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E-grāmata: Constitutive Activity in Receptors and Other Proteins, Part A

Volume editor Melvin I. Simon (The Salk Institute, La Jolla, CA, USA)

Formāts: EPUB+DRM
Sērija : Methods in Enzymology
Izdošanas datums: 26-Nov-2010
Izdevniecība: Academic Press Inc
Valoda: eng
ISBN-13: 9780123812995

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Proteins

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Formāts: EPUB+DRM
Sērija : Methods in Enzymology
Izdošanas datums: 26-Nov-2010
Izdevniecība: Academic Press Inc
Valoda: eng
ISBN-13: 9780123812995

Citas grāmatas par šo tēmu:

Proteins

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This volume of Methods in Enzymology covers the current methodology for the detection and assessment of constitutively active proteins. The chapters written by expert authors who are leaders in the field, provide hints and tricks not available in primary research publications.It is extensively referenced, with useful figures and tables throughout the volume.

Expert authors who are leaders in the field
Extensively referenced and useful figures and tables
Provides hints and tricks to facilitate reproduction of methods

This volume of Methods in Enzymology covers the current methodology for the detection and assessment of constitutively active proteins. The chapters written by expert authors who are leaders in the field, provide hints and tricks not available in primary research publications.It is extensively referenced, with useful figures and tables throughout the volume.

A. Expert authors who are leaders in the field

B. Extensively referenced and useful figures and tables

C. Provides hints and tricks to facilitate reproduction of methods

Papildus informācija

This volume of Methods in Enzymology covers the current methodology for the detection and assessment of constitutively active proteins.

Contributors

Preface

xxv

Volumes in Series

xvii

Section I Identification and Measurement of Constitutive Activity

1 Constitutive Activity at the Cannabinoid CB1 Receptor and Behavioral Responses

(28)

Katherine E. Hanlon

Todd W. Vanderah

1 Introduction

(2)

2 Modifying CB1 Activity

(6)

3 Behavioral Models

(14)

References

(5)

2 Detecting Constitutive Activity and Protean Agonism at Cannabinoid-2 Receptor

(22)

Massimiliano Beltramo

Rossella Brusa

Isabella Mancini

Paola Scandroglio

1 Introduction

(2)

2 General Considerations

(1)

3 Evaluation of Constitutive Activity Using GTPγS Assay

(4)

4 Evaluation of Constitutive Activity Using cAMP Assay

(3)

5 Evaluation of Constitutive Activity Using RT-CES

(4)

6 Evaluation of Protean Agonism with cAMP Assay

(2)

7 Comparison of the Methods

(1)

References

(4)

3 Modulation of the Constitutive Activity of the Ghrelin Receptor by Use of Pharmacological Tools and Mutagenesis

(22)

Jacek Mokrosinski

Birgitte Holst

1 Introduction

(1)

2 The Ghrelin Receptor and its Constitutive Activity

(2)

3 Structural Basis of Constitutive Activity

(4)

4 Residues Responsible for the Inverse Agonism and Efficacy Swap

(4)

5 Physiological Relevance of Constitutive Activity

(1)

6 Experimental Procedures

(3)

References

(6)

4 Assessment of Constitutive Activity and Internalization of GPR54 (KISS1-R)

(20)

Macarena Pampillo

Andy V. Babwah

1 Introduction

(1)

2 Materials

(2)

3 Methods

(12)

Acknowledgment

(1)

References

(3)

5 Assessment of Constitutive Activity in E-Type Prostanoid Receptors

(14)

Hiromichi Fujino

Toshihiko Murayama

John W. Regan

1 Introduction

(1)

2 Assays Used to Assess EP3 Receptor Constitutive Activity

(8)

3 Assays Used to Assess EP4 Receptor Constitutive Activity

105

(2)

References

107

(2)

6 α1D-Adrenergic Receptors: Constitutive Activity and Reduced Expression at the Plasma Membrane

109

(18)

J. Adolfo Garcia-Sainz

M. Teresa Romero-Avila

Luz del Carmen Medina

1 Introduction

110

(1)

2 Methods

111

(3)

3 Constitutive Activity

114

(3)

4 Plasma Membrane α1D-Adrenergic Receptors

117

(2)

5 Possible Physiological Implications

119

(2)

Acknowledgments

121

(1)

References

121

(6)

7 Constitutive Activity of the Histamine H1 Receptor

127

(22)

Saskia Nijmeijer

Rob Leurs

Henry F. Vischer

1 Introduction

128

(5)

2 Methods to Study Constitutive H1R Signaling

133

(9)

3 Constitutive Activity as Tool to Elucidate Receptor Activation and Crosstalk

142

(2)

4 Conclusion

144

(1)

