Small GTPases play a key role in many aspects of contemporary cell biology: control of cell growth and differentiation; regulation of cell adhesion and cell movement; the organization of the actin cytoskeleton; and the regulation of intracellular vesicular transport. This volume plus its companions Volumes 256 and 257 cover all the biochemical and biological assays currently in use for analyzing the role of small GTPases in many aspects of cell biology at the molecular level.
General Description of the Series:
The critically acclaimed laboratory standard for more than forty years, Methods in Enzymology is one of the most highly respected publications in the field of biochemistry. Since 1955, each volume has been eagerly awaited, frequently consulted, and praised by researchers and reviewers alike. Now with more than 300 volumes (all of them still in print), the series contains much material still relevant today--truly an essential publication for researchers in all fields of life sciences.
- Expression, purification, and posttranslational modification
- Guanine nucleotide exchange and hydrolysis
- Cell expression and analysis in Vitro
- Biological activity
Guanine Nucleotide Exchange and Hydrolysis:C. Lenzen, R.H. Cool, and A. Wittinghofer, Analysis of Intrinsic and CDC25-Stimulated Guanine Nucleotide Exchange of p21ras--Nucleotide Complexes by Fluorescence Measurements.J. Downward, Measurement of Nucleotide Exchange and Hydrolysis Activities in Immunoprecipitates.J.B. Gibbs, Determination of Guanine Nucleotides Bound to Ras in Mammalian Cells.M. Frech, D. Cussac, P. Chardin, and D. Bar-Sagi, Purification of Baculovirus-Expressed Human Sos1 Protein.M. Hart and S. Powers, Ras-Cdc25 and Rho-Dbl Binding Assays: Complex Formation in Vitro.R.D. Mosteller, W. Park, and D. Broek, Analysis of Interaction between Ras and CDC25 Guanine Nucleotide Exchange Factor Using Yeast GAL4 Two-Hybrid System.T. Satoh and Y. Kaziro, Measurement of Ras-Bound Guanine Nucleotide in Stimulated Hematopoietic Cells.A.M.M. de Vries-Smits, L. van der Voorn, J. Downward, and J.L. Bos, Measurements of GTP/GDP Exchange in Permeabilized Fibroblasts.G.E. Bollag and F. McCormick, Intrinsic and GTPase-Activating Protein-Stimulated Ras GTPase Assays.M.D. Schaber and J.B. Gibbs, Determination of Ras and GTPase-Activating Protein Interactions by Kinetic Competition Assay.C. Ellis, V. Measday, and M.F. Moran, Phosphorylation-Dependent Complexes of p120 Ras-Specific GTPase-Activating Proteins with p62 and p190.
Cell Expression and Analysis:
in Vitro:A.D. Cox, P.A. Solski, J.D. Jordan, and C.J. Der, Analysis of Ras Protein Expression in Mammalian Cells.A.M.M. deVries-Smits, B.M.T. Burgering, and J.L. Bos, Vaccinia Virus Expression of p21rasAsn-17.H. Cai and G.M. Cooper, Inducible Expression of Ras N17 Dominant Inhibitory Protein.J.F. Hancock, Prenylation and Palmitoylation Analysis.C.W.M. Reuter, A.D. Catling, and M.J. Weber, Immune Complex Kinase Assays for Mitogen-Activated Protein Kinase and MEK.S. Kuroda, K. Shimizu, B. Yamamori, and Y. Takai, Cell-Free Assay System for Ras-Dependent MEK Activation.P. Dent, G. Romero, D. Castle, and T.W. Sturgill, Preparation and Use of Semiintact Mammalian Cells for Analysis of Signal Transduction.C.J. Marshall and S.J. Leevers, Mitogen-Activated Protein Kinase Activation by Scrape Loading of P21ras.D.R. Alessi, P. Cohen, A. Ashworth, S. Cowley, S.J. Leevers, and C.J. Marshall, Assay and Expression of Mitogen-Activated Protein Kinase, MAP Kinase Kinase, and Raf.C.A. Lange-Carter and G.L. Johnson, Assay of MEK Kinases.D.K. Morrison, Activation of Raf-1by Ras in Intact Cells.R. Finney and D. Herrera, Ras-Raf Complexes: Analyses of Complexes Formed in Vivo.X.-f. Zhang, M.S. Marshall, and J. Avruch, Ras-Raf Complexes in Vitro.A.B. Vojtek and S.M. Hollenberg, Ras-Raf Interaction: Two-Hybrid Analysis.J.K. Westwick and D.A. Brenner, Methods for Analyzing c-Jun Kinase.B. Margolis, E.Y. Skolnik, and J. Schlessinger, Use of Tyrosine-Phosphorylated Proteins to Screen Bacterial Expression Libraries for SH2 Domains.S.M. Feller, B. Knudsen, T.W. Wong, and H. Hanafusa, Detection of SH3-Binding Proteins in Total Cell Lysates with Glutathine S-Transferase-SH3 Fusion Proteins: SH3 Blot Assay.N.E. Kohl, F.R. Wilson, T.J. Thomas, R.L. Bock, S.D. Mosser, A. Oliff, and J.B. Gibbs, Inhibition of Ras Function in Vitro and in Vivo Using Inhibitors of Farnesyl-Protein Transferase.
Biological Activity:C. Finlay, Rat Embryo Fibroblast Complementation Assay with ras Genes.G.J. Clark, A.D. Cox, S.M. Graham, and C.J. Der, Biological Assays for Ras Transformation.C.A. Hauser, J.K. Westwick, and L.A. Quilliam, Ras-Mediated Transcription Activation: Analysis by Transient Cotransfection Assays.B. Tocque, M. Janicot, and M. Kenigsberg, Oocyte Microinjection Assay for Evaluation of Ras-Induced Signaling Pathways.D. Bar-Sagi, Mammalian Cell Microinjection Assays.A. Mikheev, R.S. Cha, and H. Zarbl, Detection of Point Mutations in Ras in Tumor Cell Lines by Denaturant Gradient Gel Electrophoresis.S.M. Kahn, W. Jiang, I.B. Weinstein, and M. Perucho, Diagnostic Detection of Mutant ras Genes in Minor Cell Populations.L. Chen and S. Powers, Ras in Yeast: Complementation Assays for Testof Function.K.A. Mintzer and J. Field, Yeast Adenylyl Cyclase Assays.M.T. Quinn, C.A. Parkos, and A.J. Jesaitis, Purification of Human Neutrophil NADPH Oxidase Cytochrome b-588 and Association with Rap1A.P. Poullet and F. Tamanoi, Use of Yeast Two-Hybrid System to Evaluate Ras Interactions with Neurofibromin-GTPase-Activating Protein.A.B. Sparks, N.B. Adey, L.A. Quilliam, J.M. Thorn, and B.K. Kay, Screening Phage-Displayed Random Peptide Libraries for SH3 Ligands.