Purification and characterization of NADP-isocitrate dehydrogenase from skeletal muscle of Urocitellus richardsonii

NADP-dependent isocitrate dehydrogenase (NADP-IDH, EC 1.1.1.42) catalyzes the oxidative decarboxylation of isocitrate to alpha-ketoglutarate with the concomitant production of NADPH. NADPH plays important roles in many biosynthesis pathways, maintenance of proper oxidation-reduction balance, and protection against oxidative damage. This present study investigated the dynamic nature of NADP-IDH during hibernation by purifying it from the skeletal muscle of Richardson's ground squirrel (Urocitellus richardsonii) and analyzing its structural and functional changes in response to hibernation. Kinetic parameters of purified NADP-IDH from euthermic and hibernating ground squirrel skeletal muscle were characterized at 22 degrees C and 5 degrees C. Relative to euthermic muscle, -NADP-IDH in hibernating muscle had a higher affinity for its substrate, isocitrate at 22 degrees C, whereas at 5 degrees C, there was a significant decrease in isocitrate affinity. Western blot analysis revealed greater serine and threonine phosphorylation in hibernator NADP-IDH as compared to euthermic NADP-IDH. In addition, Bioinformatic analysis predicted the presence of 18 threonine and 21 serine phosphorylation sites on squirrel NADP-IDH. The structural and functional changes in NADP-IDH indicate the ability of the organism to reduce energy consumption during hibernation, while emphasizing increased NADPH production, and thus antioxidant activity, during torpor arousal cycles.

Automated summary

This study investigated the dynamic nature of NADP-IDH during hibernation, revealing increased substrate affinity and protein phosphorylation levels. These changes suggest the ability of the organism to reduce energy consumption and enhance antioxidant activity during hibernation.