Acknowledgments

144

(1)

References

145

(4)

8 Constitutive Activity of Somatostatin Receptor Subtypes

149

(16)

Anat Ben-Shlomo

Kolja Wawrowsky

Shlomo Melmed

1 Introduction

150

(1)

2 Choosing Cells to be Studied

151

(1)

3 Modifying SSTR Density

152

(11)

4 Summary

163

(1)

References

163

(2)

9 Assessment of Homologous Internalization of Constitutively Active N111G Mutant of AT1 Receptor

165

(14)

Mohiuddin Ahmed Bhuiyan

Takafumi Nagatomo

1 Introduction

166

(1)

2 Preparation of Receptor Plasmid and Protein

167

(1)

3 Radioligand Binding Assay

168

(2)

4 Inositol Phosphate Accumulation Assay

170

(1)

5 Internalization Assay

171

(2)

6 Western Blot Analysis

173

(1)

7 Data Analysis

174

(1)

8 Concluding Remark

175

(1)

References

175

(4)

10 Methods to Detect Cell Surface Expression and Constitutive Activity of GPR6

179

(18)

Balakrishna M. Prasad

Bettye Hollins

Nevin A. Lambert

1 Introduction

180

(1)

2 GPR6 is Expressed in Intracellular Compartments

180

(7)

3 Comparison of the Cell Surface Protein Detection Methods

187

(5)

4 Comparison of the Constitutive Gs-Activity Detection Methods

192

(2)

5 Conclusions

194

(1)

References

194

(3)

11 β3-Adrenoceptor Agonists and (Antagonists as) Inverse Agonists: History, Perspective, Constitutive Activity, and Stereospecific Binding

197

(34)

Maria Grazia Perrone

Antonio Scilimati

1 Introduction

198

(3)

2 β3-Adrenoceptor

201

(13)

3 Methodologies

214

(10)

Acknowledgments

224

(1)

References

225

(6)

12 Constitutive Activity of the Lutropin Receptor and Its Allosteric Modulation by Receptor Heterodimerization

231

(22)

Deborah L. Segaloff

1 Introduction

232

(3)

2 General Principles for Quantifying Receptor Activation

235

(4)

3 Modifying Cell Surface Expression Levels of Recombinant hLHR and Mutants Thereof

239

(4)

4 Quantifying Cell Surface hLHR Expression

243

(2)

5 Quantifying cAMP Production in Cells Expressing the hLHR

245

(1)

6 Experimental Strategies for Characterizing the Attenuating Effects of a Signaling Inactive hLHR on a Coexpressed wt or CAM hLHR

246

(4)

Acknowledgment

250

(1)

References

250

(3)

13 Assessing Constitutive Activity of Extracellular Calcium-Sensing Receptors in Vitro and in Bone

253

(14)

Wenhan Chang

Melita Dvorak

Dolores Shoback

1 Introduction

254

(4)

2 Materials and Methods

258

(4)

3 Results

262

(1)

4 Conclusions

263

(2)

Acknowledgments

265

(1)

References

265

(2)

14 Constitutive Activity of Neural Melanocortin Receptors

267

(14)

Ya-Xiong Tao

Hui Huang

Zhi-Qiang Wang

Fan Yang

Jessica N. Williams

Gregory V. Nikiforovich

1 Introduction

268

(1)

2 Signaling Assay for the Neural Melanocortin Receptors

268

(2)

3 Naturally Occuring Constitutively Active MC4R Mutants

270

(3)

4 Inverse Agonism of AgRP at the MC3R

273

(1)

5 Computational Modeling of the Constitutively Active MC4R Mutants

273

(4)

Acknowledgments

277

(1)

References

277

(4)

15 Measurement of Constitutive Activity of BMP Type I Receptors

281

(14)

David J. J. de Gorter

Maarten van Dinther

Peter ten Dijke

1 Introduction

281

(3)

2 Determining ALK2 Constitutive Activity

284

(5)

3 Determining the Effects of ALK2 Constitutive Activity on Osteoblast Differentiation

289

(2)

4 Concluding Remarks

291

(1)

Acknowledgments

292

(1)

References

292

(3)

16 Probing the Constitutive Activity Among Dopamine D1 and D5 Receptors and Their Mutants

295

(34)

Bianca Plouffe

Jean-Philippe D'Aoust

Vincent Laquerre

Binhui Liang

Mario Tiberi

1 Introduction

296

(3)

2 Design of Genetically Modified D1-Like Receptor Constructs and Cloning Strategy

299

(2)

3 Transfection of D1R and D5R Expression Constructs in HEK293 Cells

301

(2)

4 Radioligand-Binding Assays

303

(2)

5 Whole Cell cAMP Assays

305

(8)

