Structure-function analysis of the global transcriptional regulator RcsB:
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| Format: | Thesis Book |
| Language: | English |
| Published: |
2004
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| Subjects: | |
| Online Access: | Inhaltsverzeichnis |
| Physical Description: | 119 Bl. Ill., graph. Darst. 30 cm |
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MARC
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| 100 | 1 | |a Sengupta, Kaushik |d 1974- |e Verfasser |0 (DE-588)129082945 |4 aut | |
| 245 | 1 | 0 | |a Structure-function analysis of the global transcriptional regulator RcsB |c vorgelegt von Kaushik Sengupta |
| 264 | 1 | |c 2004 | |
| 300 | |a 119 Bl. |b Ill., graph. Darst. |c 30 cm | ||
| 336 | |b txt |2 rdacontent | ||
| 337 | |b n |2 rdamedia | ||
| 338 | |b nc |2 rdacarrier | ||
| 502 | |a Frankfurt (Main), Univ., Diss., 2004 (Nicht für den Austausch) | ||
| 650 | 0 | 7 | |a Genregulation |0 (DE-588)4122166-7 |2 gnd |9 rswk-swf |
| 650 | 0 | 7 | |a Struktur-Aktivitäts-Beziehung |0 (DE-588)4183784-8 |2 gnd |9 rswk-swf |
| 655 | 7 | |0 (DE-588)4113937-9 |a Hochschulschrift |2 gnd-content | |
| 689 | 0 | 0 | |a Genregulation |0 (DE-588)4122166-7 |D s |
| 689 | 0 | 1 | |a Struktur-Aktivitäts-Beziehung |0 (DE-588)4183784-8 |D s |
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| 943 | 1 | |a oai:aleph.bib-bvb.de:BVB01-013007778 | |
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| _version_ | 1807682828314345472 |
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| adam_text |
CONTENTS
ABSTRACT
1
1
INTRODUCTION
2
1.1
THE
RCSB
REGULON
2
1.2
CHARACTERIZATION
OF
THE
RCSAB
BOX
AND
THE
RCSB
BOX
8
1.3
RCSB
PHOSPHORYLATION
BY
RCSC-YOJN-RCSB
PHOSPHORELAY
PATHWAY
10
1.4
AN
OVERVIEW
OF
THE
RCSB
PROTEIN
AND
ITS
HOMOLOGUES
11
1.5
AIM
OF
THIS
WORK
14
2
MATERIALS
16
2.1
STRAINS
AND
PLASMIDS
16
2.2
DNA
OLIGONUCLEOTIDES
16
2.3
ENZYMES
AND
THEIR
BUFFERS
18
2.4
CHEMICALS
18
2.5
STANDARDS
20
2.6
KITS
21
2.7
MEDIA
FOR
THE
GROWTH
OF
ESCHERICHIA
COLI
21
2.8
SOLUTIONS
AND
BUFFERS
USED
IN
PROTEIN
PURIFICATION
AND
ANALYSIS
23
2.9
SOLUTIONS
AND
BUFFERS
USED
IN
AGAROSE
GEL
AND
POLYACRYLAMIDE
GEL
ELECTROPHORESIS
24
2.10
BUFFERS
FOR
SPR
AND
OTHER
BIOCHEMICAL
TECHNIQUES
27
2.11
BUFFERS
FOR
PREPARING
COMPETENT
CELLS
27
2.12
INSTRUMENTS
27
2.13
CHROMATOGRAPHIC
COLUMNS
AND
MATERIALS
29
2.14
OTHER
MATERIALS
29
3
METHODS
31
3.1
ISOLATION,
AMPLIFICATION,
