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Duarte Miguel F. Prazeres and Sofi a Aires M. Martins (eds.), G Protein-Coupled Receptor Screening Assays, Methods in Molecular Biology, vol. 1272, DOI 10.1007/978-1-4939-2336-6, © Springer Science+Business Media New York 2015

A

Adenylate cyclase ......................................................5 , 10, 66 Adrenoceptors

α-adrenergic ....................................................................6 β-adrenergic receptor ......................................................6

Agonist biased agonism ....................................................2 16, 228 inverse ............................................8 , 9, 13, 104, 211, 235 partial .............................................5 , 9, 66, 211, 234, 235

Antagonist ...................................... 4 –6, 9, 13, 24, 72, 81, 83, 86, 109, 114, 119, 134, 184, 186, 205, 210, 211, 217, 219, 220, 223, 228, 238–241, 261, 268, 272, 275, 278, 279, 281–285, 287, 289

biased antagonism ....................................... 2 28, 238, 239 Antibody .........................................................8 , 9, 25, 27, 28,

121, 123–127, 129–131, 174, 176–178, 180–182, 184–186, 195, 248

β-arrestin ................................................................1 2, 14 Assays

complementation .......................................... 1 4, 108, 110 functional .................................................. 1 2, 16, 38, 258 impedance-based ................................................2 15–226 kinetic ...........................................................................1 2 ligand binding ...............................................................1 5

B

Baculoviruses BacMam ..................................................... 6 6, 67, 71, 76 budded ........................................................ 3 7–50, 70, 74

β-galactosidase .............................1 4, 74, 92, 93, 99, 101, 108 complementation assay .........................................1 4, 108

Binding experiments competition ........................................... 2 8, 31–32, 45–46 saturation .............................................. 3 1, 44–45, 48, 49

Bioluminescence ........................................ 1 7, 108, 258, 259, 261, 263–264, 267, 268

Bioluminescence resonance energy transfer (BRET) ........................................ 108, 133–141, 258

Biosensor label-free .............................................................1 99–212 microfluidic .................................................2 29–233, 243 optical .........................................................................2 28 resonant waveguide grating ................................1 99, 228

BRET. See Bioluminescence resonance energy transfer (BRET)

C

Calcium assay ....................................................................1 51–153 dye .......................................................... 1 6, 80, 145, 151 intracellular ............................1 1, 13, 79, 80, 85, 144–146 mobilization .......................................... 1 3, 79–82, 85–87 release ............................................................. 1 3, 80, 216

Calmodulin .......................................................................2 00 Carbon nanotube

fabrication ........................................... 1 90, 191, 193–194 single-walled carbon nanotube (SWNT) ............90–194,

196, 197 cDNA ..................................................4 1, 259, 260, 266, 267 Cell line

activation ......................................................................6 6 airway smooth muscle ................................. 8 1, 83, 85–87 CHO ............................................................................5 2 clone selection ...............................................................5 3 contact ....................................................................5 6, 63 generation .........................................................5 4, 57–58 HEK293 ..........................................51–64, 204, 263, 264 HeLA ...................................................................5 2, 217 heterologous ...............................................................1 11 inducible ...............................................................5 2, 204 PC12 ..................................................................111, 116 recombinant .....................5 1–64, 206, 208, 215, 216, 241 Sf9 ................................................................................74 stably transfected ............................................ 5 2, 58, 203 T-cell ............................................................................9 3 transfection ............................3 0, 54, 55, 57–58, 127, 260 T-REx-293 ....................................................... 53, 57, 62 U2OS .................................................................1 11, 116

Chimera .................................................7 , 13, 28, 92, 93, 101 Clathrin ..............................................................................1 1 CLIP-tag ® .............................................................. 28, 30, 33 Combinatorial chemistry ..................................................2 73 Computer-aided design ......................................1 7, 271–290 Confocal imaging .....................................................1 22–125 Constitutive activity ...................................... 7 , 9, 13, 91–105 Cryptates ............................................................................2 5 Crystallography studies ........................................................8 Cyan fluorescent protein ...................................................1 36 Cyclic adenosine monophosphate (cAMP) ............. 5, 10, 11,

13, 65–76, 93, 95, 108, 144, 190, 200, 228 assay ....................................................................1 14, 144

INDEX

294 G PROTEIN-COUPLED RECEPTOR SCREENING ASSAYS

Index

D

Deorphanization .................................................................1 3 DEP. See Dielectrophoresis (DEP) Desensitization

heterologous ...........................................................1 1, 13 homologous ..................................................................1 1 receptor .......................................................1 19, 237–238

dFRAP .....................................................................1 33–140 Dielectrophoresis (DEP) .......................... 248–250, 252–255 Docking ............................................ 2 73–276, 279–283, 289 Dose-response ........................................4 , 5, 42, 71–73, 109,

114, 220–222, 258, 262, 267, 2653 Dual-Luciferase ® reporter ..........................................9 9, 110 Dye

calcium .......................................................................1 51 homogeneous time resolved fluorescence

