Electrospray ionization multiple stage quadrupole ion-trap and tandem quadrupole mass spectrometric studies on phosphatidylglycerol from arabidopsis leaves

  • Fong-Fu Hsu
  • John Turk
  • Todd D. Williams
  • Ruth Welti


Phosphatidylglycerol (PG) is the major phospholipid of plant chloroplasts. PG from Arabidopsis thaliana has an unusual fatty acyl chain, 3-trans-hexadecenoyl (Δ316:1) in the sn-2 position of the major 18:3/Δ316:1-PG species, as well as in 18:2/Δ316:1-PG and 16:0/Δ316:1-PG. Upon low-energy collisionally activated dissociation (CAD) in a tandem quadrupole or in an ion-trap mass spectrometer, the [M - H] ions of the PG molecules containing Δ316:1 give product-ion spectra that are readily distinguishable from those arising from PGs without the Δ316:1 species. The Δ316:1-fatty acyl-containing PGs are characterized by MS2 product-ion mass spectra that contain predominant [M - H - 236] ions arising from loss of the Δ316:1-fatty acyl substituent as a ketene. This is attributable to the fact that the α-hydrogen of the Δ316:1-fatty acid substituent involved in the ketene loss is an allylic hydrogen, which is very labile. This leads to preferential neutral loss of 236 and drastic decline in the neutral loss of 254 (i.e., loss as a fatty acid), the unique features that signify the presence of Δ316:1-fatty acyl containing PGs. The neutral loss scan of 236, thus, provides a sensitive tandem quadrupole mass spectrometric means to identify Δ316:1-containing PG species in lipid mixtures. This low-energy tandem mass spectrometric approach also permits the structures of the Arabidopsis PGs that consist of two isomeric structures to be unveiled.

