This volume covers the latest technological advances in the characterization of the biosynthesis and functions of Rab GTPases and their regulation by guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). The book consists of 28 chapters and starts with an overview of the Rab GTPase family. The next few chapters describe systematic approaches to the identification and classification of Rabs and Rab GAPs, as well as the detection of Rab isoprenylation and membrane distribution. The last few chapters examine the biochemical and functional properties of individual Rabs…mehr
This volume covers the latest technological advances in the characterization of the biosynthesis and functions of Rab GTPases and their regulation by guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs). The book consists of 28 chapters and starts with an overview of the Rab GTPase family. The next few chapters describe systematic approaches to the identification and classification of Rabs and Rab GAPs, as well as the detection of Rab isoprenylation and membrane distribution. The last few chapters examine the biochemical and functional properties of individual Rabs in the order of exocytic, recycling and endocytic Rabs. Written in the highly successful Methods of Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols and key tips on troubleshooting and avoiding known pitfalls.
Instructive and practical, Rab GTPases: Methods and Protocols approaches each topic with great detail and is a valuable resource for researchers and students interested in the field of Rab GTPases.
Guangpu Li, Ph.D. Department of Biochemistry and Molecular Biology University of Oklahoma Health Sciences Center 975 NE 10th Street, BRC 417B Oklahoma City, OK 73104 Tel: (405) 271-2227 ext. 61232 Fax: (405) 271-3910 E-mail: guangpu-li@ouhsc.edu
Inhaltsangabe
Rab Family of GTPases.- Bioinformatic Approaches to Identifying and Classifying Rab Proteins.- Rab-NANOPS: FRET-Biosensors for Rab Membrane Nano clustering and Prenylation Detection in Mammalian Cells.- High Throughput Assay for Profiling the Substrate Specificity of Rab GTPase-Activating Proteins.- Measuring Rab GTPase-Activating Protein (GAP) Activity in Live Cells and Extracts.- Analysis of the Interactions Between Rab GTPases and Class V Myosins.- Assaying the Interaction of the Rab Guanine Nucleotide Exchange Protein Sec2 with the Upstream Rab, a Downstream Effector and a Phosphoinositide.- Kinetic Activation of Rab8 Guanine Nucleotide Exchange Factor Rabin8 by Rab11.- Ypt1 and TRAPP Interactions: Optimization of Multi-Color Bimolecular Fluorescence Complementation in Yeast.- Identifying a Rab Effector on the Macro autophagy Pathway.- Functional Analysis of Rab27 and its Effector Slp-2a in Renal Epithelial Cells.- Small GTPases in Acrosomal Exocytosis.- Rab Antibody Characterization: Comparison of Rab14 Antibodies.- Selective Visualization of GLUT4 Storage Vesicles and Associated Rab Proteins Using IRAP-pHluorin.- 3D Time-Lapse Analysis of Rab11/FIP5 Complex: Spatiotemporal Dynamics During Apical Lumen Formation.- In Vitro and In Vivo Characterization of the Rab11-GAP Activity of Drosophila Evi5.- Characterization of the Role Rab25 in Energy Metabolism and Cancer Using Extracellular Flux Analysis and Material Balance.- Measurement of Rab35 Activity by the GTP-Rab35 Trapper RBD35.- Analysis of Connecdenn 1-3 (DENN1A-C) GEF Activity for Rab35.- Assay of Rab17 and its Guanine Nucleotide Exchange Factor Rabex-5 in Dendrites of Hippocampal Neurons.- Methods for Analysis of AP-3/Rabin4' in Regulation of Lysosome Distribution.- Determination of Rab5 Activity in the Cell by Effector Pull-Down Assay.- Identification of the Rab5 Binding Site in p110b - Assays for PI3Kb Binding to Rab5.- Role of the Rab5 Guanine Nucleotide Exchange Factor,Rme-6, in the Regulation of Clathrin-Coated Vesicle Uncoating.- Differential Effects of Overexpression of Rab5 and Rab22 on Autophagy in PC12 Cells With or Without NGF.- Determining the Role of Rab7 in Constitutive and Ligand-Mediated Epidermal Growth Factor Receptor Endocytic Trafficking Using Single Cell Assays.- Visualizing Directional Rab7 and TrkA Co-Trafficking in Axons by pTIRF Microscopy.- Quantitative Bead-Based Flow Cytometry for Assaying Rab7 GTPase Interaction with the Rab-Interacting Lysosomal Protein (RILP) Effector Protein.
Rab Family of GTPases.- Bioinformatic Approaches to Identifying and Classifying Rab Proteins.- Rab-NANOPS: FRET-Biosensors for Rab Membrane Nano clustering and Prenylation Detection in Mammalian Cells.- High Throughput Assay for Profiling the Substrate Specificity of Rab GTPase-Activating Proteins.- Measuring Rab GTPase-Activating Protein (GAP) Activity in Live Cells and Extracts.- Analysis of the Interactions Between Rab GTPases and Class V Myosins.- Assaying the Interaction of the Rab Guanine Nucleotide Exchange Protein Sec2 with the Upstream Rab, a Downstream Effector and a Phosphoinositide.- Kinetic Activation of Rab8 Guanine Nucleotide Exchange Factor Rabin8 by Rab11.- Ypt1 and TRAPP Interactions: Optimization of Multi-Color Bimolecular Fluorescence Complementation in Yeast.- Identifying a Rab Effector on the Macro autophagy Pathway.- Functional Analysis of Rab27 and its Effector Slp-2a in Renal Epithelial Cells.- Small GTPases in Acrosomal Exocytosis.- Rab Antibody Characterization: Comparison of Rab14 Antibodies.- Selective Visualization of GLUT4 Storage Vesicles and Associated Rab Proteins Using IRAP-pHluorin.- 3D Time-Lapse Analysis of Rab11/FIP5 Complex: Spatiotemporal Dynamics During Apical Lumen Formation.- In Vitro and In Vivo Characterization of the Rab11-GAP Activity of Drosophila Evi5.- Characterization of the Role Rab25 in Energy Metabolism and Cancer Using Extracellular Flux Analysis and Material Balance.- Measurement of Rab35 Activity by the GTP-Rab35 Trapper RBD35.- Analysis of Connecdenn 1-3 (DENN1A-C) GEF Activity for Rab35.- Assay of Rab17 and its Guanine Nucleotide Exchange Factor Rabex-5 in Dendrites of Hippocampal Neurons.- Methods for Analysis of AP-3/Rabin4' in Regulation of Lysosome Distribution.- Determination of Rab5 Activity in the Cell by Effector Pull-Down Assay.- Identification of the Rab5 Binding Site in p110b - Assays for PI3Kb Binding to Rab5.- Role of the Rab5 Guanine Nucleotide Exchange Factor,Rme-6, in the Regulation of Clathrin-Coated Vesicle Uncoating.- Differential Effects of Overexpression of Rab5 and Rab22 on Autophagy in PC12 Cells With or Without NGF.- Determining the Role of Rab7 in Constitutive and Ligand-Mediated Epidermal Growth Factor Receptor Endocytic Trafficking Using Single Cell Assays.- Visualizing Directional Rab7 and TrkA Co-Trafficking in Axons by pTIRF Microscopy.- Quantitative Bead-Based Flow Cytometry for Assaying Rab7 GTPase Interaction with the Rab-Interacting Lysosomal Protein (RILP) Effector Protein.
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