Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators: = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren
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| Format: | Abschlussarbeit Buch |
| Sprache: | English |
| Veröffentlicht: |
Erlangen ; Nürnberg
2016
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| Schlagworte: | |
| Online-Zugang: | Inhaltsverzeichnis |
| Beschreibung: | 200 Seiten Illustrationen, Diagramme |
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| 245 | 1 | 0 | |a Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators |b = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |c vorgelegt von Regine Brox aus Nürnberg |
| 246 | 1 | 1 | |a Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |
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Datensatz im Suchindex
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| adam_text | MOLECULAR MECHANISMS OF FUNCTIONALLY SELECTIVE AND
PROBE-DEPENDENT SIGNALING AT CXCR3 INDUCED BY NEGATIVE ALLOSTERIC
MODULATORS
MOLEKULARE MECHANISMEN DER FUNKTIONELL SELEKTIVEN UND
PROBENABHAENGIGEN SIGNALTRANSDUKTION AM CXCR3 REZEPTOR
VERANLASST DURCH NEGATIVE ALLOSTERISCHE MODULATOREN
DER NATURWISSENSCHAFTLICHEN FAKULTAET
DER FRIEDRICH-ALEXANDER-UNIVERSITAET
ERLANGEN-NUERNBERG
ZUR
ERLANGUNG DES DOKTORGRADES DR. RER. NAT.
VORGELEGT VON
REGINE BROX
AUS NUERNBERG
TABLE OF CONTENTS
1
INTRODUCTION......................................................................................................................................
1
1.1 CHEMOKINES, A FAMILY OF CHEMOTACTIC
CYTOKINES...............................................................1
1.2 CHEMOKINE
RECEPTORS.........................................................................................................3
1.2.1 STRUCTURE AND
FUNCTION...................................................................................................3
1.2.2 CHEMOKINE-RECEPTOR
INTERACTIONS.................................................................................
5
1.3 THE CHEMOKINE RECEPTOR
CXCR3........................................................................................
7
1.3.1 STRUCTURE OF
CXCR3.........................................................................................................8
1.3.2 ENDOGENOUS LIGANDS AND SIGNALING NETWORK OF CXCR3
.................................................
9
1.3.3 CXCR3 AS DRUG
TARGET....................................................................................................11
1.4 ALLOSTERY AND CHEMOKINE
RECEPTORS................................................................................
12
1.4.1 ALLOSTERIC MODULATION OF GPCRS
..................................................................................
12
1.4.2 DETECTION AND QUANTIFICATION OF ALLOSTERIC MODULATORS
..............................................
14
1.4.3 THERAPEUTIC BENEFITS OF ALLOSTERIC
MODULATION............................................................18
1.4.4 THE IMPACT OF FUNCTIONAL SELECTIVITY AND ALLOSTERISM ON CHEMOKINE
RECEPTORS
.......
23
2 OBJECTIVES OF THE
THESIS..................................................................................................................27
3 FUNCTIONAL SELECTIVE NEGATIVE ALLOSTERIC MODULATORS OF
CXCR3..............................................30
3.1 DEVELOPMENT OF SMALL MOLECULE ANTAGONISTS OF CXCR3
.................................................
30
3.2 BORONIC ACIDS AS NOVEL ALLOSTERIC MODULATORS OF CXCR3
.................................................
31
3.3 LIGAND-BIASED AND PROBE DEPENDENT MODULATION OF CXCR3
..........................................
34
3.3.1 RECEPTOR
BINDING..........................................................................................................36
3.3.2 FUNCTIONAL CHARACTERIZATION BY [35S]GTPYS INCORPORATION ASSAY AND
SS-ARRESTIN-2
RECRUITMENT
ASSAY.......................................................................................................37
3.3.3
CONCLUSION....................................................................................................................
40
4 FUNCTIONAL CHARACTERIZATION OF NEGATIVE ALLOSTERIC MODULATORS BY
BRET-BASED METHODS.41
4.1 BIOLUMINESCENCE RESONANCE ENERGY
TRANSFER..................................................................41
4.2 BRET BASED CAMP
ASSAY...................................................................................................42
4.3 BRET-BASED SS-ARRESTIN
ASSAY............................................................................................43
4.4 FUNCTIONAL CHARACTERIZATION OF NEGATIVE ALLOSTERIC MODULATORS
.....................................