ANALYSIS
AND
RECOMBINATION
OF
DNA
31
3.1.1
ISOLATION
OF
DNA
31
3.1.1.1
ISOLATION
OF
PLASMID
DNA
ON
ANALYTICAL
SCALE
31
3.1.1.2
ISOLATION
OF
PLASMID
DNA
ON
PREPARATIVE
SCALE
31
3.1.2
AMPLIFICATION
OF
DNA
FRAGMENTS
BY
POLYMERASE
CHAIN
REACTION
(PCR)
31
3.1.2.1
PURIFICATION
OF
PCR
PRODUCTS
32
3.1.3
ANALYSIS
OF
DNA
FRAGMENTS
BY
AGAROSE
GEL
ELECTROPHORESIS
32
3.1.4
RECOMBINATION
OF
DNA
33
3.1.4.1
RESTRICTION
DIGESTION
33
3.1.4.2
LIGATION
33
3.1.4.3
DNA-DNA
HYBRIDIZATION
34
3.2
TRANSFORMATION
OF
RECOMBINANT
DNA
IN
ESCHERICHIA
COLI
34
3.2.1
PREPARATION
OF
COMPETENT
CELLS
34
3.2.2
TRANSFORMATION
PROTOCOL
34
3.3
SEQUENCING
OF
DNA
35
3.4
EXPRESSION,
ISOLATION,
PURIFICATION
AND
ANALYSIS
OF
PROTEINS
36
3.4.1
EXPRESSION
OF
PROTEINS
36
3.4.1.1
EXPRESSION
OF
PROTEINS
ON
ANALYTICAL
SCALE
36
3.4.1.2
EXPRESSION
OF
PROTEINS
ON
PREPARATIVE
SCALE
36
3.4.1.3
EXPRESSION
OF
LABELLED
PROTEINS
ON
PREPARATIVE
SCALE
37
3.4.2
ISOLATION
OF
PROTEINS
37
3.4.2.1
CELL
DISRUPTION
BY
FRENCH
PRESS
37
3.4.2.2
CELL
DISRUPTION
BY
SONICATION
38
3.4.3
PURIFICATION
OF
PROTEINS
BY
AFFINITY
CHROMATOGRAPHY
38
3.4.3.1
HEPARIN
CHROMATOGRAPHY
38
3.4.3.2
NI-NTA
CHROMATOGRAPHY
38
3.4.3.3
DEXTRIN
CHROMATOGRAPHY
39
3.4.4
CONCENTRATION
OF
PROTEINS
39
3.4.5
DIALYSIS
OF
PROTEINS
40
3.4.6
SUBLIMATION
40
3.4.7
ANALYSIS
OF
PROTEINS
BY
SDS-POLYACRYLAMIDE
GEL
ELECTROPHORESIS
40
3.4.7.1
METHOD
OF
CASTING
AND
RUNNING
SDS-GELS
41
3.4.7.2
SAMPLE
PREPARATION
41
3.4.7.3
STAINING
AND
DESTAINING
OF
SDS-GELS
42
3.5
CELL-FREE
PRODUCTION
OF
LABELLED
PROTEINS
42
3.6
SPECTROSCOPIC
TECHNIQUES
43
3.6.1
NUCLEAR
MAGNETIC
RESONANCE
(NMR)
SPECTROSCOPY
43
3.6.1.1
NMR
DATA
COLLECTION
AND
PROCESSING
45
3.6.2
FLUORESCENCE
SPECTROSCOPY
46
3.6.2.1
DATA
COLLECTION
FOR
PROTEIN-PROTEIN
INTERACTION
47
3.6.3
CIRCULAR
DICHROISM
(CD)
SPECTROSCOPY
48
3.6.3.1
RECORDING
CD
SPECTRA
49
3.7
CHEMICAL
CROSS-LINKING
OF
PROTEINS
49
3.8
ELECTROPHORETIC
MOBILITY
SHIFT
ASSAY
(EMSA)
51
3.8.1
GENERATION
OF
TARGET
DNA
51
3.8.2
METHOD
OF
RADIOLABELLING
DNA
51
3.8.3
PREPARATION
OF
SAMPLES
FOR
EMSA
52
3.8.4
CASTING
AND
RUNNING
NATIVE
GELS
52
3.8.5
AUTORADIOGRAPHY
53
3.9
SURFACE
PLASMON
RESONANCE
(SPR)
53
3.9.1
GENERATION
OF
TARGET
DNA
54
3.9.2
IMMOBILIZATION
OF
TARGET
DNA
ON
THE
SA
CHIP
54
3.9.3
METHOD
OF
MONITORING
PROTEIN-DNA
INTERACTION
54
4
RESULTS
55
4.1
FERMENTATION
AND
PURIFICATION
OF
PROTEINS
55
4.1.1
THE
C-TERMINAL
RCSB
DOMAIN
(C-RCSB)
OF
ERWINIA
AMYLOVORA
55
4.1.2
C-RCSB
OF
ERWINIA
AMYLOVORA
