Comprehensive isotopomer analysis of glutamate and aspartate in small tissue samples

Feng Cai, Divya Bezwada, Ling Cai, Rohit Mahar, Zheng Wu, Mario C. Chang, Panayotis Pachnis, Chendong Yang, Sherwin Kelekar, Wen Gu, Bailey Brooks, Bookyung Ko, Hieu S. Vu, Thomas P. Mathews, Lauren G. Zacharias, Misty Martin-Sandoval, Duyen Do, K. Celeste Oaxaca, Eunsook S. Jin, Vitaly MargulisCraig R. Malloy, Matthew E. Merritt, Ralph J. DeBerardinis

Research output: Contribution to journalArticlepeer-review

2 Scopus citations

Abstract

Stable isotopes are powerful tools to assess metabolism. 13C labeling is detected using nuclear magnetic resonance (NMR) spectroscopy or mass spectrometry (MS). MS has excellent sensitivity but generally cannot discriminate among different 13C positions (isotopomers), whereas NMR is less sensitive but reports some isotopomers. Here, we develop an MS method that reports all 16 aspartate and 32 glutamate isotopomers while requiring less than 1% of the sample used for NMR. This method discriminates between pathways that result in the same number of 13C labels in aspartate and glutamate, providing enhanced specificity over conventional MS. We demonstrate regional metabolic heterogeneity within human tumors, document the impact of fumarate hydratase (FH) deficiency in human renal cancers, and investigate the contributions of tricarboxylic acid (TCA) cycle turnover and CO2 recycling to isotope labeling in vivo. This method can accompany NMR or standard MS to provide outstanding sensitivity in isotope-labeling experiments, particularly in vivo.

Original languageEnglish (US)
Pages (from-to)1830-1843.e5
JournalCell Metabolism
Volume35
Issue number10
DOIs
StatePublished - Oct 3 2023

Keywords

  • aspartate
  • cancer
  • glutamate
  • isotopomer
  • mass spectrometry
  • nuclear magnetic resonance
  • pyruvate carboxylase
  • stable isotope
  • tricarboxylic acid cycle

ASJC Scopus subject areas

  • Physiology
  • Molecular Biology
  • Cell Biology

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