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Peroxynitrite production and tyrosine nitration are present in several pathological conditions, including neurodegeneration, stroke, aging, and cancer. Nitration of the pro-survival chaperone heat shock protein 90 (Hsp90) in position 33 and 56 induces motor neuron death through a toxic gain-of-function. Here we show that nitrated Hsp90 regulates mitochondrial metabolism independently of the induction of cell death. In PC12 cells, a small fraction of nitrated Hsp90 was located on the mitochondrial outer membrane and down-regulated mitochondrial membrane potential, oxygen consumption, and ATP production. Neither endogenous Hsp90 present in the homogenate nor unmodified and fully active recombinant Hsp90 was able to compete with the nitrated protein for the binding to mitochondria. Moreover, endogenous or recombinant Hsp90 did not prevent the decrease in mitochondrial activity but supported nitrated Hsp90 mitochondrial gain-of-function. Nitrotyrosine in position 33, but not in any of the other four tyrosine residues prone to nitration in Hsp90, was sufficient to down-regulate mitochondrial activity. Thus, in addition to induction of cell death, nitrated Hsp90 can also regulate mitochondrial metabolism, suggesting that depending on the cell type, distinct Hsp90 nitration states regulate different aspects of cellular metabolism. This regulation of mitochondrial homeostasis by nitrated Hsp90 could be of particular relevance in cancer cells. [ABSTRACT FROM AUTHOR]

Reduced brain metabolism is an invariant feature of Alzheimer Disease (AD) that is highly correlated to the decline in brain functions. Decreased activities of key tricarboxylic acid cycle (TA) cycle enzymes may underlie this abnormality and are highly correlated to the clinical state of the patient. The activity of the α-ketoglutarate dehydrogenase complex (KGDHC), an arguably rate-limiting enzyme of the TCA cycle, declines with AD, but the mechanism of inactivation and whether it can be reversed remains unknown. KGDHC consists of multiple copies of three subunits. KGDHC is sensitive to oxidative stress, which is pervasive in AD brain. The present studies tested the mechanism for the peroxynitrite-induced inactivation and subsequent reactivation of purified and cellular KGDHC. Peroxynitrite inhibited purified KGDHC activity in a dose-dependent manner and reduced subunit immunoreactivity and increased nitrotyrosine immunoreactivity. Nano-LC-MS/MS showed that the inactivation was related to nitration of specific tyrosine residues in the three subunits. GSH diminished the nitrotyrosine immunoreactivity of peroxynitrite-treated KGDHC, restored the activity and the immunoreactivity for KGDHC. Nano-LC-MS/MS showed this was related to de-nitration of specific tyrosine residues, suggesting KGDHC may have a denitrase activity. Treatment of N2a cells with peroxynitrite for 5 mm followed by recovery of cells for 24 h reduced KGDHC activity and increased nitrotyrosine immunoreactivity. Increasing cellular GSH in peroxynitrite-treated cells rescued KGDHC activity to the control level. The results suggest that restoring KGDHC activity is possible and may be a useful therapeutic approach in neurodegenerative diseases. [ABSTRACT FROM AUTHOR]