47
4.5
CONCLUSION.........................................................................................................................53
5 MOLECULAR MECHANISMS OF BIASED AND PROBE DEPENDENT SIGNALING AT CXCR3
INDUCED BY
NEGATIVE ALLOSTERIC
MODULATORS..........................................................................................................55
5.1 BINDING MODE OF NEGATIVE ALLOSTERIC MODULATORS USING SITE-DIRECTED
MUTAGENESIS
....
58
5.1.1 CELL SURFACE RECEPTOR EXPRESSION
.................................................................................
59
5.1.2 EFFECTS OF AMINO ACID SUBSTITUTIONS ON THE BINDING OF NEGATIVE
ALLOSTERIC
MODULATORS..................................................................................................................
60
5.1.3 INVESTIGATION OF THE BINDING MODE BY COMPUTATIONAL STUDY
......................................
64
5.1.4 PROPOSED BINDING MODE OF BD064
..............................................................................
64
5.1.5 PROPOSED BINDING MODE OF
BD103..............................................................................
66
5.1.6
CONCLUSION....................................................................................................................
68
5.2 EFFECTS OF MUTATIONS ON SIGNALING OF CHEMOKINES CXCL11 AND CXCL10
.........................
68
5.2.1 G PROTEIN
ACTIVATION.....................................................................................................69
5.2.2 THE RECRUITMENT OF SS-ARRESTIN 1 AND SS-ARRESTIN 2
........................................................71
5.2.3 DISCUSSION AND
CONCLUSION...........................................................................................73
5.3 EFFECTS OF MUTATIONS ON THE ABILITY OF ALLOSTERIC MODULATORS TO
INHIBIT CHEMOKINE-
MEDIATED
SIGNALING...........................................................................................................
76
5.3.1 INHIBITION OF G PROTEIN
ACTIVATION................................................................................
76
5.3.2 INHIBITION OF SS-ARRESTIN 1 AND SS-ARRESTIN 2
RECRUITMENT...............................................83
5.3.3 DISCUSSION AND
CONCLUSION...........................................................................................89
6 FUNCTIONAL SELECTIVE POSITIVE ALLOSTERIC MODULATORS OF CXCR3
...............................................
92
6.1 DEVELOPMENT OF SMALL MOLECULE AGONISTS OF CXCR3
92
6.2 NOVEL CLASS OF BIASED ALLOSTERIC AGONISTS OF
CXCR3........................................................... 93
6.2.1 G PROTEIN
ACTIVATION......................................................................................................95
6.2.2 THE RECRUITMENT OF SS-ARRESTIN 2
..................................................................................
96
6.2.3
INTERNALIZATION..............................................................................................................
98
6.2.4 RECEPTOR
BINDING..........................................................................................................99
6.2.5 CHEMOTAXIS.....................................................
102
6.2.6
CONCLUSION..................................................................................................................
103
7 FUTURE
DIRECTIONS.............................................................................................................................105
7.1 BREAST
CANCER..................................................................................................................
107
7.2 CXCR3 AND ITS LIGANDS IN BREAST
CANCER.........................................................................
107
7.3 SMALL-MOLECULE ANTAGONISTS SUPPRESS METASTASIS IN BREAST
CANCER.............................110
7.4 ALDEHYDE DEHYDROGENASE AS MARKER OF
BCSLC..............................................................114
7.5 WESTERN BLOT
ANALYSIS.....................................................................................................115
7.6
CONCLUSION......................................................................................................................
117
8
SUMMARY...........................................................................................................................................
119
9
ZUSAMMENFASSUNG..........................................................................................................................
126
10 EXPERIMENTAL
PART............................................................................................................................134
10.1
MATERIALS...........................................................................................................................134
LO.L.LTECHNICAL
INSTRUMENTS................................................................................................134
10.1.2TEST
COMPOUNDS.........................................................................................................135
10.1.3 CHEMICALS AND KITS FOR
MICROBIOLOGY.........................................................................
139
10.1.4 PLASTIC /SINGLE USE
MATERIALS......................................................................................
