Measuring protein synthesis using metabolic 2H labeling, high-resolution mass spectrometry, and an algorithm

Takhar Kasumov, Serguey Ilchenko, Ling Li, Nadia Rachdaoui, Rovshan G. Sadygov, Belinda Willard, Arthur J. McCullough, Stephen Previs

Research output: Contribution to journalArticle

42 Scopus citations

Abstract

We recently developed a method for estimating protein dynamics in vivo with heavy water (2H2O) using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) [16], and we confirmed that 2H labeling of many hepatic free amino acids rapidly equilibrated with body water. Although this is a reliable method, it required modest sample purification and necessitated the determination of tissue-specific amino acid labeling. Another approach for quantifying protein kinetics is to measure the 2H enrichments of body water (precursor) and protein-bound amino acid or proteolytic peptide (product) and to estimate how many copies of deuterium are incorporated into a product. In the current study, we used nanospray linear trap Fourier transform ion cyclotron resonance mass spectrometry (LTQ FT-ICR MS) to simultaneously measure the isotopic enrichment of peptides and protein-bound amino acids. A mathematical algorithm was developed to aid the data processing. The most notable improvement centers on the fact that the precursor/product labeling ratio can be obtained by measuring the labeling of water and a protein (or peptide) of interest, thereby minimizing the need to measure the amino acid labeling. As a proof of principle, we demonstrate that this approach can detect the effect of nutritional status on albumin synthesis in rats given 2H2O.

Original languageEnglish (US)
Pages (from-to)47-55
Number of pages9
JournalAnalytical Biochemistry
Volume412
Issue number1
DOIs
StatePublished - May 1 2011

Keywords

  • Albumin
  • Heavy water
  • High-resolution mass spectrometry
  • Isotopomers
  • Mass isotopomer distribution
  • Modeling
  • Protein synthesis
  • Rat

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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