Simulation of malate and citrate isotopologues and their dependence on Citrate-Malate Shuttle (CMS) activity or anaplerosis

Published: 12 April 2022| Version 1 | DOI: 10.17632/nhp95cx53y.1


This dataset contains supplementary data to the article entitled: “Krebs Takes a Turn at Cell Differentiation”, by Stanislaw Deja, Peter Crawford and Shawn Burgess. The supplementary data contain metabolic flux analysis simulations of Malate and Citrate isotopologues, as well as the Malate M+2 / Citrate M+2 ratio during [U-13C6]glucose metabolism across a range of anaplerotic and Citrate-Malate Shuttle (CMS) flux. Details of the metabolic flux analysis (MFA) model are provided. For a given malate M+2 / citrate M+2 ratio there are innumerable combinations of solutions for dilution by anaplerosis and Citrate-Malate Shuttle activity. For a stable level of anaplerosis, malate M+2 / citrate M+2 ratio will decrease with the increasing activity of CMS, although in nonlinear fashion.


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Simulations were conducted in INCA 2.0 ( and graphs were generated in MATLAB using in house routines. The Metabolic Flux Analysis (MFA) model was constructed to match the proposed model by Arnold et al., 2022 (PMID: 35264789). The model was fixed to 40% [U-13C]pyruvate labeling which was found to generate mass isotoplogue distributions similar to experimentally measured data. Citrate Synthase (CS) flux was fixed to 100. Citrate partitioning between the Tricarboxylic acid (TCA) cycle and Citrate-Malate Shuttle (CMS) was simulated using values between 0-100%, where 0 indicates that all citrate is oxidized in the TCA cycle and 100 indicates that all citrate is diverted to the CMS. For each citrate partitioning value, variable rates of anaplerotic flux were simulated (as indicated in the legend); where the rate is expressed relative to CS. Model was constructed with following assumptions: 1) CS is the only source of citrate formation 2) PDH is the only source of mitochondrial acetyl-CoA 3) Pyruvate is diluted to maintain acetyl-CoA enrichment = 40% 4) CO2 is unbalanced 5) TCA cycle reactions are simulated in the forward direction 6) Fumarate is defined as a symmetric molecule 7) All anaplerosis is unlabeled and originates from glutamate 8) Anaplerotic carbon is removed from the TCA cycle by generic cataplerotic reaction at the oxaloacetate node 9) When CMS operates, all cytosolic oxaloacetate generated by ACL is returned back to the mitochondria in the form of malate


University of Texas Southwestern Medical Center at Dallas


Mitochondrial Metabolism, Metabolic Flux Analysis, Metabolism, Metabolic Flux, Stable Isotope