Article ID Journal Published Year Pages File Type
1174541 Analytical Biochemistry 2011 9 Pages PDF
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.

Related Topics
Physical Sciences and Engineering Chemistry Analytical Chemistry
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