WITH
A
D-PROTEIN
FUSION
TAG
(D-C-RCSB)
56
4.1.3
THE
N-TERMINAL
DOMAIN
OF
RCSB
58
4.1.4
RCSB
OF
ERWINIA
AMYLOVORA
AND
ESCHERICHIA
COLI
AND
R
CS
B
DUA
OF
ERWINIA
AMYLOVORA
60
4.1.5
RCSA
OF
ERWINIA
AMYLOVORA
WITH
MBP
AS
A
SOLUBILITY
ENHANCEMENT
TAG
(SET)
61
4.2
SOLUTION
STRUCTURE
OF
C-RCSB
OF
ERWINIA
AMYLOVORA
62
4.3
INTERDOMAIN
INTERACTIONS
OF
RCSB
64
4.3.1
SPECTRAL
PROPERTIES
OF
DIFFERENT
TRYPTOPHAN
ANALOGUES
64
4.3.2
DNA-BINDING
ANALYSIS
OF
DIFFERENT
TRYPTOPHAN
ANALOGUES
67
4.3.3
N-RCSB/RCSB,
C-RCSB/RCSB
AND
N-RCSB/C-RCSB
INTERACTION
ANALYSIS
68
4.3.3.1
N-RCSB/RCSB
AND
C-RCSB/RCSB
INTERACTION
ANALYSIS
BY
FLUORESCENCE
SPECTROSCOPY
68
4.3.3.2
N-RCSB/
RCSB
INTERACTION
ANALYSIS
BY
CHEMICAL
CROSS-LINKING
68
4.3.3.3
C-RCSB/RCSB
INTERACTION
ANALYSIS
BY
CHEMICAL
CROSS-LINKING
69
4.3.3.4
C-RCSB/N-RCSB
INTERACTION
ANALYSIS
BY
CHEMICAL
CROSS-LINKING
70
4.3.3.5
C-RCSB/N-RCSB
INTERACTION
ANALYSIS
BY
NMR
SPECTROSCOPY
71
4.4
ANALYSIS
OF
THE
DNA-BINDING
ACTIVITY
OF
C-RCSB
74
4.4.1
DNA-BINDING
ACTIVITY
ANALYSIS
BY
NMR
SPECTROSCOPY
74
4.4.2
DNA-BINDING
STUDY
BY
CD
SPECTROSCOPY
77
4.4.3
C-RCSB/DNA
COMPLEX
ANALYSIS
BY
EMSA
78
4.4.4
KINETICS
OF
C-RCSB/DNA
COMPLEX
FORMATION
BY
SPR
82
4.5
DNA-BINDING
ACTIVITY
OF
RCSB
IN
PRESENCE
OF
LOW
MOLECULAR
WEIGHT
PHOSPHORYLATION
MIMICS
85
4.5.1
DNA-BINDING
ACTIVITY
OF
MODIFIED
RCSB
BY
EMSA
85
4.5.2
DNA-BINDING
KINETICS
OF
MODIFIED
RCSB
BY
SPR
88
5
DISCUSSION
92
5.1
SOLUTION
STRUCTURE
OF
THE
C-TERMINAL
DOMAIN
OF
RCSB
PROTEIN
92
5.2
INTERACTION
OF
THE
C-RCSB
PROTEIN
WITH
THE
RCSAB
BOX
93
5.3
INTERDOMAIN
INTERACTIONS
IN
RCSB
PROTEIN
96
5.4
DNA-BINDING
PROPERTY
OF
MODIFIED
RCSB
PROTEIN
98
LIST
OF
ABBREVIATIONS
101
REFERENCES
102
ZUSAMMENFASSUNG
114
LEBENSLAUF
119 |
| any_adam_object | 1 |
| author | Sengupta, Kaushik 1974- |
| author_GND | (DE-588)129082945 |
| author_facet | Sengupta, Kaushik 1974- |
| author_role | aut |
| author_sort | Sengupta, Kaushik 1974- |
| author_variant | k s ks |
| building | Verbundindex |
| bvnumber | BV019679779 |
| ctrlnum | (OCoLC)615548081 (DE-599)BVBBV019679779 |
| discipline | Biologie |
| format | Thesis Book |
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| genre | (DE-588)4113937-9 Hochschulschrift gnd-content |
| genre_facet | Hochschulschrift |
| id | DE-604.BV019679779 |
| illustrated | Illustrated |
| indexdate | 2024-08-18T00:24:43Z |
| institution | BVB |
| language | English |
| oai_aleph_id | oai:aleph.bib-bvb.de:BVB01-013007778 |
