Data for Intramolecular Carbon Isotope Fractionations Associated with Glucose Metabolism in Escherichia coli

To better understand metabolic controls on carbon isotope patterns within acetogenic lipids and their building blocks, we determined intramolecular δ13C values of acetate excreted from E. coli grown on glucose at different growth stages. These analyses revealed a stepwise reversal in the isotopic difference between the carboxyl and methyl carbons of acetate sampled in exponential versus stationary growth. Optimization of a method using GC-Orbitrap mass spectrometry enabled sufficiently accurate and precise measurements of low mM concentrations for natural abundance isotope ratios. To aid in interpretation, an isotope model for E. coli central metabolism using glucose was run with varying relative carbon fluxes through pyruvate dehydrogenase (ƒPDH). The model was also modified to account for acetate re-uptake through the phosphate transacetylase/acetate kinase (pta-ackA) pathway. Models employed carbon flux and enzymatic fractionation data from published studies and the QIRN model framework.

During exponential growth, extracellular acetate’s carboxyl carbon was depleted in 13C by -11.3 ± 1.9‰ relative to its methyl carbon. This difference is consistent with a high relative carbon flux commitment from pyruvate to acetate, as modeled with ƒPDH = 0.89. However, as glucose was further consumed and acetate accumulated, intramolecular patterns reversed to +7.8± 1.9‰, representing 13C enrichment in the carboxyl carbon relative to the methyl carbon. This intramolecular isotope reversal cannot be accounted for by changes in ƒPDH. Instead, our data and modeling illustrate that it resulted from a reversible “acetate switch” metabolism, which has implications for understanding isotope patterns within larger microbial biomolecules synthesized from acetyl-CoA.