Identification of Single and Double Sites of Phosphorylation by ECD FT-ICR/MS in Peptides Related to the Phosphorylation Site Domain of the Myristoylated Alanine-Rich C Kinase Protein

Kellie A. Woodling, John R. Eyler, Yury O. Tsybin, Carol L. Nilsson, Alan G. Marshall, Arthur S. Edison, Iman M. Al-Naggar, Michael R. Bubb

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

A series of phosphorylated test peptides was studied by electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry (ECD FT-ICR MS). The extensive ECD-induced fragmentation made identification of phosphorylation sites for these peptides straightforward. The site(s) of initial phosphorylation of a synthetic peptide with a sequence identical to that of the phosphorylation site domain (PSD) of the myristoylated alanine-rich C kinase (MARCKS) protein was then determined. Despite success in analyzing fragmentation of the smaller test peptides, a unique site on the PSD for the first step of phosphorylation could not be identified because the phosphorylation reaction produced a heterogeneous mixture of products. Some molecules were phosphorylated on the serine closest to the N-terminus, and others on one of the two serines closest to the C-terminus of the peptide. Although no definitive evidence for phosphorylation on either of the remaining two serines in the PSD was found, modification there could not be ruled out by the ECD fragmentation data.

Original languageEnglish
Pages (from-to)2137-2145
Number of pages9
JournalJournal of the American Society for Mass Spectrometry
Volume18
Issue number12
DOIs
StatePublished - Dec 2007
Externally publishedYes

Fingerprint

Phosphorylation
Alanine
Protein Kinase C
Peptides
Serine
Cyclotrons
Cyclotron resonance
Fourier Analysis
Mass spectrometry
Mass Spectrometry
Fourier transforms
Electrons
Ions
Molecules

ASJC Scopus subject areas

  • Structural Biology
  • Spectroscopy

Cite this

Identification of Single and Double Sites of Phosphorylation by ECD FT-ICR/MS in Peptides Related to the Phosphorylation Site Domain of the Myristoylated Alanine-Rich C Kinase Protein. / Woodling, Kellie A.; Eyler, John R.; Tsybin, Yury O.; Nilsson, Carol L.; Marshall, Alan G.; Edison, Arthur S.; Al-Naggar, Iman M.; Bubb, Michael R.

In: Journal of the American Society for Mass Spectrometry, Vol. 18, No. 12, 12.2007, p. 2137-2145.

Research output: Contribution to journalArticle

Woodling, Kellie A. ; Eyler, John R. ; Tsybin, Yury O. ; Nilsson, Carol L. ; Marshall, Alan G. ; Edison, Arthur S. ; Al-Naggar, Iman M. ; Bubb, Michael R. / Identification of Single and Double Sites of Phosphorylation by ECD FT-ICR/MS in Peptides Related to the Phosphorylation Site Domain of the Myristoylated Alanine-Rich C Kinase Protein. In: Journal of the American Society for Mass Spectrometry. 2007 ; Vol. 18, No. 12. pp. 2137-2145.
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abstract = "A series of phosphorylated test peptides was studied by electron capture dissociation Fourier transform ion cyclotron resonance mass spectrometry (ECD FT-ICR MS). The extensive ECD-induced fragmentation made identification of phosphorylation sites for these peptides straightforward. The site(s) of initial phosphorylation of a synthetic peptide with a sequence identical to that of the phosphorylation site domain (PSD) of the myristoylated alanine-rich C kinase (MARCKS) protein was then determined. Despite success in analyzing fragmentation of the smaller test peptides, a unique site on the PSD for the first step of phosphorylation could not be identified because the phosphorylation reaction produced a heterogeneous mixture of products. Some molecules were phosphorylated on the serine closest to the N-terminus, and others on one of the two serines closest to the C-terminus of the peptide. Although no definitive evidence for phosphorylation on either of the remaining two serines in the PSD was found, modification there could not be ruled out by the ECD fragmentation data.",
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AU - Woodling, Kellie A.

AU - Eyler, John R.

AU - Tsybin, Yury O.

AU - Nilsson, Carol L.

AU - Marshall, Alan G.

AU - Edison, Arthur S.

AU - Al-Naggar, Iman M.

AU - Bubb, Michael R.

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