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

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

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

We recently developed a method for estimating protein dynamics in vivo with heavy water ( 2H 2O) 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 2H 2O.

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

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Labeling
Mass spectrometry
Mass Spectrometry
Amino Acids
Body Water
Proteins
Amino Acids, Peptides, and Proteins
Peptides
Water
Product Labeling
Deuterium Oxide
Cyclotrons
Protein Precursors
Deuterium
Fourier Analysis
Cyclotron resonance
Nutritional Status
Albumins
Lasers
Ionization

Keywords

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

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology
  • Cell Biology

Cite this

Measuring protein synthesis using metabolic 2H labeling, high-resolution mass spectrometry, and an algorithm. / Kasumov, Takhar; Ilchenko, Serguey; Li, Ling; Rachdaoui, Nadia; Sadygov, Rovshan; Willard, Belinda; McCullough, Arthur J.; Previs, Stephen.

In: Analytical Biochemistry, Vol. 412, No. 1, 01.05.2011, p. 47-55.

Research output: Contribution to journalArticle

Kasumov, T, Ilchenko, S, Li, L, Rachdaoui, N, Sadygov, R, Willard, B, McCullough, AJ & Previs, S 2011, 'Measuring protein synthesis using metabolic 2H labeling, high-resolution mass spectrometry, and an algorithm', Analytical Biochemistry, vol. 412, no. 1, pp. 47-55. https://doi.org/10.1016/j.ab.2011.01.021
Kasumov, Takhar ; Ilchenko, Serguey ; Li, Ling ; Rachdaoui, Nadia ; Sadygov, Rovshan ; Willard, Belinda ; McCullough, Arthur J. ; Previs, Stephen. / Measuring protein synthesis using metabolic 2H labeling, high-resolution mass spectrometry, and an algorithm. In: Analytical Biochemistry. 2011 ; Vol. 412, No. 1. pp. 47-55.
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