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This volume describes modern approaches for the detection and analysis of inositol phosphates from various biological, clinical, and environmental samples. Chapters focus on methods to study enzymes that modify inositol signals, chemical biology of inositol phosphates, and methods to synthesize and purify inositol phosphates and their analogs. Written in the highly successful Methods in 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 tips on troubleshooting and avoiding known pitfalls.
Authoritative and cutting-edge, Inositol Phosphates: Methods and Protocol aims to ensure successful results in the further study of this vital field.
Sommario
A Short Historical Perspective of Methods in Inositol Phosphate Research.- Quantitation of Inositol Phosphates by HPLC-ESI-MS.- Isolation of Inositol Hexakisphosphate from Soils by Alkaline Extraction and Hypobromite Oxidation.- Detection of Inositol Phosphates by Split PH Domains.- Dictyostelium discoideum as a Model to Study Inositol Pyrophosphate and Inorganic Polyphosphate.- Characterising Enzymes of the Diphosphoinositol Polyphosphate Phosphohydrolase (DIPP) Family.- Metabolic Labeling of Inositol Phosphates and Phosphatidylinositols in Yeast and Mammalian Cells.- Back-pyrophosphorylation Assay to Detect in vivo InsP7-dependent Protein Pyrophosphorylation in Mammalian Cells.- Analyses of Ligand Binding to IP3 receptors using Fluorescence Polarization.- Investigating the InsP3 Receptor in Living Cells by Caged InsP3.- Optical Measurement of Synaptic Vesicle Fusion and its Inhibition by Inositol Pyrophosphate in Primary Cultured Hippocampal Neurons.- Competitive Fluorescent Ligand Assay for Inositol 1,4,5-Trisphosphate.- Studying the Effects of Inositol Pyrophosphates in an In Vitro Vesicle-vesicle Fusion Assay.- Engineering Protease Resistant and Highly Active Phytases.- Direct Cloning Method for Expression of Recombinant Proteins with an Inositol Hexakisphosphate Inducible Self-cleaving Tag.- A Short Historical Perspective of Methods in Inositol Phosphate Research.
