Structural studies on the association of filamentous proteins in the human M-Bands:
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| Format: | Thesis Book |
| Language: | English |
| Published: |
2011
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| Online Access: | Volltext Inhaltsverzeichnis |
| Item Description: | Zsfassung in dt. und engl. Sprache |
| Physical Description: | 137, XVI S. Ill., graph. Darst. |
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| 100 | 1 | |a Sauer, Florian |d 1982- |e Verfasser |0 (DE-588)1026248485 |4 aut | |
| 245 | 1 | 0 | |a Structural studies on the association of filamentous proteins in the human M-Bands |c von Florian Sauer |
| 246 | 1 | 3 | |a Strukturelle Studien zur Zusammenlagerung filamentöser Protein in humanen M-Banden |
| 264 | 1 | |c 2011 | |
| 300 | |a 137, XVI S. |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
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| 500 | |a Zsfassung in dt. und engl. Sprache | ||
| 502 | |a Würzburg, Univ., Diss., 2012 | ||
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Record in the Search Index
| _version_ | 1804149495802363904 |
|---|---|
| adam_text | IMAGE 1
TABLE O F CONTENTS
TABLE OF CONTENTS
ABSTRACT I
ZUSAMMERIFASSUNG ILL
TABLE OF CONTENTS V
LIST OF FIGURES VII
LIST OF TABLES VIII
ABBREVIATIONS IX
1 INTRODUCTION 1
1.1 VERTEBRATE STRIATED MUSCLE 1
1.1.1 GENERAL ARCHITECTURE 1
1.1.2 THE SARCOMERE 2
1.1.3 SARCOMERE CONTRACTION: T H E SLIDING F I L A M E N T M O D E L 5
1.2 THE M-BAND: A CROSSLINK FOR THE THICK FILAMENTS 5
1.2.1 COMPONENTS O F T H E SARCOMERIC CYTOSKELETON ASSOCIATE IN T H E M
- B A N D 8
1.2.1.1 TITIN: THE THIRD FILAMENT 8
1.2.1.1.1 TITIN CONNECTS Z-DISKS TO THE M-BANDS 9
1.2.1.1.2 THE CENTRAL PORTION O F TITIN ACTS AS SARCOMERIC RULER AND
SPRING 13
1.2.1.2 MEMBERS O F THE MYOMESIN FAMILY: THICK FILAMENT CROSS-LINKERS 15
1.2.1.3 OBSCURIN AND OBSL1 18
1.2.1.3.1 OBSCURIN AND OBSL1: NOVEL SARCOMERIC FILAMENTS 18
1.2.1.3.2 OBSCURIN AND OBSL1 ASSEMBLE WITH TITIN AND MYOMESIN-1 IN THE
M-BANDS 19
1.2.2 ALTERATIONS O F M - B A N D PROTEINS ARE INVOLVED IN T H E D E V E
L O P M E N T O F HEREDITARY
DISEASES 2 1
1.2.3 A C U R R E N T MODEL O F T H E VERTEBRATE M - B A N D 22
1.3 AIM OF THE WORK 24
2 MATERIALS AND METHODS 25
2.1 MATERIALS 25
2.1.1 CHEMICALS, KITS AND CONSUMABLES 25
2.1.2 CULTURE MEDIA 2 6
2.1.3 ENZYMES, PRIMERS, VECTORS AND EXPRESSION STRAINS 2 6
2.1.4 BUFFERS F O R PROTEIN PURIFICATION AND CHARACTERIZATION 27
2.1.5 MACHINES AND COLUMNS 2 8
2.1.6 SOFTWARE AND DATABASES 2 9
2.2 METHODS 30
2.2.1 CLONING O F EXPRESSION CONSTRUCTS AND SITE DIRECTED MUTAGENESIS 3
0
2.2.2 PROTEIN EXPRESSION IN E. COLI 32
2.2.3 PROTEIN PURIFICATION 33
2.2.3.1 IMAC AFFINITY PURIFICATION OF POLY-HISTIDINE TAGGED PROTEINS 33
