کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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1174541 | 961757 | 2011 | 9 صفحه PDF | دانلود رایگان |
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.
Journal: Analytical Biochemistry - Volume 412, Issue 1, 1 May 2011, Pages 47–55