Function of the trigger loop in distinct steps of the transcription cycle:
Gespeichert in:
| 1. Verfasser: | |
|---|---|
| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
2013
|
| Schlagworte: | |
| Online-Zugang: | Volltext https://nbn-resolving.org/urn:nbn:de:bvb:355-epub-291180 Inhaltsverzeichnis |
| Beschreibung: | III, 95 Bl. Ill., graph. Darst. |
Internformat
MARC
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| 035 | |a (DE-599)BVBBV041636647 | ||
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| 245 | 1 | 0 | |a Function of the trigger loop in distinct steps of the transcription cycle |c von Thomas Fouqueau |
| 264 | 1 | |c 2013 | |
| 300 | |a III, 95 Bl. |b Ill., graph. Darst. | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
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| 912 | |a ebook | ||
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Datensatz im Suchindex
| _version_ | 1804151856786571264 |
|---|---|
| adam_text | TABLE OF CONTENTS
TABLE OF CONTENTS
TABLES OF CONTENTS L-LLL
I) INTRODUCTION
A. DNA-DEPENDENT RNA POLYMERASE 2
B. STRUCTURE OF MULTISUBUNIT RNAPS 5
1. STALK (E/F SUBCOMPLEX) 6
2. CLAMP DOMAIN 6
3. SWITCH REGION 8
4. ACTIVE SITE
9
A. THE TRIGGER LOOP
9
B. THE BRIDGE HELIX
11
C. FURTHER ACTIVE SITE ELEMENTS
12
C. TRANSCRIPTION CYCLE 12
1. INITIATION OF TRANSCRIPTION
14
2. ELONGATION
18
A. NUCLEOTIDE ADDITION CYCLE
19
B. NUCLEOTIDE SELECTION
20
C. PROOFREADING
21
A. INTRINSIC RNA CLEAVAGE
22
3. FACTOR-STIMULATED RNA CLEAVAGE
23
D. PROCESSIVITY
26
3. TERMINATION
27
D. AIMS OF THIS
THESIS 28
II) MATERIALS
A. SUPPLIERS 29
1. CHEMICALS
29
2. ENZYMES AND OTHER PROTEINS
30
3. COLUMN CHROMATOGRAPHY
30
B. GENETIC MATERIALS 31
1. STRAINS
31
2. PLASMIDS
31
3. PRIMERS FOR MUTAGENESIS
31
I
HTTP://D-NB.INFO/104935849X
TABLE OF CONTENTS
4. PRIMERS FOR PROMOTER MUTAGENESIS AND OLIGONUCLEOTIDES
32
III) METHODS
A. CLONING 33
1. GEL PURIFICATION OF PRIMERS 33
2. SEQUENCE-SPECIFIC MUTAGENESIS OF PLASMID
33
3. LIGATION OF LINEAR PLASMID
34
4. TRANSFORMATION OF E. COLI
34
B. PROTEIN OVEREXPRESSION AND
PURIFICATION 34
1. PROTEIN OVEREXPRESSION
34
2. PURIFICATION OF RECOMBINANT P. FURIOSUS RNAP SUBUNIT
35
A. PURIFICATION OF THE SUBUNIT FROM INCLUSION BODIES
35
A. PURIFICATION OF A AND K SUBUNITS
35
P. PURIFICATION OF A SUBUNIT
35
B. PURIFICATION OF SOLUBLE SUBUNIT
36
3. PURIFICATION OF RECOMBINANT TFS
36
4. RECONSTITUTION OF RNAP FROM P. FUSIOSUS
36
C. DNA TEMPLATES PREPARATION 37
1. STANDARD PROMOTER-DEPENDENT TRANSCRIPTION TEMPLATES 37
2. PRE-OPENED TEMPLATES
37
3. KMNCU-FOOTPRINT TEMPLATE
37
4.
