Guide to the Book Plant Kinases
The growing number of fully annotated genomes of model and nonmodel plant species, such as Arabidopsis, Brachypodium, grapevine, maize, rice, rape seed, soybean, tomato, and others, has led to a tremendous increase in sequence information. The novel geno
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Dissmeyer and A. Schnittger (eds.), Plant Kinases: Methods and Protocols, Methods in Molecular Biology, vol. 779, DOI 10.1007/978-1-61779-264-9_1, © Springer Science+Business Media, LLC 2011
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N. Dissmeyer and A. Schnittger
Fig. 1. Approaches to the study of protein kinase function found in this book. All examples are taken from this book, indicating in which chapter the respective methodologies can be found.
Plant Kinases is divided into four major sections comprising a total of 17 chapters. It starts with (I) an introductory chapter (Chapter 2) where we review general structural and functional aspects of protein kinase function highlighting several regulatory signaling networks related to this methods book. In addition, we provide a compendium of databases and Web tools for plant kinase research. Part (II) covers expression, purification, activity monitoring, and reporter gene assays of plant protein kinases; part (III) addresses the study of kinase function at the cellular level by chemical genetics, microscopy- and spectroscopy-based techniques; and part (IV) provides information on the identification of kinase substrates and phosphorylation-site detection via interaction assays, substrate labeling protocols, and proteomics (Fig. 1). Part II (Chapters 3–8) starts off by providing several methods for protein production and isolation, for instance, through immunoprecipitation from plant extracts (Chapters 4 and 5). Of particular relevance to the study of kinases and substrates for which only low amounts can be isolated from plant extracts, in vitro protein production, such as cell-free systems (Chapter 3), or bacterial and viral expression systems (Chapter 4), might be considered.
1 Guide to the Book Plant Kinases
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Protein expression is typically followed by protein purification steps and protocols are provided in Chapters 3 and 4. The ultimate step is a functional assay, for instance, by affinity-coupled kinase (Chapters 4 and 6) and phosphatase assays (Chapter 8). In parallel, activity can be monitored in planta by reporter gene assays (Chapter 5) or in heterologous systems such as bacteria (Chapter 7; Fig. 1). Part (III) (Chapters 9–13) addresses the study of kinase function at the cellular level through the lens of chemical genetics (Chapters 9 and 10), microscopy, and spectroscopy (Chapters 11–13). The modulation of kinase activity with inhibitor molecules has become a very powerful tool as outlined in Chapter 9. Recent advances in inhibitor studies make use of ATP-competitive molecules and engineered inhibitor-sensitive protein kinase variants that are replacing the wild-type copy (Chapter 10). Highly sensitive microscopical techniques are applied to reach a quantitative level in detecting and monitor protein–protein interactions. Standard fluorescence or confocal laser scanning microscopy (CLSM; Chapters 12 and 14) are used to elaborate the transient associations of proteins by Förster/fluorescence resonance energy transfer measurements (FRET; Chapters 11 and 13) and fluorescence lifetime ima
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