Metabolically stable isotope labeling prior to electrophoretic protein separation reveals differences in fractional synthesis rates between mitochondrial aldehyde dehydrogenase isoforms

Michael Cahill, Josef A. Vogt, Jorg Servos, Wojciech Wozny, Gerhard P. Schwall, Karlfried Groebe, Andre Schrattenholz, Werner Stegmann

Research output: Contribution to journalArticle

1 Citation (Scopus)

Abstract

Living mice were subjected to whole body labeling by intravenous infusion of [(13)C]glucose as the sole carbon source. After 10 h infusion the mice were sacrificed, and liver proteins were separated by two-dimensional polyacrylamide gel electrophoresis. Five spots were found to contain mitochondrial aldehyde dehydrogenase (ALDH2) by matrix assisted time of flight mass spectrometry protein identification. By measuring the isotopologue mass distributions of peptide ions, and modeling the (13)C content of the precursor amino acid pool, the fractional synthesis rate of ALDH2 molecules synthesized during the labeling period was determined. One of the five spots was observed to have a five-fold higher fraction of (13)C-containing newly synthesized ALDH2 than the spot with the highest ALDH2 content, and contained more than 60% of newly synthesized ALDH2 although it accounted for less than 20% of the total ALDH2 detected. The total range in the fraction of (13)C-containing proteins between different ALDH2 spots approached 50-fold. The ability to quantitatively characterize different protein isoforms of biological origin for ALDH2 and other proteins from living animals provides new avenues for the exploration of protein function.
Original languageEnglish
Pages (from-to)67-70
Number of pages4
JournalJournal of Chromatography
Volume1161
Issue number1-2
DOIs
Publication statusPublished - 2007

Fingerprint

Isotope Labeling
Aldehyde Dehydrogenase
Isotopes
Labeling
Protein Isoforms
Proteins
Electrophoresis, Gel, Two-Dimensional
Protein C
Intravenous Infusions
Mass Spectrometry
Carbon
Ions
Amino Acids
Glucose
Peptides
Electrophoresis
Liver
Mass spectrometry
Animals
Mitochondrial Aldehyde Dehydrogenase

Cite this

Cahill, Michael ; Vogt, Josef A. ; Servos, Jorg ; Wozny, Wojciech ; Schwall, Gerhard P. ; Groebe, Karlfried ; Schrattenholz, Andre ; Stegmann, Werner. / Metabolically stable isotope labeling prior to electrophoretic protein separation reveals differences in fractional synthesis rates between mitochondrial aldehyde dehydrogenase isoforms. In: Journal of Chromatography. 2007 ; Vol. 1161, No. 1-2. pp. 67-70.
@article{94f4317d7a4a40009de3861822db9ea1,
title = "Metabolically stable isotope labeling prior to electrophoretic protein separation reveals differences in fractional synthesis rates between mitochondrial aldehyde dehydrogenase isoforms",
abstract = "Living mice were subjected to whole body labeling by intravenous infusion of [(13)C]glucose as the sole carbon source. After 10 h infusion the mice were sacrificed, and liver proteins were separated by two-dimensional polyacrylamide gel electrophoresis. Five spots were found to contain mitochondrial aldehyde dehydrogenase (ALDH2) by matrix assisted time of flight mass spectrometry protein identification. By measuring the isotopologue mass distributions of peptide ions, and modeling the (13)C content of the precursor amino acid pool, the fractional synthesis rate of ALDH2 molecules synthesized during the labeling period was determined. One of the five spots was observed to have a five-fold higher fraction of (13)C-containing newly synthesized ALDH2 than the spot with the highest ALDH2 content, and contained more than 60{\%} of newly synthesized ALDH2 although it accounted for less than 20{\%} of the total ALDH2 detected. The total range in the fraction of (13)C-containing proteins between different ALDH2 spots approached 50-fold. The ability to quantitatively characterize different protein isoforms of biological origin for ALDH2 and other proteins from living animals provides new avenues for the exploration of protein function.",
keywords = "Aldehyde Dehydrogenase/*biosynthesis/isolation, Animals, Electrophoresis, Gel, Two-Dimensional/*methods, Isoenzymes/*biosynthesis/isolation, Isotope Labeling, Mice, Inbred C57BL, Mice, Male, Mitochondria/*enzymology, Purification, Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization",
author = "Michael Cahill and Vogt, {Josef A.} and Jorg Servos and Wojciech Wozny and Schwall, {Gerhard P.} and Karlfried Groebe and Andre Schrattenholz and Werner Stegmann",
note = "Imported on 12 Apr 2017 - DigiTool details were: Journal title (773t) = Journal of Chromatography A. ISSNs: 0021-9673;",
year = "2007",
doi = "10.1016/j.chroma.2007.04.019",
language = "English",
volume = "1161",
pages = "67--70",
journal = "Journal of Chromatography",
issn = "0021-9673",
publisher = "Elsevier",
number = "1-2",

