Issue 20, 2019

Harnessing 13C-labeled myo-inositol to interrogate inositol phosphate messengers by NMR

Abstract

Inositol poly- and pyrophosphates (InsPs and PP-InsPs) are an important group of metabolites and mediate a wide range of processes in eukaryotic cells. To elucidate the functions of these molecules, robust techniques for the characterization of inositol phosphate metabolism are required, both at the biochemical and the cellular level. Here, a new tool-set is reported, which employs uniformly 13C-labeled compounds ([13C6]myo-inositol, [13C6]InsP5, [13C6]InsP6, and [13C6]5PP-InsP5), in combination with commonly accessible NMR technology. This approach permitted the detection and quantification of InsPs and PP-InsPs within complex mixtures and at physiological concentrations. Specifically, the enzymatic activity of IP6K1 could be monitored in vitro in real time. Metabolic labeling of mammalian cells with [13C6]myo-inositol enabled the analysis of cellular pools of InsPs and PP-InsPs, and uncovered high concentrations of 5PP-InsP5 in HCT116 cells, especially in response to genetic and pharmacological perturbation. The reported method greatly facilitates the analysis of this otherwise spectroscopically silent group of molecules, and holds great promise to comprehensively analyze inositol-based signaling molecules under normal and pathological conditions.

Graphical abstract: Harnessing 13C-labeled myo-inositol to interrogate inositol phosphate messengers by NMR

Supplementary files

Article information

Article type
Edge Article
Submitted
10 jan. 2019
Accepted
17 apr. 2019
First published
23 apr. 2019
This article is Open Access

All publication charges for this article have been paid for by the Royal Society of Chemistry
Creative Commons BY-NC license

Chem. Sci., 2019,10, 5267-5274

Harnessing 13C-labeled myo-inositol to interrogate inositol phosphate messengers by NMR

R. K. Harmel, R. Puschmann, M. Nguyen Trung, A. Saiardi, P. Schmieder and D. Fiedler, Chem. Sci., 2019, 10, 5267 DOI: 10.1039/C9SC00151D

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