Determination of Fractional Synthesis Rates of Mouse Hepatic Proteins via Metabolic 13C-Labeling, MALDI-TOF MS and Analysis of Relative Isotopologue Abundances Using Average Masses
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- 作者：Josef A. Vogt ; Christian Hunzinger ; Klaus Schroer ; Kerstin Hö ; lzer ; Anke Bauer ; André ; Schrattenholz ; Michael A. Cahill ; Simone Schillo ; Gerhard Schwall ; Werner Stegmann ; and Gerd Albuszies
- 刊名：Analytical Chemistry
- 出版年：2005
- 出版时间：April 1, 2005
- 年：2005
- 卷：77
- 期：7
- 页码：2034 - 2042
- 全文大小：188K
- 年卷期：v.77,no.7(April 1, 2005)
- ISSN：1520-6882
文摘
Proteins of a liver extract taken from a metabolically 13C-labeled mouse were separated by 2D-PAGE and identifiedafter tryptic digestion by MALDI-TOF MS peptide massfingerprinting. 13C-Labeling of proteins was achieved byan infusion of U-13C-glucose, which is metabolized tolabeled nonessential amino acids. The labeling was analyzed using the relative isotopologue abundances of themeasured isotope pattern of tryptic peptides and quantified by their increase in the average molecular mass(
AVM). Fractional synthesis rates (FSR) of proteins weredetermined from corresponding peptides using measuredAVM values as well as AVM values deduced fromtRNA-precursor amino acid labeling, which in turn wasderived from proteins showing high 13C enrichments. The8-h FSR values of 43 proteins were determined to rangefrom 0 ± 0.6 to 95 ± 1%/8 h, with typical errors given asSEM values, which depend on the number of peptides ofa specific protein usable for calculation. The methoddemonstrates that FSR values as an indicator for proteinturnover in the liver proteome can be estimated withinnarrow error margins, providing baseline values fromwhich treatment-dependent deviations could be detectedwith high statistical certainty.
AVM). Fractional synthesis rates (FSR) of proteins weredetermined from corresponding peptides using measuredAVM values as well as AVM values deduced fromtRNA-precursor amino acid labeling, which in turn wasderived from proteins showing high 13C enrichments. The8-h FSR values of 43 proteins were determined to rangefrom 0 ± 0.6 to 95 ± 1%/8 h, with typical errors given asSEM values, which depend on the number of peptides ofa specific protein usable for calculation. The methoddemonstrates that FSR values as an indicator for proteinturnover in the liver proteome can be estimated withinnarrow error margins, providing baseline values fromwhich treatment-dependent deviations could be detectedwith high statistical certainty.