6 Results Validating Experimental Approaches

313

(11)

7 Concluding Remarks

324

(2)

Acknowledgments

326

(1)

References

327

(2)

17 Identification of Gain-of-Function Variants of the Human Prolctin Receptor

329

(28)

Vincent Goffin

Roman L. Bogorad

Philippe Touraine

1 Introduction

330

(3)

2 Experimental Procedures

333

(12)

3 Identification of Constitutive Activity: Results and Discussion

345

(8)

4 Conclusions

353

(1)

Acknowledgments

353

(1)

References

353

(4)

18 Investigations of Activated ACVR1/ALK2, a Bone Morphogenetic Protein Type I Receptor, That Causes Fibrodysplasia Ossificans Progressiva

357

(18)

Frederick S. Kaplan

Petra Seemann

Julia Haupt

Meiqi Xu

Vitali Y. Lounev

Mary Mullins

Eileen M. Shore

1 Introduction

358

(1)

2 Patient Methodologies

359

(3)

3 Cellular Methodologies

362

(4)

4 Tissue Methodologies

366

(3)

5 In Vivo Methodologies

369

(2)

Acknowledgments

371

(1)

References

371

(4)

19 Identification and Evaluation of Constitutively Active Thyroid Stimulating Hormone Receptor Mutations

375

(22)

Joaquin Lado-Abeal

Leah R. Quisenberry

Isabel Castro-Piedras

1 Introduction

376

(6)

2 TSHR Gene Mutational Screening

382

(4)

3 Determination of TSHR Constitutive Activity In Vitro

386

(2)

4 Measurement of TSHR Expression at Cell Surface by Flow Cytometry Analysis

388

(1)

5 TSH-TSHR Binding Assays

389

(2)

6 TSHR Phosphorylation Analysis

391

(1)

Acknowledgments

392

(1)

References

392

(5)

20 Assessment of Constitutive Activity of a G Protein-Coupled Receptor, Cpr2, in Cryptococcus neoformans by Heterologous and Homologous Methods

397

(16)

Chaoyang Xue

Yina Wang

Yen-Ping Hsueh

1 Introduction of Receptors and Constitutive Receptors

398

(1)

2 Identification of Cpr2 as a Natural Occuring Constitutively Active Receptor

399

(10)

3 Additional Constitutively Active Receptors Identified in Fungi

409

(1)

Acknowledgments

409

(1)

References

409

(4)

21 In Vitro and In Vivo Assessment of Mu Opioid Receptor Constitutive Activity

413

(32)

Edward J. Bilsky

Denise Giuvelis

Melissa D. Osborn

Christina M. Dersch

Heng Xu

Richard B. Rothman

1 Introduction

414

(1)

2 Measuring Opioid Receptor Constitutive Activity in Vitro

415

(10)

3 cAMP Quantification Assay in CHO Cells Expressing Cloned Opioid Receptors

425

(5)

4 In Vivo Assessment of Antagonist Potency in Opioid Naive Subjects

430

(5)

5 In Vivo Assessment of Antagonist Potency to Precipitate Withdrawal

435

(5)

6 Summary

440

(1)

Acknowledgments

441

(1)

References

441

(4)

22 Constitutively Active μ-Opioid Receptors

445

(26)

Mark Connor

John Traynor

1 Introduction

446

(1)

2 Methods for Measuring Constitutive Activity

447

(18)

3 Conclusions

465

(1)

Acknowledgments

465

(1)

References

466

(5)

23 Protein Kinase CK2 is a Constitutively Active Enzyme that Promotes Cell Survival: Strategies to Identify CK2 Substrates and Manipulate its Activity in Mammalian Cells

471

(24)

Jacob P. Turowec

James S. Duncan

Ashley C. French

Laszlo Gyenis

Nicole A. St. Denis

Greg Vilk

David W. Litchfield

1 Introduction

472

(2)

2 Purification of CK2 for In Vitro Studies

474

(7)

3 Assays for CK2 Activity

481

(4)

4 Modulation of CK2 in Mammalian Cells

485

(6)

5 Conclusions

491

(1)

Acknowledgments

491

(1)

References

492

(3)

24 Assessment of CK2 Constitutive Activity in Cancer Cells

495

Maria Ruzzene

Giovanni Di Maira

Kendra Tosoni

Lorenzo A. Pinna

1 Introduction

496

(3)

2 Assay of CK2 in Crude Biological Samples

499

(5)

3 In-Cell Assay of Endogenous CK2 Activity

504

(2)

4 Identification/Validation of in Vivo CK2 Targets with Specific Inhibitors

506

(5)

Acknowledgments

511

(1)

References

511

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E-grāmata: Constitutive Activity in Receptors and Other Proteins, Part A

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