Characterization of phosphatidylinositol, phosphatidylinositol-4-phosphate, and phosphatidylinositol-4,5-bisphosphate by electrospray ionization tandem mass spectrometry: A mechanistic study

Authors
Citation
Ff. Hsu et J. Turk, Characterization of phosphatidylinositol, phosphatidylinositol-4-phosphate, and phosphatidylinositol-4,5-bisphosphate by electrospray ionization tandem mass spectrometry: A mechanistic study, J AM SOC M, 11(11), 2000, pp. 986-999
Citations number
13
Categorie Soggetti
Spectroscopy /Instrumentation/Analytical Sciences
Journal title
JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY
ISSN journal
10440305 → ACNP
Volume
11
Issue
11
Year of publication
2000
Pages
986 - 999
Database
ISI
SICI code
1044-0305(200011)11:11<986:COPP>2.0.ZU;2-3
Abstract
Structural characterization of phosphatidylinositol (PI), phosphatidylinosi tol-4-phosphate CPI-LP), and phosphatidylinositol-4,5-bisphosphate (PI-4,5- P-2) by collisionally activated dissociation (CAD) tandem mass spectrometry with electrospray ionization is described. In negative ion mode, the major fragmentation pathways under low energy CAD for PI arise from neutral loss of free fatty acid substituents ([M - H - RxCO2H](-)) and neutral loss of the corresponding ketenes ([M - H - R-x'CH=C=O](-)), followed by consecutiv e loss of the inositol head group. The intensities of the ions arising from neutral loss of the sn-2 substutient as a free fatty acid ([M - H - R2CO2H ](-)) or as a ketene ([M - H - R-2'CH=C=O](-)) are greater than those of io ns reflecting corresponding losses of the sn-l substutient. This is consist ent with our recent finding that ions reflecting those losses arise from ch arge-driven processes that occur preferentially at the sn-2 position. These features permit assignment of the position of the fatty acid substituents on the glycerol backbone. Nucleophilic attack of the anionic phosphate onto the C-l or the C-2 of the glycerol to which the fatty acids attached expel s sn-l (R1CO2-) or sn-2 (R2CO2-) carboxylate anion, respectively. This path way is sterically more favorable at sn-2 than at sn-l. However, further dis sociations of [M - H - RxCO2H - inositol](-), [M - H - RxCO2H](-) and [M - H - R-x'CH=C=O](-) precursor ions also yield RxCO2- ions, whose abundance a re affected by the collision energy applied. Therefore, relative intensitie s of the RxCO2- ions in the spectrum do not reflect their positions on the glycerol backbone and determination of their regiospecificities based on th eir ion intensities is not reliable. The spectra also contain specific ions at m/z 315, 279, 259, 241, and 223, reflecting the Inositol head group. Th e last three ions are also observed in the tandem spectra of the [M - H](-) ions of phosphatidylinositol monophosphate (PI-P) and phosphatidylinositol bisphosphate (PI-P-2), in addition to the ions at mit 321 and 303, reflect ing the doubly phosphorylated inositol ions. The PI-P-2 also contains uniqu e ions at m/z 401 and 383 that reflect the triply phosphorylated inositol i ons. The [M - H](-) ions of PI-P and PI-P-2 undergo fragmentation pathways similar to that of PI upon CAD. However, the doubly charged ([M - 2H](2-)) molecular ions undergo fragmentation pathways that are typical of the [M - H] ions of glycerophosphoethanolamine, which are basic. These results sugge st that the further deprotonated gaseous [M - 2H](2-) ions of PI-P and PI-P -2 are basic precursors. (C) 2000 American Society for Mass Spectrometry.