TY - JOUR
T1 - Expanding the Coverage of the Metabolome with Nitrogen-Based NMR
AU - Bhinderwala, Fatema
AU - Lonergan, Samantha
AU - Woods, Jade
AU - Zhou, Chunyi
AU - Fey, Paul D.
AU - Powers, Robert
N1 - Funding Information:
We thank Dr. Martha Morton, the Director of the Research Instrumentation Facility in the Department of Chemistry at the University of Nebraska-Lincoln for her assistance with the NMR experiments. This material is based upon work supported by the National Science Foundation under Grant Number (1660921). This work was supported in part by funding from the Redox Biology Center (P30 GM103335, NIGMS); the Nebraska Center for Integrated Biomolecular Communication (P20 GM113126, NIGMS), and the National Institutes of Health grant (P01 AI083211, NIAID) to PDF and RP. The research was performed in facilities renovated with support from the National Institutes of Health (RR015468-01). Any opinions, findings, and conclusions or recommendations expressed in this material are those of the author(s) and do not necessarily reflect the views of the National Science Foundation.
Publisher Copyright:
© 2018 American Chemical Society.
PY - 2018/4/3
Y1 - 2018/4/3
N2 - Isotopically labeling a metabolite and tracing its metabolic fate has provided invaluable insights about the role of metabolism in human diseases in addition to a variety of other issues. 13C-labeled metabolite tracers or unlabeled 1H-based NMR experiments are currently the most common application of NMR to metabolomics studies. Unfortunately, the coverage of the metabolome has been consequently limited to the most abundant carbon-containing metabolites. To expand the coverage of the metabolome and enhance the impact of metabolomics studies, we present a protocol for 15N-labeled metabolite tracer experiments that may also be combined with routine 13C tracer experiments to simultaneously detect both 15N- and 13C-labeled metabolites in metabolic samples. A database consisting of 2D 1H-15N HSQC natural-abundance spectra of 50 nitrogen-containing metabolites are also presented to facilitate the assignment of 15N-labeled metabolites. The methodology is demonstrated by labeling Escherichia coli and Staphylococcus aureus metabolomes with 15N1-ammonium chloride, 15N4-arginine, and 13C2-acetate. Efficient 15N and 13C metabolite labeling and identification were achieved utilizing standard cell culture and sample preparation protocols.
AB - Isotopically labeling a metabolite and tracing its metabolic fate has provided invaluable insights about the role of metabolism in human diseases in addition to a variety of other issues. 13C-labeled metabolite tracers or unlabeled 1H-based NMR experiments are currently the most common application of NMR to metabolomics studies. Unfortunately, the coverage of the metabolome has been consequently limited to the most abundant carbon-containing metabolites. To expand the coverage of the metabolome and enhance the impact of metabolomics studies, we present a protocol for 15N-labeled metabolite tracer experiments that may also be combined with routine 13C tracer experiments to simultaneously detect both 15N- and 13C-labeled metabolites in metabolic samples. A database consisting of 2D 1H-15N HSQC natural-abundance spectra of 50 nitrogen-containing metabolites are also presented to facilitate the assignment of 15N-labeled metabolites. The methodology is demonstrated by labeling Escherichia coli and Staphylococcus aureus metabolomes with 15N1-ammonium chloride, 15N4-arginine, and 13C2-acetate. Efficient 15N and 13C metabolite labeling and identification were achieved utilizing standard cell culture and sample preparation protocols.
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U2 - 10.1021/acs.analchem.7b04922
DO - 10.1021/acs.analchem.7b04922
M3 - Article
C2 - 29505241
AN - SCOPUS:85044996735
VL - 90
SP - 4521
EP - 4528
JO - Analytical Chemistry
JF - Analytical Chemistry
SN - 0003-2700
IS - 7
ER -