143
10.1.5 CELL
LINES......................................................................................................................
144
10.1.6
PLASMIDS......................................................................................................................
145
10.1.7 BUFFER
COMPOSITIONS...................................................................................................149
10.1.8 COMPUTER SOFTWARE
151
10.2
METHODS...........................................................................................................................
152
10.2.1 MOLECULAR
BIOLOGY......................................................................................................
152
10.2.2 MAMMALIAN CELL
CULTURE.............................................................................................156
10.2.3 RADIOLIGAND BINDING
STUDIES.......................................................................................158
10.2.4 FUNCTIONAL
ASSAYS.......................................................................................................
160
10.2.5 BIOLUMINESCENCE RESONANCE ENERGY
TRANSFER..........................................................162
10.2.6 ALDEFLUOR
ASSAY...........................................................................................................
165
10.2.7 WESTERN BLOT ANALYSIS............................................
166
10.2.8 CONCENTRATIONS USED FOR TERNARY COMPLEX
MODEL.....................................................167
11
APPENDIX...........................................................................................................................................170
11.1
ABBREVIATIONS..................................................................................................................170
11.2 LIST OF
FIGURES.................................................................................................................
173
11.3 LIST OF
TABLES...................................................................................................................
176
12
LITERATURE..........................................................................................................................................178
CURRICULUM VITAE/LEBENSLAUF,
.200
|
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| author | Brox, Regine 1988- |
| author_GND | (DE-588)1124800840 |
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| spelling | Brox, Regine 1988- (DE-588)1124800840 aut Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren vorgelegt von Regine Brox aus Nürnberg Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren Erlangen ; Nürnberg 2016 200 Seiten Illustrationen, Diagramme txt rdacontent n rdamedia nc rdacarrier Dissertation Friedrich-Alexander-Universität Erlangen-Nürnberg 2016 Ligand Biochemie (DE-588)4606532-5 gnd rswk-swf Allosterischer Effektor (DE-588)4291850-9 gnd rswk-swf Chemokine (DE-588)4429989-8 gnd rswk-swf (DE-588)4113937-9 Hochschulschrift gnd-content Chemokine (DE-588)4429989-8 s Ligand Biochemie (DE-588)4606532-5 s Allosterischer Effektor (DE-588)4291850-9 s DE-604 DNB Datenaustausch application/pdf http://bvbr.bib-bvb.de:8991/F?func=service&doc_library=BVB01&local_base=BVB01&doc_number=029437433&sequence=000001&line_number=0001&func_code=DB_RECORDS&service_type=MEDIA Inhaltsverzeichnis |
| spellingShingle | Brox, Regine 1988- Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren Ligand Biochemie (DE-588)4606532-5 gnd Allosterischer Effektor (DE-588)4291850-9 gnd Chemokine (DE-588)4429989-8 gnd |
| subject_GND | (DE-588)4606532-5 (DE-588)4291850-9 (DE-588)4429989-8 (DE-588)4113937-9 |
| title | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |
| title_alt | Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |
| title_auth | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |
| title_exact_search | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |
| title_full | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren vorgelegt von Regine Brox aus Nürnberg |
| title_fullStr | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren vorgelegt von Regine Brox aus Nürnberg |
| title_full_unstemmed | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren vorgelegt von Regine Brox aus Nürnberg |
| title_short | Molecular mechanisms of functionally selective and probe-dependent signaling at CXCR3 induced by negative allosteric modulators |
| title_sort | molecular mechanisms of functionally selective and probe dependent signaling at cxcr3 induced by negative allosteric modulators molekulare mechanismen der funktionell selektiven und probenabhangigen signaltransduktion am cxcr3 rezeptor veranlasst durch negative allosterische modulatoren |
| title_sub | = Molekulare Mechanismen der funktionell selektiven und probenabhängigen Signaltransduktion am CXCR3 Rezeptor veranlasst durch negative allosterische Modulatoren |
| topic | Ligand Biochemie (DE-588)4606532-5 gnd Allosterischer Effektor (DE-588)4291850-9 gnd Chemokine (DE-588)4429989-8 gnd |
| topic_facet | Ligand Biochemie Allosterischer Effektor Chemokine Hochschulschrift |
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