Protein'Protein Interactions: Methods and ApplicationsHaian Fu As the mysteries stored in our DNA have been more completely revealed, scientists have begun to face the extraordinary challenge of unraveling the int- cate network of protein–protein interactions established by that DNA fra- work. It is increasingly clear that proteins continuously interact with one another in a highly regulated fashion to determine cell fate, such as proliferation, diff- entiation, or death. These protein–protein interactions enable and exert str- gent control over DNA replication, RNA transcription, protein translation, macromolecular assembly and degradation, and signal transduction; essentially all cellular functions involve protein–protein interactions. Thus, protein–p- tein interactions are fundamental for normal physiology in all organisms. Alt- ation of critical protein–protein interactions is thought to be involved in the development of many diseases, such as neurodegenerative disorders, cancers, and infectious diseases. Therefore, examination of when and how protein–p- tein interactions occur and how they are controlled is essential for understa- ing diverse biological processes as well as for elucidating the molecular basis of diseases and identifying potential targets for therapeutic interventions. Over the years, many innovative biochemical, biophysical, genetic, and computational approaches have been developed to detect and analyze p- tein–protein interactions. This multitude of techniques is mandated by the diversity of physical and chemical properties of proteins and the sensitivity of protein–protein interactions to cellular conditions. |
Contents
15 | |
Characterization of ProteinProtein Interactions | 35 |
Norma J Greenfield 5 ProteinProtein Interaction Analysis by Nuclear Magnetic | 55 |
Resonance Spectroscopy | 79 |
Measuring RhodopsinGProtein Interactions | 93 |
Using Light Scattering to Determine the Stoichiometry | 113 |
Sedimentation Equilibrium Studies | 119 |
Analysis of ProteinProtein Interactions by Simulation | 137 |
The SplitUbiquitin MembraneBased Yeast TwoHybrid System | 297 |
Saccharomyces cerevisiae | 313 |
Mammalian TwoHybrid Assay for Detecting ProteinProtein | 327 |
CoImmunoprecipitation from Transfected Cells | 337 |
Microscopic Analysis of Fluorescence Resonance Energy | 351 |
Monitoring Molecular Interactions in Living Cells Using | 371 |
A New Tool | 383 |
Confocal Microscopy for Intracellular CoLocalization of Proteins | 399 |
Fluorescence Gel Retardation Assay to Detect | 155 |
Fluorescence Polarization Assay to Quantify | 161 |
GlutathioneSTransferaseFusion Based Assays for Studying | 175 |
Affinity Capillary Electrophoresis Analyses of ProteinProtein | 187 |
Mapping ProteinLigand Interactions by HydroxylRadical | 199 |
Use of Phage Display and Polyvalency to Design Inhibitors | 213 |
A Bacterial TwoHybrid System Based on Transcriptional Activation | 231 |
Using the Yeast TwoHybrid System to Identify | 247 |
Yeast TwoHybrid System | 263 |
Mapping Biochemical Networks with ProteinFragment | 411 |
In Vivo Protein CrossLinking | 427 |
Computational Prediction of ProteinProtein Interactions | 445 |
Affinity Methods for PhosphorylationDependent Interactions | 469 |
TwoDimensional Gel Electrophoresis for Analysis | 479 |
Sample Preparation of Gel Electrophoretically Separated | 499 |
Quantitative Protein Analysis by Solid Phase Isotope Tagging | 511 |
Internet Resources for Studying ProteinProtein Interactions | 519 |
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Common terms and phrases
acceptor activation affinity amino acid analysis antibody approx assay association constant bait beads binding Biol cells centrifugation Chem cloning coli colonies column concentration containing cross-linking database detection determined dilution dimer dissociation domain donor electrophoresis elution equilibrium Escherichia coli experimental expression filter fluorescence fluorophore FRET fusion proteins G-protein green fluorescent protein Gẞy HSQC Incubate inhibitor injection interac kinase labeled laser ligand lysate lysis buffer measurements medium molecular weight molecules monomer motif mutants NaCl Note parameters peptide phage phage display phosphorylation plasmid plates probe protein complexes protein interactions protein of interest protein–protein interactions purified reaction reagent receptor reporter genes residues resuspend rhodopsin room temperature sample scan Scansite screening SDS-PAGE selection sequence signal solution specific step strain structure Subheading subunit target tein tion titration transfection transformation Tris-HCl tube two-hybrid system vector vivo volume wash Western blot X-Gal yeast
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