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Selection of tracers for 13C-Metabolic Flux Analysis using Elementary Metabolite Units (EMU) basis vector methodology

Paper ID Volume ID Publish Year Pages File Format Full-Text
31584 44821 2012 12 PDF Available
Title
Selection of tracers for 13C-Metabolic Flux Analysis using Elementary Metabolite Units (EMU) basis vector methodology
Abstract

Metabolic flux analysis (MFA) is a powerful technique for elucidating in vivo fluxes in microbial and mammalian systems. A key step in 13C-MFA is the selection of an appropriate isotopic tracer to observe fluxes in a proposed network model. Despite the importance of MFA in metabolic engineering and beyond, current approaches for tracer experiment design are still largely based on trial-and-error. The lack of a rational methodology for selecting isotopic tracers prevents MFA from achieving its full potential. Here, we introduce a new technique for tracer experiment design based on the concept of elementary metabolite unit (EMU) basis vectors. We demonstrate that any metabolite in a network model can be expressed as a linear combination of so-called EMU basis vectors, where the corresponding coefficients indicate the fractional contribution of the EMU basis vector to the product metabolite. The strength of this approach is the decoupling of substrate labeling, i.e. the EMU basis vectors, from the dependence on free fluxes, i.e. the coefficients. In this work, we demonstrate that flux observability inherently depends on the number of independent EMU basis vectors and the sensitivities of coefficients with respect to free fluxes. Specifically, the number of independent EMU basis vectors places hard limits on how many free fluxes can be determined in a model. This constraint is used as a guide for selecting feasible substrate labeling. In three example models, we demonstrate that by maximizing the number of independent EMU basis vectors the observability of a system is improved. Inspection of sensitivities of coefficients with respect to free fluxes provides additional constraints for proper selection of tracers. The present contribution provides a fresh perspective on an important topic in metabolic engineering, and gives practical guidelines and design principles for a priori selection of isotopic tracers for 13C-MFA studies.

► A new technique for tracer experiment design based on elementary metabolite unit basis vectors. ► Any metabolite in a network model can be expressed as a linear combination of EMU basis vectors. ► The strength of the approach is decoupling of substrate labeling from dependence on fluxes. ► Flux observability inherently depends on the number of independent EMU basis vectors and sensitivities of coefficients. ► We provide practical guidelines and design principles for a priori selection of isotopic tracers for 13C-MFA.

Keywords
MFA, metabolic flux analysis; EMU, elementary metabolite unit; MS, mass spectrometry; MS/MS, tandem mass spectrometry; NMR, nuclear magnetic resonance; MID, mass isotopomer distribution; TCA, tricarboxylic acid cycle; SSR, variance-weighted sum of squared
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Publisher
Database: Elsevier - ScienceDirect
Journal: Metabolic Engineering - Volume 14, Issue 2, March 2012, Pages 150–161
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us