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Kathleen A. McCann; David W. Williams; Edward E. McKee (Profiled Author: Edward E. McKee)
Journal of Neurochemistry. 2012;122(1):126-137.Abstract
The goal of this project was to characterize deoxypyrimidine salvage pathways used to maintain deoxynucleoside triphosphate pools in isolated brain mitochondria and to determine the extent that antiviral pyrimidine analogs utilize or affect these pathways. Mitochondria from rat brains were incubated in media with labeled and unlabeled deoxynucleosides and deoxynucleoside analogs. Products were analyzed by HPLC coupled to an inline UV monitor and liquid scintillation counter. Isolated mitochondria transported thymidine and deoxycytidine into the matrix, and readily phosphorylated both of these to mono-, di-, and tri-phosphate nucleotides. Rates of phosphorylation were much higher than rates observed in mitochondria from heart and liver. Deoxyuridine was phosphorylated much more slowly than thymidine and only to dUMP. 3'-azido-3'-deoxythymidine, zidovudine (AZT), an antiviral thymidine analog, was phosphorylated to AZT-MP as readily as thymidine was phosphorylated to TMP, but little if any AZT-DP or AZT-TP was observed. AZT at 5.5 ± 1.7 μM was shown to inhibit thymidine phosphorylation by 50%, but was not observed to inhibit deoxycytidine phosphorylation except at levels > 100 μM. Stavudine and lamivudine were inert when incubated with isolated brain mitochondria. The kinetics of phosphorylation of thymidine, dC, and AZT were significantly different in brain mitochondria compared to mitochondria from liver and heart. Fishing in the brain mitochondrial deoxynucleotide pools.Our goal was to characterize deoxypyrimidine salvage pathways used to maintain deoxynucleoside triphosphate pools in brain mitochondria and to determine the extent that antiviral pyrimidine analogs utilize or affect these pathways. We discovered very high thymidine kinase 2 activity with novel kinetics. Understanding the activation of pyrimidine analogs in the brain is crucial to understanding toxicity. © 2012 The Authors. Journal of Neurochemistry © 2012 International Society for Neurochemistry.
PMID: 22530558 PMCID: PMC3383789
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