Crane Two-Component Signaling Systems, Part C

1. Auflage 2010
ISBN: 978-0-12-381348-0
Verlag: Elsevier Science & Techn.
Format: EPUB
Kopierschutz: 6 - ePub Watermark

E-Book, Englisch, Band Volume 471, 496 Seiten

Reihe: Methods in Enzymology

Multicellular organisms must be able to adapt to cellular events to accommodate prevailing conditions. Sensory-response circuits operate by making use of a phosphorylation control mechanism known as the 'two-component system.' This volume, the third in a three-volume treatment edited by the same group of editors, includes a wide range of methods, including those dealing with the Sln-1 kinase pathway, triazole sensitivity in C. albicans, and histidine kinases in cyanobacteria circadian clock. - Includes time-tested core methods and new innovations applicable to any researcher studing two-component signaling systems or histidine kinases - Methods included are useful to both established researchers and newcomers to the field - Relevant background and reference information given for procedures can be used as a guide to developing protocols in a number of disciplines

Autoren/Hrsg.

Crane, Brian

Crane, Alexandrine

Inhaltsverzeichnis

1;Front Cover;1
2;Methods in Enzymology Two-Component Signaling Systems, Part C;4
3;Copyright Page;5
4;Contents;6
5;Contributors;14
7;Chapter 1: Characterizing Cross-Talk In Vivo: Avoiding Pitfalls and Overinterpretation;48
7.1;Abstract;48
7.2;1. Overview;49
7.3;2. Sources of Cross-Talk;50
7.4;3. Cross-Talk Suppression;51
7.5;4. Transcriptional Reporters;53
7.6;5. Response Regulator Localization;55
7.7;6. Phosphatase Cross-Talk;60
7.8;7. Signal Response in Cross-Talk Networks;61
7.9;8. Concluding Remarks;61
7.10;Acknowledgments;62
7.11;References;62
8;Chapter 2: Inference of Direct Residue Contacts in Two-Component Signaling;64
8.1;Abstract;64
8.2;1. Introduction;65
8.3;2. Extraction Tools;71
8.4;3. DCA: Direct Coupling Analysis;75
8.5;Appendix: Nonstandard Linear Algebra Functions;85
8.6;Acknowledgments;86
8.7;References;86
9;Chapter 3: Computational Modeling of Phosphotransfer Complexes in Two-Component Signaling;90
9.1;Abstract;90
9.2;1. Introduction;91
9.3;2. Methods;94
9.4;3. Summary;101
9.5;Acknowledgments;102
9.6;References;102
10;Chapter 4: Kinetic Studies of the Yeast His-Asp Phosphorelay Signaling Pathway;106
10.1;Abstract;106
10.2;1. Introduction;107
10.3;2. Materials and Methods;109
10.4;3. Conclusion;120
10.5;Acknowledgments;121
10.6;References;121
11;Chapter 5: Purification of MBP-EnvZ Fusion Proteins Using an Automated System;124
11.1;Abstract;124
11.2;1. Introduction;125
11.3;2. Comparison Between the Cytoplasmic Domains of E. coli and Typhi EnvZ Proteins;127
11.4;3. Purification of the Recombinant MBP Proteins by FPLC;129
11.5;4. SDS-PAGE Analysis of Recombinant Expressed MBP-EnvZc Protein;129
11.6;5. Results;131
11.7;6. Discussion;132
11.8;Acknowledgments;132
11.9;References;132
12;Chapter 6: Measurement of Response Regulator Autodephosphorylation Rates Spanning Six Orders of Magnitude;136
12.1;Abstract;137
12.2;1. Overview of Response Regulator Autodephosphorylation;137
12.3;2. Purification of Response Regulator Proteins;138
12.4;3. General Considerations for Autodephosphorylation Assays;139
12.5;4. Assay of Autodephosphorylation by Loss of 32P;141
