Inositol Phosphates, 1st ed. 2020
Methods and Protocols
Methods in Molecular Biology Series, Vol. 2091

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.

Part I: Isolation and Measurement

1. A Short Historical Perspective of Methods in Inositol Phosphate Research

Stephen B. Shears

2. Quantitation of Inositol Phosphates by HPLC-ESI-MS

Quynh H. Duong and Ronald B. Pegg

3. Isolation of Inositol Hexakisphosphate from Soils by Alkaline Extraction and Hypobromite Oxidation

Benjamin L. Turner

4. Detection of Inositol Phosphates by Split PH Domains

5. Dictyostelium discoideum as a Model to Study Inositol Pyrophosphate and Inorganic Polyphosphate

Yann Desfougères and Adolfo Saiardi

Part II: Assays for Function

6. Characterising Enzymes of the Diphosphoinositol Polyphosphate Phosphohydrolase (DIPP) Family

Lucinda Winward, Rajagopal Sharada Kilari, and Stephen T. Safrany

7. Metabolic Labeling of Inositol Phosphates and Phosphatidylinositols in Yeast and Mammalian Cells

Andrew T. Hale, Bradley P. Clarke, and John D. York

8. Back-pyrophosphorylation Assay to Detect in vivo InsP₇-dependent Protein Pyrophosphorylation in Mammalian Cells

Manasa Chanduri and Rashna Bhandari

9. Analyses of Ligand Binding to IP₃ receptors using Fluorescence Polarization

Ana M. Rossi and Colin W. Taylor

10. Investigating the InsP₃ Receptor in Living Cells by Caged InsP₃

Xin Hui and Peter Lipp

11. Optical Measurement of Synaptic Vesicle Fusion and its Inhibition by Inositol Pyrophosphate in Primary Cultured Hippocampal Neurons

Sung Hyun Kim

12. Competitive Fluorescent Ligand Assay for Inositol 1,4,5-Trisphosphate

Akihiko Tanimuraand Satoshi Shuto

13. Studying the Effects of Inositol Pyrophosphates in an In Vitro Vesicle-vesicle Fusion Assay

Mi Sook Sungand Tae-Young Yoon

Part III: Chemical Tools

14. Engineering Protease Resistant and Highly Active Phytases

Canfang Niu, Peilong Yang, and Bin Yao

15. Direct Cloning Method for Expression of Recombinant Proteins with an Inositol Hexakisphosphate Inducible Self-cleaving Tag

Keehwan Kwon, Marco Biancucci, Patrick J. Woida, and Karla J. F. Satchell

Includes cutting-edge methods and protocols

Provides step-by-step detail essential for reproducible results

Contains key notes and implementation advice from the experts