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• W1992048153 abstract "Methionine is an essential proteogenic amino acid. In addition, it is a methyl donor for DNA and protein methylation and a propylamine donor for polyamine biosynthesis. Both the methyl and propylamine donation pathways involve metabolic cycles, and methods are needed to quantitate these cycles. Here, we describe an analytical approach for quantifying methionine metabolic fluxes that accounts for the mixing of intracellular and extracellular methionine pools. We observe that such mixing prevents isotope tracing experiments from reaching the steady state due to the large size of the media pools and hence precludes the use of standard stationary metabolic flux analysis. Our approach is based on feeding cells with (13)C methionine and measuring the isotope-labeling kinetics of both intracellular and extracellular methionine by liquid chromatography-mass spectrometry (LC-MS). We apply this method to quantify methionine metabolism in a human fibrosarcoma cell line and study how methionine salvage pathway enzyme methylthioadenosine phosphorylase (MTAP), frequently deleted in cancer, affects methionine metabolism. We find that both transmethylation and propylamine transfer fluxes amount to roughly 15% of the net methionine uptake, with no major changes due to MTAP deletion. Our method further enables the quantification of flux through the pro-tumorigenic enzyme ornithine decarboxylase, and this flux increases 2-fold following MTAP deletion. The analytical approach used to quantify methionine metabolic fluxes is applicable for other metabolic systems affected by mixing of intracellular and extracellular metabolite pools." @default.
• W1992048153 created "2016-06-24" @default.
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• W1992048153 date "2014-01-16" @default.
• W1992048153 modified "2023-09-30" @default.
• W1992048153 title "Quantitation of Cellular Metabolic Fluxes of Methionine" @default.
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• W1992048153 doi "https://doi.org/10.1021/ac4032093" @default.
• W1992048153 hasPubMedCentralId "https://www.ncbi.nlm.nih.gov/pmc/articles/4060246" @default.
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