RADIOACTIVELY 5 END LABELED
EMSA
TEMPLATE
38
D. ASSAYS 38
1. IN VITRO PROMOTER DEPENDENT TRANSCRIPTION ASSAYS
38
2. BAND SHIFT ASSAYS (EMSA)
39
3. KMNCVFOOTPRINT ASSAYS
39
4. BEAD-BASED RNA EXTENSION AND TFS INDUCED CLEAVAGE ASSAYS
39
5. BEAD-BASED RNA INTRINSIC CLEAVAGE ASSAYS
40
6. DATA ANALYSIS
40
IV) RESULTS
A. RECOMBINANT TL MUTANT RNAPS 41
B. RECONSTITUTION OF TL
MUTANTS RNAPS AND BINDING
ON THE PROMOTER 42
C. FUNCTION OF THE
TL IN
TRANSCRIPTION INITIATION 43
D. TL FUNCTION IN CATALYSIS 46
II
TABLE OF CONTENTS
1. COMPLEMENT UTP ADDITION
46
2. COMPLEMENT ATP ADDITION
47
E. TL FUNCTION IN NTP SELECTION AND
TRANSCRIPTION FIDELITY 48
F. TL FUNCTION IN NTP OVER 2 DNTP DISCRIMINATION 50
G. TL IS
NOT REQUIRED FOR INTRINSIC RNA CLEAVAGE 51
H. TL IS
NOT REQUIRED FOR TFS-STIMULATED
RNA CLEAVAGE 54
I. THE TL FUNCTIONS
IN SUPPRESSING ABNORMAL
TRANSCRIPTION TERMINATION 56
V) DISCUSSION
A. THE ESSENTIAL
ROLE OF THE TL DURING
TRANSCRIPTION INITIATION 58
B. THE FUNCTION OF A L83 IN TRANSCRIPTION
FIDELITY 58
C. SUBSTRATE BINDING
AND CATALYSIS 59
D. DISCRIMINATION
AGAINST THE WRONG
NUCLEOTIDE 59
E. TL-DEPENDENT AND TL-INDEPENDENT
RNA PROOFREADING 60
F. IMPLICATIONS FOR THE MECHANISM OF TRANSCRIPTION TERMINATION 61
G. TL DYNAMICS
IN THE TRANSCRIPTION CYCLE 62
VI) BIBLIOGRAPHY 64
VII) APPENDIX 85
A. ABBREVIATIONS 85
B. SUPPLEMENTAL FIGURES 87
SUMMARY
93
ACKNOWLEDGEMENTS
94
ERKLARUNG
95
IN
|
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| discipline | Biologie |
| format | Thesis Book |
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| language | English |
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| spelling | Fouqueau, Thomas Verfasser aut Function of the trigger loop in distinct steps of the transcription cycle von Thomas Fouqueau 2013 III, 95 Bl. Ill., graph. Darst. txt rdacontent n rdamedia nc rdacarrier Regensburg, Univ., Diss., 2013 Aktives Zentrum (DE-588)4337771-3 gnd rswk-swf DNS-abhängige-RNS-Polymerasen (DE-588)4273322-4 gnd rswk-swf Pyrococcus furiosus (DE-588)4314680-6 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Pyrococcus furiosus (DE-588)4314680-6 s DNS-abhängige-RNS-Polymerasen (DE-588)4273322-4 s Aktives Zentrum (DE-588)4337771-3 s b DE-604 Erscheint auch als Online-Ausgabe urn:nbn:de:bvb:355-epub-291180 http://epub.uni-regensburg.de/29118/ Verlag kostenfrei Volltext https://nbn-resolving.org/urn:nbn:de:bvb:355-epub-291180 Resolving-System DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=027077526&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Fouqueau, Thomas Function of the trigger loop in distinct steps of the transcription cycle Aktives Zentrum (DE-588)4337771-3 gnd DNS-abhängige-RNS-Polymerasen (DE-588)4273322-4 gnd Pyrococcus furiosus (DE-588)4314680-6 gnd |
| subject_GND | (DE-588)4337771-3 (DE-588)4273322-4 (DE-588)4314680-6 (DE-588)4113937-9 |
| title | Function of the trigger loop in distinct steps of the transcription cycle |
| title_auth | Function of the trigger loop in distinct steps of the transcription cycle |
| title_exact_search | Function of the trigger loop in distinct steps of the transcription cycle |
| title_full | Function of the trigger loop in distinct steps of the transcription cycle von Thomas Fouqueau |
| title_fullStr | Function of the trigger loop in distinct steps of the transcription cycle von Thomas Fouqueau |
| title_full_unstemmed | Function of the trigger loop in distinct steps of the transcription cycle von Thomas Fouqueau |
| title_short | Function of the trigger loop in distinct steps of the transcription cycle |
| title_sort | function of the trigger loop in distinct steps of the transcription cycle |
| topic | Aktives Zentrum (DE-588)4337771-3 gnd DNS-abhängige-RNS-Polymerasen (DE-588)4273322-4 gnd Pyrococcus furiosus (DE-588)4314680-6 gnd |
| topic_facet | Aktives Zentrum DNS-abhängige-RNS-Polymerasen Pyrococcus furiosus Hochschulschrift |
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