(HTRF ® ) ................................................................28 Dynamic mass redistribution (DMR) ..................... 200–212,

228–230, 232–244

E

Efficacy .................5 , 24, 27, 28, 205, 228, 235, 241, 258, 265 Electrical signals ...............................................................1 90 Electroporation ...................................................................7 4 Endocytosis ........................................................................1 1 Epac2-camps ................................................................6 6–68 Expression

stable .............................................................................6 7 transient ......................... 5 2, 103, 123, 125, 127, 260, 262

F

Field effect transistor ................................................1 94, 197 Firefly luciferase .............................................. 9 2, 93, 95, 99,

108–111, 113, 115, 117, 259, 260 FLAG-tagged ........................................... 1 20, 123–125, 130 FlexStation ................................................. 8 0–82, 84, 88, 89 FLIPR .................................................................... 8 0, 83, 87 Flow-cytometry ...........6 3, 120–122, 125–127, 176–182, 185 Fluorescence

acceptor ................. 2 4–26, 31, 34, 65, 134, 136, 138, 139 anisotropy ...............................................................3 7–50 donor ...........................................2 4–26, 31, 65, 136, 139 life-time ...................................................... 2 5, 38, 46, 65 photobleaching ...................................................1 39, 167 resonance ................................................................1 4, 24 resonance energy transfer ......................................2 4, 134

Fluorophores ............1 3, 24–26, 38, 39, 65, 72, 129, 131, 136 Förster distance ..................................................................2 4 Förster resonance energy transfer (FRET) ............ 23–35, 65,

66, 73, 134, 136, 139 time-resolved ..........................................................2 3–65

Fragment complementation ......................................1 07–117 Free energy perturbation (FEP) ............... 249, 276–278, 288 Functional selectivity .................................. 1 5, 202, 227–244

G

G-protein chimeric ..............................................................1 00, 102 G 12/13 .......................................................................11, 15 G 15 ................................................................................13 G 16 ................................................................................13 Gα ............................................................................. 9 , 10 Gβ/γ .......................................................................... 9 , 10 G i ........................................................................ 5, 11, 13 G o ................................................................................. 11 G q ....................................................................... 6, 11, 13 G s .................................................................... 5, 9, 11, 13 G t ................................................................................ 5, 6 heterotrimeric .....................................................6 , 9, 200 independent signaling .................................................1 20

G protein-coupled receptors (GPCRs) endogenous ..............................9 2, 93, 146, 216, 217, 272 expression ................................1 3, 30, 52, 53, 67, 92, 185 immobilization ...........................................1 34, 173–186 internalization ................................... 7 , 14, 119–131, 263 kinases ...............................................1 1, 14, 93, 108, 109 labeling .............................................................2 6–30, 38 ligand-gated ....................................................................8 orphan .................................7 , 9, 14, 91–93, 98–103, 105 signaling mechanism ................................................3 –17

Guanosine triphosphate nucleotide (GTP) ................ 5, 9, 10

H

HaloTag ® .....................................................................28, 33 Homology modeling ........................ 2 73, 274, 276, 278–282 Hydrodynamic tail vein (HTV) method .......... 2 58, 265, 268

I

Imaging binary ...........................................1 45, 147, 149, 154, 169 ImageJ ................................................................1 43–172 in vivo ......................................... 2 58, 261–262, 264–266 macro .................................................. 1 45, 146, 162–165 plug-ins ............................................... 1 44, 146, 147, 149 single cell .................................... 1 49, 258, 261, 263–264 threshold ......................1 45, 147–149, 154–158, 169, 171

Immunofluorescence ...........1 4, 120, 121, 123–127, 131, 267 Impedance ..........................................................1 5, 215–226 In silico ................................................................. 1 6, 17, 273 IP 3 .................................................................................10, 79

L

Label-free biosensor ............................................... 1 5, 199–212, 228 holistic readout ...........................................................2 06 integrated cell response ...............................................2 02

Lanthanides ........................................................................2 5 Lentivirus ...................................................................7 6, 175

particles ..............................................................1 75–176

G PROTEIN-COUPLED RECEPTOR SCREENING ASSAYS

295

Index

Ligand bias ......................................................................2 28, 233 binding dynamics ....................................................3 7–50 design ..........................................2 71, 272, 281, 286, 287 docking ............................................... 2 73–275, 282–283 endogenous ...........................................................9 2, 272 labeling ...................................................................2 6–27 surrogate .......................................................................9 2

M

Microfluidics ............................................... 1 5, 16, 228–232, 237–238, 241, 243, 247–255

Microscopy .....................................6 3, 65, 67, 108, 130, 137, 143–145, 176, 177, 180, 182, 248, 250, 258, 261

bioluminescence .................................. 2 58, 261, 263–264 Modelling .......... 2 31, 271–276, 278–280, 284, 286, 288, 289 Molecular

dynamics ............................................. 1 77, 273, 281–282 modeling .............................................................2 71, 286

Mouse model ....................................................................1 , 5

N

Nanovesicles .............................................................1 89–197

