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Phosphate group-driven fragmentation of multiply charged phosphopeptide anions. Improved recognition of peptides phosphorylated at serine, threonine, or tyrosine by negative ion electrospray tandem mass spectrometry.
Anal Chem. 2006 Feb 15; 78(4):1249-56.AC

Abstract

The nanoelectrospray product ion spectra of multiply charged phosphopeptide anions reveal the occurrence of phosphate-specific high-mass fragment ions of the type [M - nH - 79](n-1)-. These so far unrecognized fragments, which are observed for phosphoserine-, phosphothreonine-, and phosphotyrosine-containing peptides, are the counterparts of the established inorganic phosphopeptide marker ion found at m/z 79 = [PO3]-. The high-mass marker ions are formed with high efficiency at moderate collision offset values and are particularly useful for sensitive recognition of pSer-, pThr-, and pTyr-peptides due to the low background level in MS/MS spectra at m/z values above those of the precursor ions. By virtue of this feature, the detection of the new phosphorylation-specific fragment ions appears to be more sensitive than the detection of the low-mass phosphate marker ion at m/z 79, where a higher interference by nonspecific background signals is generally observed. The number of phosphate groups within a phosphopeptide can also be estimated on the basis of the [M - nH - 79](n-1)- ions, since these exhibit an effective, sequential neutral loss of H3PO4 of the residing phosphate groups. A mechanistic explanation for the formation of the [M - nH - 79](n-1)- ions from multiply charged phosphopeptides is given. The high-mass marker ions are proposed to originate from phosphopeptide anions, which carry two negative charges located at the phosphate group. A new search tool denominated "variable m/z gain analysis", which utilizes these newly recognized high-mass fragments for spotting of phosphopeptides in a negative ion parent scan, is proposed. The findings strengthen the value of negative ion ESI-MS/MS for analysis of protein phosphorylation.

Authors+Show Affiliations

Central Spectroscopy and Central Peptide Synthesis Unit, German Cancer Research Center, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.No affiliation info availableNo affiliation info available

Pub Type(s)

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

Language

eng

PubMed ID

16478119

Citation

Edelson-Averbukh, Marina, et al. "Phosphate Group-driven Fragmentation of Multiply Charged Phosphopeptide Anions. Improved Recognition of Peptides Phosphorylated at Serine, Threonine, or Tyrosine By Negative Ion Electrospray Tandem Mass Spectrometry." Analytical Chemistry, vol. 78, no. 4, 2006, pp. 1249-56.
Edelson-Averbukh M, Pipkorn R, Lehmann WD. Phosphate group-driven fragmentation of multiply charged phosphopeptide anions. Improved recognition of peptides phosphorylated at serine, threonine, or tyrosine by negative ion electrospray tandem mass spectrometry. Anal Chem. 2006;78(4):1249-56.
Edelson-Averbukh, M., Pipkorn, R., & Lehmann, W. D. (2006). Phosphate group-driven fragmentation of multiply charged phosphopeptide anions. Improved recognition of peptides phosphorylated at serine, threonine, or tyrosine by negative ion electrospray tandem mass spectrometry. Analytical Chemistry, 78(4), 1249-56.
Edelson-Averbukh M, Pipkorn R, Lehmann WD. Phosphate Group-driven Fragmentation of Multiply Charged Phosphopeptide Anions. Improved Recognition of Peptides Phosphorylated at Serine, Threonine, or Tyrosine By Negative Ion Electrospray Tandem Mass Spectrometry. Anal Chem. 2006 Feb 15;78(4):1249-56. PubMed PMID: 16478119.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Phosphate group-driven fragmentation of multiply charged phosphopeptide anions. Improved recognition of peptides phosphorylated at serine, threonine, or tyrosine by negative ion electrospray tandem mass spectrometry. AU - Edelson-Averbukh,Marina, AU - Pipkorn,Rüdiger, AU - Lehmann,Wolf D, PY - 2006/2/16/pubmed PY - 2007/3/30/medline PY - 2006/2/16/entrez SP - 1249 EP - 56 JF - Analytical chemistry JO - Anal Chem VL - 78 IS - 4 N2 - The nanoelectrospray product ion spectra of multiply charged phosphopeptide anions reveal the occurrence of phosphate-specific high-mass fragment ions of the type [M - nH - 79](n-1)-. These so far unrecognized fragments, which are observed for phosphoserine-, phosphothreonine-, and phosphotyrosine-containing peptides, are the counterparts of the established inorganic phosphopeptide marker ion found at m/z 79 = [PO3]-. The high-mass marker ions are formed with high efficiency at moderate collision offset values and are particularly useful for sensitive recognition of pSer-, pThr-, and pTyr-peptides due to the low background level in MS/MS spectra at m/z values above those of the precursor ions. By virtue of this feature, the detection of the new phosphorylation-specific fragment ions appears to be more sensitive than the detection of the low-mass phosphate marker ion at m/z 79, where a higher interference by nonspecific background signals is generally observed. The number of phosphate groups within a phosphopeptide can also be estimated on the basis of the [M - nH - 79](n-1)- ions, since these exhibit an effective, sequential neutral loss of H3PO4 of the residing phosphate groups. A mechanistic explanation for the formation of the [M - nH - 79](n-1)- ions from multiply charged phosphopeptides is given. The high-mass marker ions are proposed to originate from phosphopeptide anions, which carry two negative charges located at the phosphate group. A new search tool denominated "variable m/z gain analysis", which utilizes these newly recognized high-mass fragments for spotting of phosphopeptides in a negative ion parent scan, is proposed. The findings strengthen the value of negative ion ESI-MS/MS for analysis of protein phosphorylation. SN - 0003-2700 UR - https://www.unboundmedicine.com/medline/citation/16478119/Phosphate_group_driven_fragmentation_of_multiply_charged_phosphopeptide_anions__Improved_recognition_of_peptides_phosphorylated_at_serine_threonine_or_tyrosine_by_negative_ion_electrospray_tandem_mass_spectrometry_ L2 - https://doi.org/10.1021/ac051649v DB - PRIME DP - Unbound Medicine ER -