INFORMATIONEN HTTP://D-NB.INFO/1017394784
DIGITALISIERT DURCH
IMAGE 2
TABLE OF CONTENTS
2.2.3 PARASITE SYNCHRONIZATION 36
2.2.4 PARASITE FREEZING 36
2.2.5 THAWING OF PARASITES 36
2.2.6 TRANSFECTION OF P. FALCIPARUM 37
2.2.7 SAPONIN LYSIS 37
2.2.8 ISOLATION OF GENOMIC P. FALCIPARUM DNA 38
2.2.9 DNA PRECIPITATION 38
2.2.10 PERCOLL GRADIENT 38
2.2.11 TETANOLYSIN LYSIS 39
2.2.12 SOLUBILITY ASSAYS 39
2.3 MICROSCOPY 40
2.3.1 LIVE CELL IMAGING 40
2.3.2 IFA WITH ACETONE FIXED PARASITES 40
2.3.3 BODIPY-TR-C5-CERAMIDE STAINING 40
2.3.4 INHIBITOR TREATMENTS OF RING STAGES 41
2.3.5 CONFOCAL MICROSCOPY AND 4D IMAGING 41
2.3.6 DENDRA2 CONVERSION 42
2.4 BACTERIAL CULTURES 42
2.4.1 PRODUCTION OF CHEMICAL COMPETENT E. COLL BACTERIA 42
2.4.2 TRANSFORMATION OF E. COLI 42
2.4.3 FREEZING OF E. COLI 43
2.5 MOLECULAR BIOLOGICAL METHODS 43
2.5.1 AMPLIFICATION OF DNA SEQUENCES 43
2.5.2 PCR PRODUCT PURIFICATION 44
2.5.3 DNA RESTRICTION DIGEST 44
2.5.4 DNAIIGATION 44
2.5.5 SCREENING OF BACTERIAL CLONES 45
2.5.6 ISOLATION OF BACTERIAL PLASMIDS (MINI- AND MIDIPREP) 45
2.6 PROTEIN ANALYSIS 46
2.6.1 SDSPAGE 46
2.6.2 WESTERN BLOT ANALYSIS 46
CHAPTER 3: RESULTS 47
3.1 FOUR DIMENSIONAL IMAGING TO STUDY PARASITE DEVELOPMENT, HOST CELL
MODIFICATIONS AND PROTEIN EXPORT IN P. FALCIPARUM 47
3.1.1 ESTABLISHING 4D IMAGING 47
3.1.2 RING STAGE DEVELOPMENT 47
3.1.3 MORPHOLOGICAL CHANGES OF RING STAGE PARASITES 49
3.1.4 INFLUENCE OF CYTOSKELETON PERTURBING DRUGS ON RINGS STAGE
MORPHOLOGY ..50 3.1.5 TRANSITION FROM THE RING- TO THE TROPHOZOITE STAGE
52
3.1.6 FROM THE TROPHOZOITE STAGE TO EGRESS 54
3.1.7 INVESTIGATION OF THE CAVITY 55
3.1.8 MAURER S CLEFTS DEVELOPMENT CAN BE DIVIDED INTO 3 PHASES 57 3.1.9
EXPLORING PROTEIN TRANSPORT TO THE MAURER S CLEFTS 59
3.2 ROLE OF THE TRANSMEMBRANE DOMAIN IN THE EXPORT OF REX2 64 3.2.1
PROPERTIES OF TMS IN PNEP EXPORT 65
3.2.2 EXPORT DOES NOT DEPEND ON TM LENGTH 66
3.2.3 SCRAMBLING THE TM SEQUENCE LEADS TO A REDUCED EXPORT 67
3.2.4 THE HYDROPHOBICITY PATTERN IS NOT RESPONSIBLE FOR EXPORT 68
IMAGE 3
TABLE OF CONTENTS
3.2.5 THE N-TERMINAL HALF OF THE REX2 TM APPEARS TO BE MORE IMPORTANT
FOR EXPORT THAN THE C-TERMINAL HALF 68
3.2.6 THE TM OF A CLASSICAL TAIL-ANCHORED PROTEIN DOES NOT PROMOTE
EXPORT ....69 3.2.7 SOLUBILITY PROFILES OF TM REPLACEMENTS CONSTRUCTS 70
3.2.8 QUANTIFICATION OF TM SUBSTITUTION PHENOTYPES 71
3.3 EXPORT REQUIREMENTS FOR PNEPS 73
3.3.1 TRAFFICKING OF PNEPS 73
3.3.2 A REPORTER TO TEST FOR SEQUENCES MEDIATING EXPORT 73
3.3.3 COMMON REGIONS IN PNEPS PROMOTE EXPORT 75
3.3.4 THE MATURE N-TERMINUS OF PEXEL PROTEINS CAN FUNCTIONALLY REPLACE
PNEP N-TERMINI 77
3.3.5 N-TERMINI OF NON-EXPORTED SECRETORY PROTEINS MIMICKING MATURE
PEXELN-TERMINI FAIL TO PROMOTE EXPORT 79
