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Sequencing of peptides phosphorylated on serines and threonines by post-source decay in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
J Mass Spectrom. 1999 Nov; 34(11):1195-204.JM

Abstract

In the era of complete genome sequences, biochemical and medical research will focus more on the dynamic proteome of a cell. Regulation of proteins by post-translational modifications, which are not determined by the gene sequence, are already intensively studied. One example is phosphorylation of serines and threonines, probably the single most common cellular regulatory mechanism. In this paper we describe the sequencing of mono- and bisphosphorylated peptides, including identification of the phosphorylation sites, by post-source decay (PSD) in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. In addition to dephosphorylation of the parent ions, we studied the influence of the phosphate group on the fragmentation of peptides. Generally, peptides phosphorylated on serine and threonine residues displayed no difference in their fragmentation patterns. The intensities of the resulting fragment ion signals depend only on the peptide sequence and not on either the phosphorylated amino acid or its position in the peptide chain. Phosphorylation increased the bond cleavage C-terminal to the phosphorylation site more than 10-fold, resulting in abundant signals, which typically dominated the PSD spectra. The produced C-terminally phosphorylated b-type fragment ions showed characteristic dephosphorylated fragment ions b(n) -H(3)PO(4) (-98 Da) and b(n) -HPO(3) (-80 Da) of higher abundances than the phosphorylated fragment ion. As a second layer to identify the phosphorylation site, all internally phosphorylated fragment ions were accompanied by minor, but always detectable, signals of the dephosphorylated fragment ions. Interpretation of PSD spectra of phosphopeptides was not more complicated than for unphosphorylated peptides, despite the increased number of obtained fragment ion signals.

Authors+Show Affiliations

Biologisch-Medizinisches Forschungszentrum (BMFZ), Heinrich-Heine-Universität, Düsseldorf, Germany.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

10548813

Citation

Hoffmann, R, et al. "Sequencing of Peptides Phosphorylated On Serines and Threonines By Post-source Decay in Matrix-assisted Laser Desorption/ionization Time-of-flight Mass Spectrometry." Journal of Mass Spectrometry : JMS, vol. 34, no. 11, 1999, pp. 1195-204.
Hoffmann R, Metzger S, Spengler B, et al. Sequencing of peptides phosphorylated on serines and threonines by post-source decay in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. J Mass Spectrom. 1999;34(11):1195-204.
Hoffmann, R., Metzger, S., Spengler, B., & Otvos, L. (1999). Sequencing of peptides phosphorylated on serines and threonines by post-source decay in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. Journal of Mass Spectrometry : JMS, 34(11), 1195-204.
Hoffmann R, et al. Sequencing of Peptides Phosphorylated On Serines and Threonines By Post-source Decay in Matrix-assisted Laser Desorption/ionization Time-of-flight Mass Spectrometry. J Mass Spectrom. 1999;34(11):1195-204. PubMed PMID: 10548813.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Sequencing of peptides phosphorylated on serines and threonines by post-source decay in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AU - Hoffmann,R, AU - Metzger,S, AU - Spengler,B, AU - Otvos,L,Jr PY - 1999/11/5/pubmed PY - 2001/3/28/medline PY - 1999/11/5/entrez SP - 1195 EP - 204 JF - Journal of mass spectrometry : JMS JO - J Mass Spectrom VL - 34 IS - 11 N2 - In the era of complete genome sequences, biochemical and medical research will focus more on the dynamic proteome of a cell. Regulation of proteins by post-translational modifications, which are not determined by the gene sequence, are already intensively studied. One example is phosphorylation of serines and threonines, probably the single most common cellular regulatory mechanism. In this paper we describe the sequencing of mono- and bisphosphorylated peptides, including identification of the phosphorylation sites, by post-source decay (PSD) in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. In addition to dephosphorylation of the parent ions, we studied the influence of the phosphate group on the fragmentation of peptides. Generally, peptides phosphorylated on serine and threonine residues displayed no difference in their fragmentation patterns. The intensities of the resulting fragment ion signals depend only on the peptide sequence and not on either the phosphorylated amino acid or its position in the peptide chain. Phosphorylation increased the bond cleavage C-terminal to the phosphorylation site more than 10-fold, resulting in abundant signals, which typically dominated the PSD spectra. The produced C-terminally phosphorylated b-type fragment ions showed characteristic dephosphorylated fragment ions b(n) -H(3)PO(4) (-98 Da) and b(n) -HPO(3) (-80 Da) of higher abundances than the phosphorylated fragment ion. As a second layer to identify the phosphorylation site, all internally phosphorylated fragment ions were accompanied by minor, but always detectable, signals of the dephosphorylated fragment ions. Interpretation of PSD spectra of phosphopeptides was not more complicated than for unphosphorylated peptides, despite the increased number of obtained fragment ion signals. SN - 1076-5174 UR - https://www.unboundmedicine.com/medline/citation/10548813/Sequencing_of_peptides_phosphorylated_on_serines_and_threonines_by_post_source_decay_in_matrix_assisted_laser_desorption/ionization_time_of_flight_mass_spectrometry_ DB - PRIME DP - Unbound Medicine ER -