engleski

The effect of ganglioside composition on enzyme activity, protein expression, and submembrane localization of Na+/K+ -ATPase in mouse brain

The Na+ /K+ -ATPase (NKA) is an enzyme that asymmetrically distributes Na+ and K+ ions across the plasma membrane to generate and maintain the membrane potential, also having a role in signaling processes. NKA dysfunction has been implicated in several neurodegenerative disorders. Its positioning as well as different functions of active and inactive pools depend on interactions with neighboring membrane lipids. Ganglioside enriched lipid rafts (LR) are housing active NKA pool, while the bulk membrane contains the inactive pool. Gangliosides are known to modulate the structure, function and localization of membrane proteins thus having an impact on ion homeostasis. The aim of this study was to investigate the effect of altered ganglioside composition on activity, protein expression and submembrane localization of NKA, using St8sia1 null mice with impaired synthesis of gangliosides. Adult wild type (WT) and null mice littermates were sacrificed, brains neuroanatomically dissected and cortical and cerebellar homogenates prepared. NKA activity was measured spectrophotometrically. Protein expression of NKA in homogenates was analyzed by Western blotting. LR and non-raft (nLR) fractions from cortices and cerebella were isolated by ultracentrifugation in discontinuous sucrose gradients, and submembrane localization of NKA analyzed by Western blotting. Data revealed statistically lower NKA activity in cortices of null mice compared to the WT mice, whilst there was no disparity in the cerebella. Total protein amount of NKA was statistically lower in null mice cortices compared to their WTs, whilst it was unvaried in the cerebella. Analysis of submembrane localization has shown higher amount of NKA to be positioned within LR of cortices than those derived from the cerebella. These results demonstrate that altered ganglioside composition may contribute to lower NKA expression influencing NKA activity and cellular ion homeostasis.

glycosphingolipids ; sodium potassium pump ; sodium potassium ATPase

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