3.3.6 SEQUENCES OF MATURE N-TERMINI INVOLVED IN EXPORT 80
3.3.7 MATURE N-TERMINUS BASED DISCRIMINATION FOR EXPORT STILL OCCURS
AFTER PEXEL CLEAVAGE 84
3.3.8 SOLUBILITY SHIFT DURING EXPORT OF TM PROTEINS 86
3.3.9 REX2-GFPMDHFR NEEDS TO BE UNFOLDED IN ORDER TO BE EXPORTED 89
CHAPTER 4: DISCUSSION 91
4.1 MAJOR FINDINGS 92
4.1.1 VISUALIZATION OF PARASITE DEVELOPMENT 92
4.1.2 ESTABLISHING 4D IMAGING IN P. FALCIPARUM 92
4.1.3 PROTEIN EXPORT OF P. FALCIPARUM 99
4.1.4 PNEPS AND PEXEL PROTEINS: CONVERGING EXPORT PATHWAYS? 107
CHAPTERS: REFERENCES 109
ACKNOWLEDGEMENTS 124
APPENDIX I 125
APPENDIX IT 127
PUBLICATIONS 128
|
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| spelling | Grüring, Christof Verfasser aut Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) vorgelegt von Christof Grüring Berlin Mensch und Buch Verl. 2011 128 S. Ill., graph. Darst., Kt. txt rdacontent n rdamedia nc rdacarrier Biological science Hamburg, Univ., Diss., 2011 Proteintransport (DE-588)4273577-4 gnd rswk-swf Virulenzfaktor (DE-588)4354782-5 gnd rswk-swf Plasmodium falciparum (DE-588)4250521-5 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Plasmodium falciparum (DE-588)4250521-5 s Virulenzfaktor (DE-588)4354782-5 s Proteintransport (DE-588)4273577-4 s DE-604 DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=024652315&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Grüring, Christof Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) Proteintransport (DE-588)4273577-4 gnd Virulenzfaktor (DE-588)4354782-5 gnd Plasmodium falciparum (DE-588)4250521-5 gnd |
| subject_GND | (DE-588)4273577-4 (DE-588)4354782-5 (DE-588)4250521-5 (DE-588)4113937-9 |
| title | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) |
| title_auth | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) |
| title_exact_search | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) |
| title_full | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) vorgelegt von Christof Grüring |
| title_fullStr | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) vorgelegt von Christof Grüring |
| title_full_unstemmed | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) vorgelegt von Christof Grüring |
| title_short | Characterization and visualization of protein export in the human malaria parasite Plasmodium falciparum (Welch, 1897) |
| title_sort | characterization and visualization of protein export in the human malaria parasite plasmodium falciparum welch 1897 |
| topic | Proteintransport (DE-588)4273577-4 gnd Virulenzfaktor (DE-588)4354782-5 gnd Plasmodium falciparum (DE-588)4250521-5 gnd |
| topic_facet | Proteintransport Virulenzfaktor Plasmodium falciparum Hochschulschrift |
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