2.2.3.2 PURIFICATION BY SIZE-EXCLUSION CHROMATOGRAPHY 34
2.2.4 PROTEIN AND PROTEIN COMPLEX CHARACTERIZATION 3 4
2.2.4.1 ESTIMATION O F PROTEIN CONCENTRATIONS 34
2.2.4.2 ESTIMATION OF PROTEIN WEIGHT AND PURITY BY SDS-PAGE 35
2.2.4.3 ESTIMATION OF SECONDARY STRUCTURE CONTENT BY CIRCULAR DICHROISM
35
2.2.4.4 COMPLEX CHARACTERIZATION BY ITC 36
2.2.4.5 X-RAY CRYSTALLOGRAPHY 37
2.2.4.5.1 PRINCIPLES 37
2.2.4.5.2 PHASE DETERMINATION 39
2.2.4.5.3 ESTIMATION O F DIFFRACTION DATA QUALITY 43
- V -
HTTP://D-NB.INFO/1026910498
IMAGE 2
TABLE O F CONTENTS
2.2.4.5.4 PROTEIN CRYSTALLIZATION 44
2.2.4.5.5 DATA COLLECTION, STRUCTURE SOLUTION AND REFINEMENT 46
2.2.4.5.6 STRUCTURE ANALYSIS AND VISUALIZATION 47
3 RESULTS AND DISCUSSION 4 8
3.1 CHARACTERIZATION OF THE HUMAN TITIN - OBSL1 / OBSCURIN HEAD - TO -
TAIL COMPLEX ... 49 3.1.1 RESULTS 4 9
3.1.1.1 TITIN M10 AND OBSCURIN/OBSLL I G L FORM A BINARY COMPLEX 49
3.1.1.2 CRYSTALLIZATION 50
3.1.1.3 DATA COLLECTION AND STRUCTURE SOLUTION 5 1
3.1.1.4 OVERALL STRUCTURE 53
3.1.1.5 COMPLEX FORMATION BY INTERMOLECULAR 15-SHEET FORMATION 54
3.1.1.6 CHARACTERIZATION O F THE INTERFACE AND DISEASE RELATED MUTATIONS
57
3.1.1.6.1 HOMOLOGY BETWEEN OBSL1 AND OBSCURIN 57
3.1.1.6.2 CALORIMETRIC CHARACTERIZATION 59
3.1.1.6.3 CHARACTERIZATION O F DISEASE RELATED MUTATIONS 59
3.1.2 DISCUSSION 65
3.1.2.1 A NOVEL VARIATION O F A KNOWN ASSEMBLY MODE 65
3.1.2.2 A SAFEGUARD FOR TITIN KINASE FUNCTION? 67
3.1.2.3 TWO HYPOTHETICAL MODELS FOR THE ORIGIN O F M10 LINKED MYOPATHIES
70
3.2 CHARACTERIZATION OF THE DIRECT ASSEMBLY BETWEEN TITIN AND MYOMESIN-1
74
3.2.1 RESULTS 7 4
3.2.1.1 PURIFICATION O F THE COMPLEX PARTNERS 74
3.2.1.2 IN VITRO TITIN DOES NOT SPONTANEOUSLY FORM A COMPLEX WITH
MYOMESIN-1 76
3.2.2 DISCUSSION 7 8
3.3 CHARACTERIZATION OF THE CENTRAL ASSEMBLY SITE OF MYOMESIN-1 80
3.3.1 RESULTS 8 0
3.3.1.1 MYOMESIN-1 DIMERIZATION IS MEDIATED BY DOMAIN 5 80
3.3.1.2 CRYSTALLIZATION OF THE MYOMESIN-1 MY5 DOMAIN 84
3.3.1.3 DATA COLLECTION, STRUCTURE SOLUTION AND REFINEMENT 86
3.3.1.4 CRYSTAL STRUCTURE O F THE MY5 DIMER 88
3.3.2 DISCUSSION 9 2
3.3.2.1 DIMERIZATION BY THREE DIMENSIONAL DOMAIN SWAPPING 92
3.3.2.2 BIOLOGICAL IMPLICATIONS: A HYPOTHETICAL MODEL FOR THE
INCORPORATION O F TETRAMERIC MYOMESIN-1 IN THE M-BANDS 95
3.4 CRYSTAL STRUCTURE OF TITIN M 4 100
3.4.1 RESULTS 100
3.4.1.1 CRYSTALLIZATION 100
3.4.1.2 DATA COLLECTION, STRUCTURE SOLUTION AND MODEL REFINEMENT 101
3.4.1.3 OVERALL STRUCTURE 106
3.4.1.4 DIMERIZATION O F M4 109
3.4.1.5 THE POSITION O F CYS22 IS UNIQUE WITHIN TITIN IG-LIKE DOMAINS
AND CONSERVED AMONG VERTEBRATES I L L
3.4.2 DISCUSSION 117
3.4.2.1 THE M4 ASSEMBLY MAY REPRESENT A TEMPORARY STABLE ANCHOR O F
TITIN IN THE M-BANDS 117
4 CONCLUSIONS AND FUTURE PERSPECTIVES 122
5 LITERATURE 125
