The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti:
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1. Verfasser: | |
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Format: | Abschlussarbeit Buch |
Sprache: | English |
Veröffentlicht: |
2012
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Schlagworte: | |
Online-Zugang: | Inhaltsverzeichnis |
Beschreibung: | III, 125 S. Ill., graph. Darst. 30 cm |
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Datensatz im Suchindex
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adam_text | IMAGE 1
TABLE O F CONTENTS
TABLE OF CONTENTS 1. INTRODUCTION 1
1.1. AEDES AEGYPTI AS A MODEL ORGANISM 1
1.2 THE ALIMENTARY CANAL O F ADULT AEDES AEGYPTI 2
1.3 THE MALPIGHIAN TUBULES O F AEDES AEGYPTI 3
1.4 DIURESIS AND DIURETIC HORMONES 4
1.4.1 THE CALCITONIN-LIKE DIURETIC HORMONE DH31 5
1.4.2 THE AEDES AEGYPTI LEUCOKININ-LIKE PEPTIDE HORMONES - AEDESKININS 6
1.4.3 THE CORTICOTROPIN-RELEASING FACTOR (CRF)-RELATED PEPTIDE DH 44 6
1.4.4 CAPA-PEPTIDES 7
1.5 MOLECULAR BASICS O F ION TRANSPORT IN MALPIGHIAN TUBULES O F AEDES
AEGYPTI 8
1.6 THE V-ATPASE 10
1.6.1 STRUCTURE AND FUNCTION 10
1.6.2 REGULATION 12
1.6.2.1 ISOFORMS 12
1.6.2.2 REVERSIBLE DISSOCIATION 14
1.6.2.3 REVERSIBLE INSERTION O F FULLY ASSEMBLED V-ATPASES 16
1.6.2.4 INTRINSIC REGULATORY MECHANISMS 17
1.6.2.5 V-ATPASE AND PH 18
1.6.2.6 THE INFLUENCE O F CA 2+ -DEPENDENT PATHWAYS 19
1.7 AIM O F THIS THESIS 19
2. MATERIAL AND METHODS 21
2.1 MATERIAL 21
2.1.1 CHEMICALS, ENZYMES AND REAGENTS 21
2.1.2 ANTIBODIES 22
2.1.3 ESCHERICHIA COLI STRAINS AND GROWTH MEDIA 23
2.1.4 PLASMIDS AND OLIGONUCLEOTIDES 23
2.2 AEDES AEGYPTI AND TISSUE ISOLATION 24
2.3 MOLECULAR BIOLOGICAL AND GENETIC METHODS 25
2.3.1 ISOLATION O F TOTAL RNA AND CDNA SYNTHESIS 25
2.3.2 IDENTIFICATION O F FULL-LENGTH MRNA O F THE AEDES AEGYPTI V-ATPASE
SUBUNIT C 26
2.3.3 MRNA EXPRESSION ANALYSIS 27
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TABLE O F CONTENTS
2.3.4 AGAROSE GEL ELECTROPHORESIS AND DNA GEL EXTRACTION 28
2.3.5 PLASMID CONSTRUCTION 28
2.3.6 PREPARATION O F PLASMID DNA 29
2.3.7 TRANSFORMATION O F DNA IN CHEMICAL COMPETENT E. COLI AND LONG TERM
STORAGE 29
2.3.8 EXPRESSION O F RECOMBINANT V-ATPASE SUBUNIT C 30
2.4 BIOCHEMICAL METHODS 30
2.4.1 ATPASE ASSAYS AND PHOTOMETRIC DETERMINATION O F INORGANIC
PHOSPHATE 30
2.4.2 DETERMINATION O F INTRACELLULAR CAMP AND CGMP CONTENT 32
2.4.3 PURIFICATION O F RECOMBINANT V-ATPASE SUBUNIT C 33
2.4.4 PROTEIN DETERMINATION 33
2.4.5 IN VITRO PHOSPHORYLATION O F RECOMBINANT V-ATPASE C SUBUNITS 33
2.4.6 SDS-PAGE, PHOS-TAG-PAGE AND PROTEIN STAINING 34
2.4.7 ELECTRO-BLOT, QUANTITATIVE SLOT BLOT, OVERLAY BLOT AND
IMMUNOSTAINING 34
2.4.8 ENRICHMENT O F PHOSPHORYLATED PROTEINS 36
2.4.9 PROTEIN PRECIPITATION WITH TRICHLORIC ACID 36
2.4.10 AFFINITY CHROMATOGRAPHY USING THE RECOMBINANT V-ATPASE C SUBUNIT
37
2.5 BIOPHYSICAL METHODS 37
2.5.1 ELECTROPHYSIOLOGY 37
2.5.2 FLUORESCENCE MICROSCOPY FOR IMMUNOCYTOCHEMISTRY 38
2.5.3 INTRACELLULAR PH MEASUREMENTS 39
2.5.4 CIRCULAR DICHROISM SPECTROSCOPY 40
2.5.5 MASS SPECTROSCOPY AFTER IN-GEL DIGESTION 41
2.6 OTHER METHODS 42
2.7 STATISTICS 42
3. RESULTS 43
3.1 V-ATPASE ACTIVITIES IN UNSTIMULATED MALPIGHIAN TUBULES 43
3.2 DIURETIC HORMONES INFLUENCE CYCLIC NUCLEOTIDE LEVELS AND AFFECT
MEMBRANE AND TRANSEPITHELIAL POTENTIALS 44
3.3 NATRIURETIC STIMULATION INCREASES V-ATPASE ACTIVITY 46
3.4 CYCLIC NUCLEOTIDE-INDUCED ACTIVATION O F ATP-CONSUMERS CORRELATES
WITH STIMULATION O F V-ATPASE ACTIVITY 48
3.5 TOWARDS THE REGULATORY MECHANISMS O F THE V-ATPASE IN MALPIGHIAN
TUBULES OF AEDES AEGYPTI..... 50
3.5.1 CYCLIC NUCLEOTIDE-INDUCED ASSEMBLY O F THE V-ATPASE 50
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TABLE O F CONTENTS
