One scaffold, two conformations: The ring-flip of the messenger InsP8 occurs under cytosolic conditions

Inositol poly- and pyrophosphates (InsPs and PP-InsPs) are central eukaryotic messengers. These very highly phosphorylated molecules can exist in two distinct conformations, a canonical one with five phosphoryl groups in equatorial positions, and a “flipped” axial conformation. Using ¹³C-labeled InsPs / PP-InsPs, the behavior of these molecules was investigated by 2D-NMR under solution conditions reminiscent of a cytosolic environment. Remarkably, the most highly phosphorylated messenger InsP₈ readily adopts both conformations at physiological conditions. Environmental factors - such as pH, metal cation composition, and temperature - strongly influence the conformational equilibrium. Thermodynamic data revealed that the transition of InsP₈ from the equatorial to the axial conformation is, in fact, an exothermic process. The speciation of InsPs and PP-InsPs also affects their interaction with protein binding partners; addition of Mg²⁺ decreased the binding constant K_d of InsP₈ to an SPX protein domain. The results illustrate that PP-InsP speciation reacts very sensitively to solution conditions, suggesting it might act as an environment-dependent molecular switch.