12.6;5. Assay of Autodephosphorylation by Fluorescence;148
12.7;6. Assay of Autodephosphorylation by Pi Release;153
12.8;7. Assay of Autodephosphorylation from Systems of Reactions;155
12.9;8. Future Prospects;157
12.10;Acknowledgments;158
12.11;References;158
13;Chapter 7: Transmembrane Receptor Chimeras to Probe HAMP Domain Function;162
13.1;Abstract;162
13.2;1. Introduction;163
13.3;2. Design of Chimeras of HAMP Domains Fused to the AC Rv3645 Catalytic Domain;164
13.4;3. Choice of Vector and Cloning Strategy;165
13.5;4. Expression and Purification of the Chimeras;165
13.6;5. AC Assay;167
13.7;6. Example of Applications of the Method and Results;167
13.8;7. Concluding Remarks;168
13.9;Acknowledgments;169
13.10;References;169
14;Chapter 8: Light-Activated Bacterial LOV-Domain Histidine Kinases;172
14.1;Abstract;172
14.2;1. Introduction;173
14.3;2. Description of Method;174
14.4;3. Concluding Remarks and Future Perspectives;179
14.5;Acknowledgments;180
14.6;References;180
15;Chapter 9: Characterization of Bacteriophytochromes from Photosynthetic Bacteria;182
15.1;Abstract;183
15.2;1. Introduction;183
15.3;2. Cloning BphP in Expression Vector;186
15.4;3. Overexpression and Purification of BphPs;187
15.5;4. Autophosphorylation;188
15.6;5. Phosphotransfer;190
15.7;6. Gel Mobility Shift Assay and DNase I Footprint;190
15.8;7. Gene Disruption;192
15.9;8. Analysis of the Photosynthetic Phenotypes of the BphP Mutants;194
15.10;9. Photochemical Measurements;195
15.11;References;205
16;Chapter 10: Biophysical Assays for Protein Interactions in the Wsp Sensory System and Biofilm Formation;208
16.1;Abstract;208
16.2;1. Introduction;209
16.3;2. Analyses of the Shape and Molecular Weight of Proteins and Protein Complexes;210
16.4;3. Experimental Considerations;218
16.5;4. Case Studies;221
16.6;5. Concluding Remarks;227
16.7;Acknowledgments;228
16.8;References;228
17;Chapter 11: High-Throughput Screening of Bacterial Protein Localization;232
17.1;Abstract;233
17.2;1. Introduction;233
17.3;2. Pipeline Overview;234
17.4;3. Construction of a Caulobacter ORFeome;235
17.5;4. Construction of the Fluorescently Tagged Protein Library;237
17.6;5. Imaging the Caulobacter Protein Localization Library;242
17.7;6. Image Scoring and Analysis;244
17.8;7. Conclusion;248
17.9;Acknowledgments;249
17.10;References;250
18;Chapter 12: In Vitro and In Vivo Analysis of the ArcB/A Redox Signaling Pathway;252
18.1;Abstract;252
18.2;1. Introduction;253
18.3;2. In Vitro Characterization of the Arc TCS;255
18.4;3. In Vivo Characterization of the Arc TCS;267
18.5;4. Conclusions;273
18.6;Acknowledgments;273
18.7;References;273
19;Chapter 13: Potassium Sensing Histidine Kinase in Bacillus subtilis;276
19.1;Abstract;276
19.2;1. Introduction;277
19.3;2. Screen for Molecules that Stimulate KinC Sensor Kinase;278
19.4;3. Quantitative Analysis of the Activation of KinC;283
19.5;4. Structural Analysis of KinC;287
19.6;5. Monitoring the Signals Using Indirect Measurements;291
19.7;6. Applications of the System Signal-Kinase;292
19.8;7. Conclusions;295
19.9;References;296
20;Chapter 14: Two-Component Systems and Regulation of Developmental Progression in Myxococcus xanthus;300
20.1;Abstract;301
20.2;1. Introduction;301
20.3;2. Generation of In-Frame Deletions or Point Mutations in the M. xanthus Genome;304
