G Protein-Coupled Receptors (eBook)

Dr David R. Poyner, University of Aston, Birmingham, UK; School of Life and Health Sciences, Aston University, Birmingham, UK. Professor?Mark Wheatley, University of Birmingham, UK.

Preface.

Contributors.

1 Measurement of Ligand-G-Protein-coupled ReceptorInteractions (Katie Leach, Celine Valant, Patrick M. Sextonand Arthur Christopoulos).

1.1 Introduction.

1.2 Methods and Approaches.

References.

2 Second Messenger Assays for G-Protein-coupled Receptors:cAMP, Ca¯2+, Inositol Phosphates¯, ERK1/2(Karen J. Gregory, Patrick M. Sexton, Arthur Christopoulosand Caroline A. Hick).

2.1 Introduction.

2.2 Methods and Approaches.

2.3 Troubleshooting.

References.

3 Use of the [¯35S]GTPgammaS Binding Assayto Determine Ligand Efficacy at G-protein-coupled Receptors(Elodie Kara and Philip G. Strange).

3.1 Introduction.

3.2 Methods and Approaches.

3.3 Troubleshooting.

Acknowledgements.

References.

4 Quantitative Imaging of Receptor Trafficking (AndyR. James, Takeo Awaji, F. Anne Stephenson and Nicholas A.Hartell).

4.1 Introduction.

4.2 Methods and Approaches.

4.3 Troubleshooting.

References.

5 Production of Recombinant G-Protein-coupled Receptor inYeast for Structural and Functional Analysis (Richard A.J.Darby, Mohammed Jamshad, Ljuban Grgic, William J. Holmes and RoslynM. Bill).

5.1 Introduction.

5.2 Methods and Approaches.

5.3 Troubleshooting.

References.

6 Monitoring GPCR-Protein Complexes UsingBioluminescence Resonance Energy Transfer (Werner C. Jaeger,Kevin D.G. Pfleger and Karin A. Eidne).

6.1 Introduction.

6.2 Methods and Approaches.

6.3 Troubleshooting.

References.

7 Using Intramolecular Fluorescence Resonance Energy Transferto Study Receptor Conformation (Cornelius Krasel and CarstenHoffmann).

7.1 Introduction.

7.2 Methods and Approaches.

7.3 Troubleshooting.

References.

8 A Disulfide Cross-linking Strategy Useful for StudyingLigand-induced Structural Changes in GPCRs (Jian Hua Li,Stuart D.C. Ward, Sung-Jun Han, Fadi F. Hamdan and JürgenWess).

8.1 Introduction.

8.2 Methods and Approaches.

8.3 General Considerations, Caveats and Troubleshooting.

Acknowledgements.

References.

9 Use of Fluorescence Correlation Spectroscopy to Study theDiffusion of G-protein-coupled Receptors (Stephen J.Briddon, Jonathan A. Hern and Stephen J. Hill).

9.1 Introduction.

9.2 Methods and Approaches.

9.3 Troubleshooting.

References.

10 Identification and Proteomic Analysis of GPCRPhosphorylation (Kok Choi Kong, Sharad C. Mistry and AndrewB. Tobin).

10.1 Introduction.

10.2 Methods.

Acknowledgements.

References.

11 Measurement and Visualization of G-Protein-coupledReceptor Trafficking by Enzyme-linked Immunosorbent Assay andImmunofluorescence (Stuart J. Mundell, Shaista P. Nisar andEamonn Kelly).

11.1 Introduction.

11.2 Methods and Approaches.

11.3 Troubleshooting.

References.

12 Substituted Cysteine Accessibility Method (SCAM)(George Liapakis and Jonathan A. Javitch).

12.1 Introduction.

12.2 Methods and Approaches.

12.3 Troubleshooting.

References.

13 Homology Modelling of G-Protein-coupled Receptors(John Simms).

13.1 Introduction.

13.2 Methods and Approaches.

13.3 Troubleshooting.

13.4 Automated Methods for Generating Models of GPCRs.

References.

Appendix: Site-directed Mutagenesis and Chimeras (AlexConner, Mark Wheatley and David Poyner).

A.1 Introduction.

A.2 Why Mutagenesis?

A.3 Troubleshooting.

A.4 Conclusion.

References.

Index.