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Title: 3D TOCSY-HSQC NMR for metabolic flux analysis using non-uniform sampling

13C-Metabolic Flux Analysis ( 13C-MFA) is rapidly being recognized as the authoritative method for determining fluxes through metabolic networks. Site-specific 13C enrichment information obtained using NMR spectroscopy is a valuable input for 13C-MFA experiments. Chemical shift overlaps in the 1D or 2D NMR experiments typically used for 13C-MFA frequently hinder assignment and quantitation of site-specific 13C enrichment. Here we propose the use of a 3D TOCSY-HSQC experiment for 13C-MFA. We employ Non-Uniform Sampling (NUS) to reduce the acquisition time of the experiment to a few hours, making it practical for use in 13C-MFA experiments. Our data show that the NUS experiment is linear and quantitative. Identification of metabolites in complex mixtures, such as a biomass hydrolysate, is simplified by virtue of the 13C chemical shift obtained in the experiment. In addition, the experiment reports 13C-labeling information that reveals the position specific labeling of subsets of isotopomers. As a result, the information provided by this technique will enable more accurate estimation of metabolic fluxes in larger metabolic networks.
Authors:
 [1] ;  [2] ;  [3] ;  [4] ;  [1]
  1. Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
  2. Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Washington State Univ., Pullman, WA (United States)
  3. Miami Univ., Oxford, OH (United States)
  4. Duke Univ. Medical Center, Durham, NC (United States)
Publication Date:
Report Number(s):
PNNL-SA-114619
Journal ID: ISSN 0003-2700; 48535; KP1704020
Grant/Contract Number:
AC05-76RL01830
Type:
Published Article
Journal Name:
Analytical Chemistry
Additional Journal Information:
Journal Volume: 88; Journal Issue: 5; Journal ID: ISSN 0003-2700
Publisher:
American Chemical Society (ACS)
Research Org:
Pacific Northwest National Lab. (PNNL), Richland, WA (United States)
Sponsoring Org:
USDOE Office of Science (SC), Biological and Environmental Research (BER) (SC-23)
Country of Publication:
United States
Language:
English
Subject:
37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CHEMISTRY; Environmental Molecular Sciences Laboratory
OSTI Identifier:
1239174
Alternate Identifier(s):
OSTI ID: 1243206

Reardon, Patrick N., Marean-Reardon, Carrie L., Bukovec, Melanie A., Coggins, B. E., and Isern, Nancy G.. 3D TOCSY-HSQC NMR for metabolic flux analysis using non-uniform sampling. United States: N. p., Web. doi:10.1021/acs.analchem.5b04535.
Reardon, Patrick N., Marean-Reardon, Carrie L., Bukovec, Melanie A., Coggins, B. E., & Isern, Nancy G.. 3D TOCSY-HSQC NMR for metabolic flux analysis using non-uniform sampling. United States. doi:10.1021/acs.analchem.5b04535.
Reardon, Patrick N., Marean-Reardon, Carrie L., Bukovec, Melanie A., Coggins, B. E., and Isern, Nancy G.. 2016. "3D TOCSY-HSQC NMR for metabolic flux analysis using non-uniform sampling". United States. doi:10.1021/acs.analchem.5b04535.
@article{osti_1239174,
title = {3D TOCSY-HSQC NMR for metabolic flux analysis using non-uniform sampling},
author = {Reardon, Patrick N. and Marean-Reardon, Carrie L. and Bukovec, Melanie A. and Coggins, B. E. and Isern, Nancy G.},
abstractNote = {13C-Metabolic Flux Analysis (13C-MFA) is rapidly being recognized as the authoritative method for determining fluxes through metabolic networks. Site-specific 13C enrichment information obtained using NMR spectroscopy is a valuable input for 13C-MFA experiments. Chemical shift overlaps in the 1D or 2D NMR experiments typically used for 13C-MFA frequently hinder assignment and quantitation of site-specific 13C enrichment. Here we propose the use of a 3D TOCSY-HSQC experiment for 13C-MFA. We employ Non-Uniform Sampling (NUS) to reduce the acquisition time of the experiment to a few hours, making it practical for use in 13C-MFA experiments. Our data show that the NUS experiment is linear and quantitative. Identification of metabolites in complex mixtures, such as a biomass hydrolysate, is simplified by virtue of the 13C chemical shift obtained in the experiment. In addition, the experiment reports 13C-labeling information that reveals the position specific labeling of subsets of isotopomers. As a result, the information provided by this technique will enable more accurate estimation of metabolic fluxes in larger metabolic networks.},
doi = {10.1021/acs.analchem.5b04535},
journal = {Analytical Chemistry},
number = 5,
volume = 88,
place = {United States},
year = {2016},
month = {2}
}