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.

Small GTPase binding proteins (GTPases) are an ancient group of proteins that play key roles in almost every aspect of cell biology, from cell proliferation to nuclear transport. In GTPase Protocols: The Ras Superfamily, 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.