}

Metabolically stable isotope labeling prior to electrophoretic protein separation reveals differences in fractional synthesis rates between mitochondrial aldehyde dehydrogenase isoforms. / Cahill, Michael; Vogt, Josef A.; Servos, Jorg; Wozny, Wojciech; Schwall, Gerhard P.; Groebe, Karlfried; Schrattenholz, Andre; Stegmann, Werner.

In: Journal of Chromatography, Vol. 1161, No. 1-2, 2007, p. 67-70.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Metabolically stable isotope labeling prior to electrophoretic protein separation reveals differences in fractional synthesis rates between mitochondrial aldehyde dehydrogenase isoforms

AU - Cahill, Michael

AU - Vogt, Josef A.

AU - Servos, Jorg

AU - Wozny, Wojciech

AU - Schwall, Gerhard P.

AU - Groebe, Karlfried

AU - Schrattenholz, Andre

AU - Stegmann, Werner

N1 - Imported on 12 Apr 2017 - DigiTool details were: Journal title (773t) = Journal of Chromatography A. ISSNs: 0021-9673;

PY - 2007

Y1 - 2007

N2 - Living mice were subjected to whole body labeling by intravenous infusion of [(13)C]glucose as the sole carbon source. After 10 h infusion the mice were sacrificed, and liver proteins were separated by two-dimensional polyacrylamide gel electrophoresis. Five spots were found to contain mitochondrial aldehyde dehydrogenase (ALDH2) by matrix assisted time of flight mass spectrometry protein identification. By measuring the isotopologue mass distributions of peptide ions, and modeling the (13)C content of the precursor amino acid pool, the fractional synthesis rate of ALDH2 molecules synthesized during the labeling period was determined. One of the five spots was observed to have a five-fold higher fraction of (13)C-containing newly synthesized ALDH2 than the spot with the highest ALDH2 content, and contained more than 60% of newly synthesized ALDH2 although it accounted for less than 20% of the total ALDH2 detected. The total range in the fraction of (13)C-containing proteins between different ALDH2 spots approached 50-fold. The ability to quantitatively characterize different protein isoforms of biological origin for ALDH2 and other proteins from living animals provides new avenues for the exploration of protein function.

AB - Living mice were subjected to whole body labeling by intravenous infusion of [(13)C]glucose as the sole carbon source. After 10 h infusion the mice were sacrificed, and liver proteins were separated by two-dimensional polyacrylamide gel electrophoresis. Five spots were found to contain mitochondrial aldehyde dehydrogenase (ALDH2) by matrix assisted time of flight mass spectrometry protein identification. By measuring the isotopologue mass distributions of peptide ions, and modeling the (13)C content of the precursor amino acid pool, the fractional synthesis rate of ALDH2 molecules synthesized during the labeling period was determined. One of the five spots was observed to have a five-fold higher fraction of (13)C-containing newly synthesized ALDH2 than the spot with the highest ALDH2 content, and contained more than 60% of newly synthesized ALDH2 although it accounted for less than 20% of the total ALDH2 detected. The total range in the fraction of (13)C-containing proteins between different ALDH2 spots approached 50-fold. The ability to quantitatively characterize different protein isoforms of biological origin for ALDH2 and other proteins from living animals provides new avenues for the exploration of protein function.

KW - Aldehyde Dehydrogenase/biosynthesis/isolation

KW - Animals

KW - Electrophoresis, Gel, Two-Dimensional/methods

KW - Isoenzymes/biosynthesis/isolation

KW - Isotope Labeling

KW - Mice, Inbred C57BL

KW - Mice, Male

KW - Mitochondria/enzymology

KW - Purification

KW - Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization

U2 - 10.1016/j.chroma.2007.04.019

DO - 10.1016/j.chroma.2007.04.019

M3 - Article

VL - 1161

SP - 67

EP - 70

JO - Journal of Chromatography

JF - Journal of Chromatography

SN - 0021-9673

IS - 1-2

ER -