O

Oligomerization .................................................1 4, 133–140

P

Phospholipase C ............................................... 6 , 10, 79, 216 Plasmid ........................................... 2 9, 30, 33, 47, 48, 53, 54,

62, 63, 67–70, 73, 87, 95, 97, 110, 112, 113, 115, 116, 121, 127, 137, 138, 140, 175, 176, 178–180, 183, 185, 190, 191, 259–261, 265, 266, 268

Platform ...............................................1 3, 17, 28, 52, 55, 60, 113, 114, 202, 204, 208, 215, 216

Potency .................................................5 , 46, 66, 67, 76, 104, 105, 205, 228, 235, 241, 258, 273, 281, 283, 287

Protein kinase A .........................................................1 1, 200 Protein kinase C .................................................................1 1 Protein–protein interaction ......................... 1 4, 15, 108, 116,

130, 134, 259, 263

Q

QSAR.......................................................................2 76, 285

R

Ratiometric ................................................... 3 8, 67, 248, 253 Real-time ...................................... 8 , 12, 38, 65, 67, 134, 173,

197, 202, 211, 215, 216, 228, 233, 234, 236, 241–243 analysis ........................................................................2 02

Receptor adenosine .....................2 72–273, 278–281, 283, 286, 287 adrenergic .......................................................................6

cloning ........................................................................1 10 constitutive activity .................................................9 , 105 cross-talk ......................................................................1 3 degradation .................................................................1 20 dopamine1 ................................................ 12, 66, 67, 217 dopamine 2 ...........................................................12, 134 expression .............................................1 3, 30, 31, 35, 49,

59, 87, 175, 178, 182, 185, 204 histamine H1 ..............................................................217 human CXCR4 ..................................................184, 185 internalization ................ 7 , 11, 14, 35, 123–126, 130, 131 kisspeptin ............................................................1 19–131 long chain fatty acid FFA1 .........................................202 Melanocortin 4 (MC 4 ) ..........................40, 46, 48, 66, 76 muscarinic .......................................................................6 neurotensin .............................................................5 1–63 olfactory ..............................................................1 89–197 oligomerization .....................................................1 4, 134 purification .....................................................................8 recycling ..................................................................1 0, 35 rhodopsin ........................................................................9 vasopressin .................................................. 1 08, 110, 115

Renilla luciferase .................. 9 9, 110, 111, 113, 115, 117, 258 Reporter systems

fluorescence ..................................................................1 3 luminescence .................................................................1 3

Resonance waveguide grating (RWG) ...................... 15, 199, 228, 230, 231

S

Screening assays .................................................... 3 –16, 92, 189–19 high-throughput .......... 1 3, 24, 25, 91–117, 173, 241, 242

Second messengers ......................9 , 10, 12, 79, 200, 215, 216 Selection markers .............................................................1 15 Sensors

CM5 sensor chip ................................................1 77, 184 corning Epic biosensor ...............................................2 03 field-effect transistors .........................................1 94, 197 impedance based ...........................................................1 5 resonance waveguide grating (RWG) ...................15, 228 surface plasmon resonance (SPR) .......................173–186 xCELLigence real time cell analyzer ..........................2 16

Signalling mechanisms ..............................................................3 –16 pathway ..................................................4 , 12, 13, 15, 16,

24, 79, 80, 92, 100, 190, 200, 201, 210, 216, 223, 247 transduction ...............................3 , 13, 133, 197, 215, 247

Single-cell .............................................5 8, 63, 136, 149, 243, 247–255, 258, 261, 263–264

SNAP-tag ............................................................... 2 8, 30, 33 Spectroscopy .........................................................................8

fluorescence ....................................................................8 Split luciferase .................................. 2 58–263, 265, 266, 268

296 G PROTEIN-COUPLED RECEPTOR SCREENING ASSAYS

Index

Split TEV .................................................................1 07–117 SPR. See Surface plasmon resonance (SPR) Stokes shift .........................................................................2 5 Structure

based drug design ...............................................2 73, 275 crystal ..............................9 , 271–273, 276, 278, 279, 286

Surface plasmon resonance (SPR) ...................... 15, 173–186

T

Tag ........................................................................ 5 4, 56, 174 Tangential flow filtration (TFF) ............................ 41, 43, 47,

49, 68, 70, 73, 75 T Epac VV .............................................................6 6–69, 71–72 Tetracycline-inducible expression system ...........................5 2

TFF. See Tangential flow filtration (TFF) Transfection ....................................3 0, 41, 48, 54, 55, 57–58,

62, 63, 66, 68, 69, 73, 74, 87, 95, 98–100, 102, 104, 108, 110–113, 115, 116, 121, 122, 127, 128, 131, 137–139, 175, 176, 178–179, 185, 191, 195, 260

V

Virus amplification ............................................... 4 1–42, 73, 74 cytomegalovirus .................................. 5 2, 53, 67, 69, 103 titer ..................................................4 1–42, 68, 71, 73, 75

Y

Yellow fluorescent protein ........................................1 00, 136