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Corporate Overview

IsoTopics™ – November 2016

Optimal Tracers for Parallel Labeling Experiments and 13C Metabolic Flux Analysis: A New Precision and Synergy Scoring System

Antoniewicz MR and colleagues have developed a new scoring scheme to identify optimal 13C tracers in single and parallel labeling experiments of metabolic flux analysis (MFA) studies. The scheme is based on precision and synergistic metrics that, for the first time, accounts for non-linear flux intervals and avoids biases due to flux value normalization. The metrics were evaluated against hundreds of labeling schemes through extensive in silico simulations and in vivo labeling experiments of E. coli measured by GC-MS. Matched findings revealed the double labeled glucose tracers (1,6-13C,) and (1,2-13C,) to be optimal for MFA of E. coli.



13C-Metabolic flux analysis (13C-MFA) is a widely used approach in metabolic engineering for quantifying intracellular metabolic fluxes. The precision of fluxes determined by 13C-MFA depends largely on the choice of isotopic tracers and the specific set of labeling measurements. A recent advance in the field is the use of parallel labeling experiments for improved flux precision and accuracy. However, as of today, no systemic methods exist for identifying optimal tracers for parallel labeling experiments. In this contribution, we have addressed this problem by introducing a new scoring system and evaluating thousands of different isotopic tracer schemes. Based on this extensive analysis we have identified optimal tracers for 13C-MFA. The best single tracers were doubly 13C-labeled glucose tracers, including [1,6-13C]glucose, [5,6-13C]glucose and [1,2-13C]glucose, which consistently produced the highest flux precision independent of the metabolic flux map (here, 100 random flux maps were evaluated). Moreover, we demonstrate that pure glucose tracers perform better overall than mixtures of glucose tracers. For parallel labeling experiments the optimal isotopic tracers were [1,6-13C]glucose and [1,2-13C]glucose. Combined analysis of [1,6-13C]glucose and [1,2-13C]glucose labeling data improved the flux precision score by nearly 20-fold compared to widely use tracer mixture 80% [1-13C]glucose +20% [U-13C]glucose.

Crown SB, Long CP, Antoniewicz MR

Reat article



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