3.5.2 IS THERE A PHOSPHORYLATION O F V-ATPASE SUBUNITS IN MALPIGHIAN
TUBULES OF AEDES AEGYPTIL 52
3.5.3 THE V-ATPASE SUBUNIT C 57
3.5.3.1 CLONING O F SUBUNIT C 57
3.5.3.2 PHOSPHORYLATION O F THE RECOMBINANT SUBUNIT C BY PKA AND
DETERMINATION O F THE PHOSPHORYLATION SITE 59
3.5.3.3 BINDING PARTNERS O F SUBUNIT C 61
3.5.4 ROLE O F THE CYTOSKELETON 64
3.5.5 ROLE O F THE INTRACELLULAR PH 67
3.5.5.1 METABOLIC INHIBITORS DUPLICATE CYCLIC NUCLEOTIDE EFFECTS 67
3.5.5.2 INDICATION O F A PHOSPHORYLATION-INDEPENDENT MECHANISM 68
3.5.5.3 QUALITATIVE MEASUREMENT O F INTRACELLULAR PH IN PRINCIPAL CELLS
OF MALPIGHIAN TUBULES 70
3.5.6 TISSUE-SPECIFIC ISOFORM EXPRESSION O F V-ATPASE SUBUNITS 71
3.5.6.1 SUBUNIT A 72
3.5.6.2 SUBUNIT G 74
3.5.7 V-ATPASE ACTIVITY AND REGULATION IN MIDGUT AND HINDGUT O F ADULT
AEDES AEGYPTI 76
4. DISCUSSION 78
4.1 BASAL ATPASE ACTIVITY 78
4.2 ENERGIZATION O F NATRIURESIS 81
4.3 CA 2+ -INDUCED MECHANISMS 90
4.4 THE EFFECT O F CGMP 91
4.5 DO V-ATPASE ISOFORMS DETERMINE V-ATPASE REGULATION? 94
5. SUMMARY 98
6. REFERENCES 100
7. APPENDIX 119
7.1 ABBREVIATIONS 119
7.2 CURRICULUM VITAE 122
7.3 PUBLICATIONS 123
7.4 ERKLARUNG IIBER DIE EIGENSTANDIGKEIT DER ERBRACHTEN
WISSENSCHAFTLICHEN LEISTUNG 124
8. ACKNOWLEDGEMENTS 125
III
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any_adam_object | 1 |
author | Tiburcy, Felix 1981- |
author_GND | (DE-588)1032629193 |
author_facet | Tiburcy, Felix 1981- |
author_role | aut |
author_sort | Tiburcy, Felix 1981- |
author_variant | f t ft |
building | Verbundindex |
bvnumber | BV041167402 |
ctrlnum | (OCoLC)853279820 (DE-599)DNB1033875473 |
dewey-full | 610 570 |
dewey-hundreds | 600 - Technology (Applied sciences) 500 - Natural sciences and mathematics |
dewey-ones | 610 - Medicine and health 570 - Biology |
dewey-raw | 610 570 |
dewey-search | 610 570 |
dewey-sort | 3610 |
dewey-tens | 610 - Medicine and health 570 - Biology |
discipline | Biologie Medizin |
format | Thesis Book |
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indexdate | 2024-07-10T00:41:10Z |
institution | BVB |
language | English |
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physical | III, 125 S. Ill., graph. Darst. 30 cm |
publishDate | 2012 |
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spelling | Tiburcy, Felix 1981- Verfasser (DE-588)1032629193 aut The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti vorgelegt von Felix Tiburcy 2012 III, 125 S. Ill., graph. Darst. 30 cm txt rdacontent n rdamedia nc rdacarrier Osnabrück, Univ., Diss., 2012 (DE-588)4113937-9 Hochschulschrift gnd-content DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=026142684&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
spellingShingle | Tiburcy, Felix 1981- The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti |
subject_GND | (DE-588)4113937-9 |
title | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti |
title_auth | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti |
title_exact_search | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti |
title_full | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti vorgelegt von Felix Tiburcy |
title_fullStr | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti vorgelegt von Felix Tiburcy |
title_full_unstemmed | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti vorgelegt von Felix Tiburcy |
title_short | The plasma membrane V-ATPase in Malpighian tubules of Aedes aegypti |
title_sort | the plasma membrane v atpase in malpighian tubules of aedes aegypti |
topic_facet | Hochschulschrift |
url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=026142684&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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