20.4;3. Phenotype Assays for Analysis of M. xanthus Development;310
20.5;4. Expression Analysis;314
20.6;5. In Vitro Biochemical Analysis of TCS Proteins;317
20.7;Acknowledgments;322
20.8;References;322
21;Chapter 15: Two-Component Signaling to the Stress MAP Kinase Cascade in Fission Yeast;326
21.1;Abstract;326
21.2;1. Introduction;327
21.3;2. Detection of Protein Interactions in the H2O2 Signaling Pathway;329
21.4;3. Detection of Cysteine S-Thiolation in Tdh1 GAPDH;333
21.5;Acknowledgments;335
21.6;References;335
22;Chapter 16: Genetic and Biochemical Analysis of the SLN1 Pathway in Saccharomyces cerevisiae;338
22.1;Abstract;338
22.2;1. Introduction;339
22.3;2. Materials and Methods;344
22.4;Acknowledgments;362
22.5;References;362
23;Chapter 17: Analysis of Mitogen-Activated Protein Kinase Phosphorylation in Response to Stimulation of Histidine Kinase Signa;366
23.1;Abstract;366
23.2;1. Introduction;367
23.3;2. Growth of Cultures and Exposure to Hyperosmotic Conditions or Fungicide;369
23.4;3. Mitogen-Activated Protein Kinase Assay;371
23.5;4. Adapting the MAPK Assay;378
23.6;5. Discussion;378
23.7;Acknowledgments;379
23.8;References;379
24;Chapter 18: Biochemical Characterization of Plant Hormone Cytokinin-Receptor Histidine Kinases Using Microorganisms;382
24.1;Abstract;382
24.2;1. Introduction;383
24.3;2. Characterization of Plant TCS Components in E. coli;387
24.4;3. Protocol for Histidine Kinase Assays in E. coli;390
24.5;4. Protocol of Cytokinin-Binding Assay with Intact E. coli Cells;392
24.6;5. Preparation of Radioactive Phospho-HPt Factor, and In Vitro Assay of Phosphotransfer to RR;392
24.7;6. Characterization of Plant TCS Components in S. cerevisiae;394
24.8;7. Protocol of Histidine Kinase Assay in S. cerevisiae;396
24.9;8. Protocol of HPt Factor Assay in S. cerevisiae;397
24.10;9. Protocol of Cytokinin-Binding Assay by Using S. pombe Membranes Enriched in AHK4/CRE1;397
24.11;Acknowledgments;398
24.12;References;398
25;Chapter 19: Characterization of Pseudo-Response Regulators in Plants;404
25.1;Abstract;404
25.2;1. The Arabidopsis Circadian Clock;405
25.3;2. Detection of PRR Proteins;407
25.4;3. Localization of PRR Proteins;411
25.5;4. Exploring the Circadian Phenotypes of prr Mutants;414
25.6;5. Concluding Remarks;422
25.7;Acknowledgments;422
25.8;References;422
26;Chapter 20: Reversible Histidine Phosphorylation in Mammalian Cells;426
26.1;Abstract;426
26.2;1. Introduction;427
26.3;2. Analysis of Phosphorylation and Dephosphorylation of Histidine Residues In Vitro;428
26.4;3. Functional Analysis of NDPK/PHPT-1 Regulated Systems in Living Cells;434
26.5;References;447
27;Chapter 21: Histidine Phosphorylation in Histones and in Other Mammalian Proteins;450
27.1;Abstract;450
27.2;1. Introduction;451
27.3;2. Chemical Phosphorylation of Histone H4 Proteins and Peptides;452
27.4;3. Detection of Phosphohistidine-Phosphoamino Acid Analysis;453
27.5;4. Filter-Based Assay of Alkali-Stable, Acid-Labile Protein Phosphorylation (Nytran Assay);462
27.6;5. In-Gel Kinase Assay;463
27.7;6. Phosphorylation and Thiophosphorylation Site Analysis by Edman Sequencing;466
27.8;7. Mass Spectrometric Phosphopeptide Analysis;468
27.9;Acknowledgments;471
27.10;References;471
28;Author Index;474
29;Subject Index;482