LC-MS/MS measurements of ketoacids isotopologues distribution in HeLa cells after 1 day of intramitochondrial pyruvate influx by Grubraw

We analysed 13C metabolic inclusion into ketoacids in HeLa cell lines stably expressing Pseudomonas aeruginosa DadA (FAD-dependent D-amino acid dehydrogenase) gene with mitochondrial targeting (named Grubraw) after the addition of 12C D-alanine comparing to no D-alanine. We used HeLa with functional DadA and mutated DadA for control. In cells expressing functionally active DadA, D-alanine generates additional influx of intra-mitochondrial pyruvate. If cells grows on labelled carbone source and unlabeled D-alanine is used, this upcoming pyruvate shifts original mass-isotopologue distribution that can be registered by LC-MS.

Our cells were cultured for in a medium with labeled glucose or glutamine. To obtain current data, we changed growth media to pure one, added D-alanine, and incubated cells under these conditions for 24 hours before extraction. Using this scheme, we focused on a long-time effects of Grubraw activation.

We publish original RAW data from Orbitrap mass-spectrometer of intra- and extracellular ketoacid analysis. In “G” folders we put experiments with 1,2-[13]C-glucose and unlabelled glutamine. In “Q” folders, there are measurements with 5-[13]C-glutamine and unlabelled glucose. All files are named in a similar way. C12/C13 in the beginning indicates using of isotopically labeled carbon source or non-labeled one. “M”/“D” at the second position indicate using a cell line with mutated (M) or functional (D) DadA. “Plus”/“minus” indicates the addition of D-alanine. First number is a biological replicate, second number - technical replicate.