Supplementary material

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  1. 1.
    Sakurai, I.; Hagio, M.; Gombos, Z.; Tyystjarvi, T.; Paakkarinen, V.; Aro, E. M.; Wada, H. Requirement of Phosphatidylglycerol for Maintenance of Photosynthetic Machinery. Plant Physiol. 2003, 133, 1376–1384.CrossRefGoogle Scholar
  2. 2.
    Sato, N. Roles of the Acidic Lipids Sulfoquinovosyl Diacylglycerol and Phosphatidylglycerol in Photosynthesis: Their Specificity and Evolution. J. Plant Res. 2004, 117, 495–505.CrossRefGoogle Scholar
  3. 3.
    Frentzen, M. Phosphatidylglycerol and Sulfoquinovosyldiacylglycerol: Anionic Membrane Lipids and Phosphate Regulation. Curr. Opin. Plant Biol. 2004, 7, 270–276.CrossRefGoogle Scholar
  4. 4.
    Joyard, J.; Maréchal, E.; Miège, C.; Block, M. A.; Dorne, A.; Douce, R. Structure, distribution, and biosynthesis of glycerolipids from higher plant chloroplasts. In Lipids in Photosynthesis: Structure, Function, and Genetics; Siegenthaler, P. A.; Murata, N., Eds.; Kluwer Academic Publishers: Dordrecht, The Netherlands, 1988; pp 21–52.Google Scholar
  5. 5.
    Hagio, M.; Gombos, Z.; Várkonyi, Z.; Masamoto, K.; Sato, N.; Tsuzuki, M.; Wada, H. Direct Evidence for Requirement of Phosphatidylglycerol in Photosystem II of Photosynthesis. Plant Physiol. 2000, 124, 795–804.CrossRefGoogle Scholar
  6. 6.
    Pineau, B.; Girard-Bascou, J.; Eberhard, S.; Choquet, Y.; Tremolieres, A.; Gerard-Hirne, C.; Bennardo-Connan, A.; Decottignies, P.; Gillet, S.; Wollman, F. A. A Single Mutation That Causes Phosphatidylglycerol Deficiency Impairs Synthesis of Photosystem II Cores in Chlamydomonas reinhardtii. Eur. J. Biochem. 2004, 271, 329–338.CrossRefGoogle Scholar
  7. 7.
    Haverkate, F.; de Gier, J.; van Deenen, L. L. The Occurrence of δ3-Trans-Hexadecenoic Acid in Phosphatidyl Glycerol from Spinach Leaves. Experientia 1964, 20, 511–512.CrossRefGoogle Scholar
  8. 8.
    Dubacq, J. P.; Tremolires, A. Occurrence and Function of Phosphatidylglycerol Containing Δ3-Trans-Hexadecenoic Acid in Photosynthetic Lamellae. Physiol. Veg. 1983, 21, 293–312.Google Scholar
  9. 9.
    Selstam, E. Development of Thylakoid membranes with respect to lipids. In Advances in Photosynthesis, Vol. VI; Lipids in Photosynthesis: Structure, Function, and Genetics; Siegenthaler, P.-A.; Murata, N., Eds.; Kluwer Academic Publishers: Dordrecht, The Netherlands, 1998; 209–224.Google Scholar
  10. 10.
    Browse, J.; Warwick, N.; Somerville, C. R.; Slack, C. R. Fluxes Through the Prokaryotic and Eukaryotic Pathways of Lipid Synthesis in the “16:3” Plant Arabidopsis thaliana. Biochem. J. 1986, 235, 25–31.Google Scholar
  11. 11.
    Roughan, P. G. Phosphatidyl Choline: Donor of 18-Carbon Unsaturated Fatty Acids for Glycerolipid Biosynthesis. Lipids 1975, 10, 609–614.CrossRefGoogle Scholar
  12. 12.
    Mackender, R. O.; Leech, R. M. The Galactolipid, Phospholipid, and Fatty Acid Composition of the Chloroplast Envelope Membranes of Vicia faba L. Plant Physiol. 1974, 53, 496–502.CrossRefGoogle Scholar
  13. 13.
    Ohnishi, M.; Thompson, G. A., Jr. Biosynthesis of the Unique Trans-δ3-Hexadecenoic Acid Component of Chloroplast Phosphatidylglycerol: Evidence Concerning Its Site and Mechanism of Formation. Arch. Biochem. Biophys. 1991, 288, 591–599.CrossRefGoogle Scholar
  14. 14.
    Beisson, F.; Koo, A. J. K.; Ruuska, S.; Schwender, J.; Pollard, M.; Thelen, J.; Paddock, T.; Salas, J.; Savage, L.; Milcamps, A.; Mhaske, V. B.; Cho, Y.; Ohlrogge, J. B. Arabidopsis thaliana Genes Involved in Acyl Lipid Metabolism: A 2003 Census of the Candidates, a Study of the Distribution of Expressed Sequence Tags in Organs, and a Web-Based Database. Plant Physiol 2003, 132, 681–697.CrossRefGoogle Scholar
  15. 15.
    Dubertret, G.; Mirshahi, A.; Mirshahi, M.; Gerard-Hirne, C.; Tremolieres, A. Evidence from in Vivo Manipulations of Lipid Composition in Mutants that the δ3-Trans-Hexadecenoic Acid-Containing Phosphatidylglycerol is Involved in the Biogenesis of the Light-Harvesting Chlorophyll A/B-Protein Complex of Chlamydomonas reinhardtii. Eur. J. Biochem. 1994, 226, 473–482.CrossRefGoogle Scholar
  16. 16.