APPENDIX XI
PUBLICATIONS XII
PRESENTATIONS AND PARTICIPATIONS XII
CONTRIBUTIONS XIII
ACKNOWLEDGEMENTS XIV
CURRICULUM VITAE XV
ERKLARUNG XVI
- V I -
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| spelling | Sauer, Florian 1982- Verfasser (DE-588)1026248485 aut Structural studies on the association of filamentous proteins in the human M-Bands von Florian Sauer Strukturelle Studien zur Zusammenlagerung filamentöser Protein in humanen M-Banden 2011 137, XVI S. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Zsfassung in dt. und engl. Sprache Würzburg, Univ., Diss., 2012 Muskelkontraktion (DE-588)4170858-1 gnd rswk-swf Quer gestreifte Muskulatur (DE-588)4176716-0 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Muskelkontraktion (DE-588)4170858-1 s Quer gestreifte Muskulatur (DE-588)4176716-0 s DE-604 Erscheint auch als Online-Ausgabe urn:nbn:de:bvb:20-opus-72410 https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-72410 Resolving-System kostenfrei Volltext DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=025287838&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Sauer, Florian 1982- Structural studies on the association of filamentous proteins in the human M-Bands Muskelkontraktion (DE-588)4170858-1 gnd Quer gestreifte Muskulatur (DE-588)4176716-0 gnd |
| subject_GND | (DE-588)4170858-1 (DE-588)4176716-0 (DE-588)4113937-9 |
| title | Structural studies on the association of filamentous proteins in the human M-Bands |
| title_alt | Strukturelle Studien zur Zusammenlagerung filamentöser Protein in humanen M-Banden |
| title_auth | Structural studies on the association of filamentous proteins in the human M-Bands |
| title_exact_search | Structural studies on the association of filamentous proteins in the human M-Bands |
| title_full | Structural studies on the association of filamentous proteins in the human M-Bands von Florian Sauer |
| title_fullStr | Structural studies on the association of filamentous proteins in the human M-Bands von Florian Sauer |
| title_full_unstemmed | Structural studies on the association of filamentous proteins in the human M-Bands von Florian Sauer |
| title_short | Structural studies on the association of filamentous proteins in the human M-Bands |
| title_sort | structural studies on the association of filamentous proteins in the human m bands |
| topic | Muskelkontraktion (DE-588)4170858-1 gnd Quer gestreifte Muskulatur (DE-588)4176716-0 gnd |
| topic_facet | Muskelkontraktion Quer gestreifte Muskulatur Hochschulschrift |
| url | https://nbn-resolving.org/urn:nbn:de:bvb:20-opus-72410 http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=025287838&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
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