Analyses of inositol phosphates and phosphoinositides by strong anion exchange (SAX)-HPLC

Laha, D and Kamleitner, M and Johnen, P and Schaaf, G (2021) Analyses of inositol phosphates and phosphoinositides by strong anion exchange (SAX)-HPLC. In: Methods in Molecular Biology, 2295 . pp. 365-378.

PDF Methods_Mol_Biol_2295_365_2021.pdf - Published Version Restricted to Registered users only Download (354kB) | Request a copy

Abstract

The phosphate esters of myo-inositol (Ins) occur ubiquitously in biology. These molecules exist as soluble or membrane-resident derivatives and regulate a plethora of cellular functions including phosphate homeostasis, DNA repair, vesicle trafficking, metabolism, cell polarity, tip-directed growth, and membrane morphogenesis. Phosphorylation of all inositol hydroxyl groups generates phytic acid (InsP6), the most abundant inositol phosphate present in eukaryotic cells. However, phytic acid is not the most highly phosphorylated naturally occurring inositol phosphate. Specialized small molecule kinases catalyze the formation of the so-called myo-inositol pyrophosphates (PP-InsPs), such as InsP7 and InsP8. These molecules are characterized by one or several �high-energy� diphosphate moieties and are ubiquitous in eukaryotic cells. In plants, PP-InsPs play critical roles in immune responses and nutrient sensing. The detection of inositol derivatives in plants is challenging. This is particularly the case for inositol pyrophosphates because diphospho bonds are labile in plant cell extracts due to high amounts of acid phosphatase activity. We present two steady-state inositol labeling-based techniques coupled with strong anion exchange (SAX)-HPLC analyses that allow robust detection and quantification of soluble and membrane-resident inositol polyphosphates in plant extracts. These techniques will be instrumental to uncover the cellular and physiological processes controlled by these intriguing regulatory molecules in plants. © Springer Science+Business Media, LLC, part of Springer Nature 2021.

Item Type:	Journal Article
Publication:	Methods in Molecular Biology
Publisher:	Humana Press Inc.
Additional Information:	The copyright for this article belongs to Humana Press Inc.
Department/Centre:	Division of Biological Sciences > Biochemistry
Date Deposited:	18 Aug 2021 10:34
Last Modified:	19 Aug 2021 06:07
URI:	http://eprints.iisc.ac.in/id/eprint/69278

Actions (login required)

View Item