    Krupa, Z. The Action of Lipases on Chloroplast Membranes: III. The Effect of Lipid Hydrolysis on Chlorophyll-Protein Complexes in Thylakoid Membranes. Photosynth. Res. 1984, 5, 177–184.CrossRefGoogle Scholar
  17. 17.
    Chapman, D. J.; De-Felice, J.; Barber, J. Characteristics of Chloroplast Thylakoid Lipid Composition Associated with Resistance to Triazine Herbicides. Planta 1985, 166, 280–285.CrossRefGoogle Scholar
  18. 18.
    Xu, Y.; Siegenthaler, P.-A. Low Temperature Treatments Induce an Increase in the Relative Content of Both Linolenic and Δ3-Trans-Hexadecenoic Acids in Thylakoid Membrane Phosphatidylglycerol of Squash Cotyledons. Plant Cell Physiol. 1997, 38, 611–618.CrossRefGoogle Scholar
  19. 19.
    Lamberto, M.; Ackman, R. G. Confirmation by Gas Chromatography/Mass Spectrometry of Two Unusual Trans-3-Monoethylenic Fatty Acids from the Nova Scotian Seaweeds Palmaria palmata and Chondrus crispus. Lipids 1994, 29, 441–444.CrossRefGoogle Scholar
  20. 20.
    Lamberto, M.; Ackman, R. G. Positional Isomerization of Trans-3-Hexadecenoic Acid Employing 2-Amino-2-Methyl-Propanol as a Derivatizing Agent for Ethylenic Bond Location by Gas Chromatography/Mass Spectrometry. Anal. Biochem. 1995, 230, 224–228.CrossRefGoogle Scholar
  21. 21.
    Welti, R.; Wang, X.; Williams, T. D. Electrospray Ionization Tandem Mass Spectrometry Scan Modes for Plant Chloroplast Lipids. Anal. Biochem. 2003, 314, 149–152.CrossRefGoogle Scholar
  22. 22.
    Hsu, F. F.; Turk, J. Studies on Phosphatidylglycerol with Triple Quadrupole Tandem Mass Spectrometry with Electrospray Ionization: Fragmentation Processes and Structural Characterization. J. Am. Soc. Mass Spectrom. 2001, 12, 1036–1043.CrossRefGoogle Scholar
  23. 23.
    Xu, Y.; Siegenthaler, P. A. Phosphatidylglycerol Molecular Species of Photosynthetic Membranes Analyzed by High-Performance Liquid Chromatography: Theoretical Considerations. Lipids 1996, 31, 223–229.CrossRefGoogle Scholar
  24. 24.
    Christie, W. W. Lipid Analysis 2nd ed. Pergamon Press: Oxford, United Kingdom, 1982; pp 109–110.Google Scholar
  25. 25.
    Hsu, F. F.; Turk, J. Electrospray Ionization with Low-Energy Collisionally Activated Dissociation Tandem Mass Spectrometry of Complex Lipids: Structural Characterization and Mechanisms of Fragmentation. In Modern Methods for Lipid Analysis by Liquid Chromatography/Mass Spectrometry; Byrdwell, W. C., Ed.; AOCS Press: Champaign, IL. 2005; pp 61–178.Google Scholar
  26. 26.
    Hsu, F. F.; Turk, J. Charge-Driven Fragmentation Processes in Diacyl Glycerophosphatidic Acids upon Low-Energy Collisional Activation: A Mechanistic Proposal. J. Am. Soc. Mass Spectrom. 2000, 11, 797–803.CrossRefGoogle Scholar
  27. 27.
    Hong, S.; Lu, Y.; Yanag, R.; Gotlinger, K. H.; Petasis, N. P.; Serhan, C. N. Resolvin D1, Protectin D1, and Related Docosahexaenoic Acid-Derived Products: Analysis Via Electrospray/Low Energy Tandem Mass Spectrometry Based on Spectra and Fragmentation Mechanisms. J. Am. Soc. Mass Spectrom. 2006, 18, 128–144.CrossRefGoogle Scholar
  28. 28.
    Moe, M. K.; Anderssen, T.; Strøm, M. B.; Jensen, E. Total Structure Characterization of Unsaturated Acidic Phospholipids Provided by Vicinal Dihydroxylation of Fatty Acid Double Bonds and Negative Electrospray Ionization Mass Spectrometry. J. Am. Soc. Mass Spectrom. 2005, 16, 46–59.CrossRefGoogle Scholar
  29. 29.
    Thomas, M. C.; Mitchell, T. W.; Blanksby, S. J. Ozonolysis of Phospholipid Double Bonds during Electrospray Ionization: A New Tool for Structure Determination. J. Am. Chem. Soc. 2006, 128, 58–59.CrossRefGoogle Scholar

Copyright information

© American Society for Mass Spectrometry 2007

Authors and Affiliations

  • Fong-Fu Hsu
    • 1
  • John Turk
    • 1
  • Todd D. Williams
    • 2
  • Ruth Welti
    • 3
  1. 1.Mass Spectrometry Resource, Division of Endocrinology, Diabetes, Metabolism, and Lipid research, Department of Internal MedicineWashington University School of MedicineSt. LouisUSA
  2. 2.University of Kansas Mass Spectrometry LaboratoryUniversity of KansasLawrenceUSA
  3. 3.Kansas Lipidomics Research Center, Division of BiologyKansas State UniversityManhattanUSA
  4. 4.Washington University School of MedicineSt. LouisUSA

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