Quantitative peptidomics of pituitary glands from mice deficient in copper transport.

F. Y. Chei, B. A. Eipper, R. E. Mains, Lloyd D. Fricker

Research output: Contribution to journalArticle

22 Citations (Scopus)

Abstract

We previously described a method of quantitating levels of peptides in Cpe(fat)/Cpe(fat) mice using affinity chromatography to isolate peptide-processing intermediates and differential isotopic labeling/mass spectrometry. In the present study, we compared two different isotopic labels, acetic anhydride and succinic anhydride for detection and quantitation of peptides in wild type mice. As previously found for acetic anhydride, succinic anhydride efficiently labels all primary amines in various peptides. Of these two reagents, succinic anhydride provides better resolution between the heavy and light peaks of the labelled peptides due to a greater mass difference between the deuterated (heavy) and non-deuterated (light) form of this label (4 Da for succinate, 3 Da for acetate). Using succinic anhydride labeling, the accuracy of measuring 1:1 and 1:2 ratios of peptides in pituitary extracts was within 5% of the theoretical value for most peptides. The accuracy with succinic anhydride is comparable to the accuracy of acetic anhydride and more peptides could be detected and quantitated with succinic anhydride. The two labels were then used to examine pituitary peptides in mice with a defect in copper transport (Atp7a mice) vs wild type mice. Using succinic anhydride, 13 peptides could be detected, 12 of which matched the theoretical mass of known pituitary peptides. Five of the six peptides which contain C-terminal amide groups were significantly decreased in the Atp7a mice relative to wild type mice, whereas only one non-amidated peptide was significantly decreased in Atp7a mice. With acetic anhydride, only five peptides could be quantitated. The three peptides which contain C-terminal amide groups were decreased approximately 30% in the Atp7a mice. The selective decrease in amidated peptides in Atp7a mice is consistent with the copper-requirement of the enzyme that forms C-terminal amides.

Original languageEnglish (US)
Pages (from-to)713-722
Number of pages10
JournalCellular and molecular biology (Noisy-le-Grand, France)
Volume49
Issue number5
StatePublished - Jul 2003

Fingerprint

Pituitary Gland
Copper
Peptides
Labels
Amides
Labeling
Fats
Affinity chromatography
Light
Succinic Acid
succinic anhydride
Affinity Chromatography
Amines
Mass spectrometry

ASJC Scopus subject areas

  • Molecular Biology
  • Clinical Biochemistry
  • Cell Biology

Cite this

Quantitative peptidomics of pituitary glands from mice deficient in copper transport. / Chei, F. Y.; Eipper, B. A.; Mains, R. E.; Fricker, Lloyd D.

In: Cellular and molecular biology (Noisy-le-Grand, France), Vol. 49, No. 5, 07.2003, p. 713-722.

Research output: Contribution to journalArticle

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