GTPase Protocols

The Ras Superfamily

Inbunden, Engelska, 2002

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Edward J. Manser and Thomas Leung have collected the key techniques currently in use to probe the function of these ubiquitous proteins both in vitro and in vivo. Presented in a format that ensures ready reproducibility by accomplished experimentalists who have refined the various methods in their laboratories, each technique includes step-by-step instructions, tips on avoiding pitfalls and troubleshooting, and ancillary notes explaining how to adapt each procedure in the event of problems. The methods cover the spectrum of core techniques required for the five major GTPase subfamilies (Ras, Rho, Rab, Arf, and Ran) and permit a diversity of applications ranging from structural studies on a GTPase to real time in vivo analysis. Timely and highly practical, GTPase Protocols: The Ras Superfamily illuminates the powerful techniques used by investigators today to study this special family of proteins that plays such important roles in human health and disease.

Produktinformation

Utgivningsdatum2002-06-05
Mått155 x 235 x 20 mm
Vikt568 g
FormatInbunden
SpråkEngelska
SerieMethods in Molecular Biology
Antal sidor267
Upplaga2002
FörlagHumana Press Inc.
ISBN9780896039346

Tillhör följande kategorier

Biologi inom Naturvetenskap och teknik
Biokemi inom Naturvetenskap och teknik

General Protocols.- The GTPase Cycle How Dominant Inhibitory Mutants Block the Biological Functions of Small GTPases.- Preparation of GTPases for Structural and Biophysical Analysis.- Using cDNA-Representational Difference Analysis (cDNA-RDA) in Combination with Microarrays to Identify Rac Regulated Genes.- Fluorescence Methods in the Study of Small GTP-Binding Proteins.- Specific Methods.- Ras and Rac as Activators of Reactive Oxygen Species (ROS).- Interfering with Ras Signaling Using Membrane-Permeable Peptides or Drugs.- A Ras-Based Module to Generate 32P-Labeled Fusion Proteins for Blot Overlays.- Determination of the Activity of Rho-Like GTPases in Cells.- of Dominant Inhibitory Proteins Directed Against ROK and MRCK Kinases.- Effects of Rho Family GTPases on Cell-Cell Adhesion.- Cell Motility and Invasion Assays.- Isolation of Regulatory Proteins for the Rab3 Subfamily GTPases.- Analysis and Preparation of Stable Complexes between Rab GTPases, Rab Escort Protein, and Rab Geranylgeranyl Transferase.- Preparation of Myristoylated Arf1 and Arf6 Proteins.- ARF-Directed Guanine-Nucleotide-Exchange (GEP) Proteins.- Arf6 and its Role in Cytoskeletal Modulation.- ARF GTPase-Activating Protein 1.- The Use of Permeabilized Cell Systems to Study Nuclear Transport.- Analysis of Nuclear Protein Import and Export In Vitro Using Fluorescent Cargoes.- Characterization of the Effects of RanGTP on the Microtubule Cytoskeleton.

From the reviews: "The 20 protocols provide detailed procedures for working with most of the known small GTPases of the RAS superfamily. Included are those from RAS, RHO, RAB, ARF, and RAN. In each section, several different approaches are given, including methods for studying downstream effects. Assay methods range from microarray analysis to cell motility measurements. In each case, there is sufficient detail to allow investigators new to the procedure to use them effectively. In addition, the limitations of the procedures are provided as well as useful "notes" to facilitate adoption in the laboratory. A short bibliography accompanies each chapter and provides adequate primary source references."-Doody's Health Sciences Book Review Journal "In GTPase Protocols, Manser and Leung assemble the key protocols required for researchers venturing into this field ... . combination of clear practical detail with underlying theory provides the reader with the information required not only to perform the technique but also to interpret the resulting data. ... In summary, this is a collection of detailed and precise experimental methods from some of the leading research groups in this field. The authors are to be congratulated on the clarity brought to each technique ... ." (Harry Mellor, Journal of Cell Science, Vol. 116 (7-8), 2003)