(507b) Complete-MFA: Complementary Parallel Labeling Experiments Technique for Metabolic Flux Analysis

Metabolic flux analysis (MFA) has emerged as a practical tool for measuring fluxes in complex biological systems. In ¹³C-MFA, fluxes are obtained by introducing a labeled tracer into a cell culture at metabolic steady state, measuring the relative labeling of metabolic intermediates and by-products at isotopic steady-state, and computationally processing these measurements to quantify fluxes. In this work, we describe a novel, more advanced method for ¹³C-MFA that we called COMPLETE-MFA, which stands for complementary parallel labeling experiments technique for high-resolution metabolic flux analysis.

The COMPLETE-MFA approach was developed specifically for high-resolution flux measurements. Until now, metabolic fluxes have been estimated from single tracer experiments. However, it is well know that in realistic network models there is often not one optimal isotopic tracer for determining all fluxes at high resolution. The COMPLETE-MFA method that we present here relies on using complementary isotopic tracers in parallel cell cultures, where each tracer is optimal for a specific part of metabolism. While each tracer may be suboptimal for elucidating the complete network model, we demonstrate that combined analysis of the parallel labeling experiments by simultaneously fitting all data sets to a single flux model provides flux results that are superior compared to any single tracer experiment. As proof of concept, we have quantified high-resolution fluxes for wild-type E. coli in batch culture. Using our COMPLETE-MFA approach we have resolved metabolic fluxes with 10-fold better precision (i.e. smaller confidence intervals) compared to current best method.