Dynamic phosphometabolomic profiling of human tissues and transgenic models by 18O-assisted 31P NMR and mass spectrometry

Emirhan Nemutlu, Song Zhang, Anu Gupta, Nenad O. Juranic, Slobodan I. Macura, Andre Terzic, Arshad Jahangir, Petras Dzeja

Research output: Contribution to journalArticle

15 Scopus citations

Abstract

Nextgeneration screening of disease-related metabolomic phenotypes requires monitoring of both metabolite levels and turnover rates. Stable isotope 18O-assisted 31P nuclear magnetic resonance (NMR) and mass spectrometry uniquely allows simultaneous measurement of phosphometabolite levels and turnover rates in tissue and blood samples. The 18O labeling procedure is based on the incorporation of one 18O into P ifrom [ 18O]H 2O with each act of ATP hydrolysis and the distribution of 18O-labeled phosphoryls among phosphatecarrying molecules. This enables simultaneous recording of ATP synthesis and utilization, phosphotransfer fluxes through adenylate kinase, creatine kinase, and glycolytic pathways, as well as mitochondrial substrate shuttle, urea and Krebs cycle activity, glycogen turnover, and intracellular energetic communication. Application of expanded 18O-labeling procedures has revealed significant differences in the dynamics of G-6-P[ 18O] (glycolysis), G-3-P[ 18O] (substrate shuttle), and G-1-P[ 18O] (glycogenolysis) between human and rat atrial myocardium. In human atria, the turnover of G-3-P[ 18O], which defects are associated with the sudden death syndrome, was significantly higher indicating a greater importance of substrate shuttling to mitochondria. Phosphometabolomic profiling of transgenic hearts deficient in adenylate kinase (AK1-/-), which altered levels and mutations are associated to human diseases, revealed a stress-induced shift in metabolomic profile with increased CrP[ 18O] and decreased G-1-P[ 18O] metabolic dynamics. The metabolomic profile of creatine kinase M-CK/ScCKmit-/--deficient hearts is characterized by a higher G-6-[ 18O]P turnover rate, G-6-P levels, glycolytic capacity, γ/β-phosphoryl of GTP[ 18O] turnover, as well as β-[ 18O]ATP and β-[ 18O]ADP turnover, indicating altered glycolytic, guanine nucleotide, and adenylate kinase metabolic flux. Thus, 18O-assisted gas chromatographymass spectrometry and 31P NMR provide a suitable platform for dynamic phosphometabolomic profiling of the cellular energetic system enabling prediction and diagnosis of metabolic diseases states.

Original languageEnglish (US)
Pages (from-to)386-402
Number of pages17
JournalPhysiological Genomics
Volume44
Issue number7
DOIs
StatePublished - Apr 1 2012

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Keywords

  • 31P nuclear magnetic resonance
  • Adenylate kinase
  • Creatine kinase
  • Energy metabolism
  • Gas chromatography
  • Glycogenolysis
  • Glycolysis
  • Metabolic labeling
  • Metabolite turnover
  • Metabolomics
  • Mitochondria
  • Oxygen-18 stable isotopes
  • Phosphometabolomics
  • Phosphotransfer
  • System bioenergetics

ASJC Scopus subject areas

  • Physiology
  • Genetics

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