| oclc_num | 615548081 |
| open_access_boolean | |
| owner | DE-355 DE-BY-UBR |
| owner_facet | DE-355 DE-BY-UBR |
| physical | 119 Bl. Ill., graph. Darst. 30 cm |
| publishDate | 2004 |
| publishDateSearch | 2004 |
| publishDateSort | 2004 |
| record_format | marc |
| spelling | Sengupta, Kaushik 1974- Verfasser (DE-588)129082945 aut Structure-function analysis of the global transcriptional regulator RcsB vorgelegt von Kaushik Sengupta 2004 119 Bl. Ill., graph. Darst. 30 cm txt rdacontent n rdamedia nc rdacarrier Frankfurt (Main), Univ., Diss., 2004 (Nicht für den Austausch) Genregulation (DE-588)4122166-7 gnd rswk-swf Struktur-Aktivitäts-Beziehung (DE-588)4183784-8 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Genregulation (DE-588)4122166-7 s Struktur-Aktivitäts-Beziehung (DE-588)4183784-8 s DE-604 DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=013007778&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Sengupta, Kaushik 1974- Structure-function analysis of the global transcriptional regulator RcsB Genregulation (DE-588)4122166-7 gnd Struktur-Aktivitäts-Beziehung (DE-588)4183784-8 gnd |
| subject_GND | (DE-588)4122166-7 (DE-588)4183784-8 (DE-588)4113937-9 |
| title | Structure-function analysis of the global transcriptional regulator RcsB |
| title_auth | Structure-function analysis of the global transcriptional regulator RcsB |
| title_exact_search | Structure-function analysis of the global transcriptional regulator RcsB |
| title_full | Structure-function analysis of the global transcriptional regulator RcsB vorgelegt von Kaushik Sengupta |
| title_fullStr | Structure-function analysis of the global transcriptional regulator RcsB vorgelegt von Kaushik Sengupta |
| title_full_unstemmed | Structure-function analysis of the global transcriptional regulator RcsB vorgelegt von Kaushik Sengupta |
| title_short | Structure-function analysis of the global transcriptional regulator RcsB |
| title_sort | structure function analysis of the global transcriptional regulator rcsb |
| topic | Genregulation (DE-588)4122166-7 gnd Struktur-Aktivitäts-Beziehung (DE-588)4183784-8 gnd |
| topic_facet | Genregulation Struktur-Aktivitäts-Beziehung Hochschulschrift |
| url | http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=013007778&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA |
| work_keys_str_mv | AT senguptakaushik structurefunctionanalysisoftheglobaltranscriptionalregulatorrcsb |