Stabilization of Leukotriene A4 by Epithelial Fatty Acid-binding Protein in the Rat Basophilic Leukemia Cell

Jennifer S. Dickinson Zimmer, Dennis R. Voelker, David A. Bernlohr, Robert C. Murphy

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28 Scopus citations

Abstract

Leukotriene A4 (LTA4) is a chemically unstable triene epoxide product of 5-lipoxygenase metabolism of arachidonic acid. Despite this chemical reactivity and its synthesis at the perinuclear membrane, LTA4 is enzymatically converted into the cysteinyl leukotrienes and leukotriene B4. Furthermore, LTA4 participates in transcellular biosynthesis and is thus transferred between cells as an intact molecule. A cytosolic fatty acid-binding protein present in the rat basophilic leukemia cells was identified using mass spectrometry. This protein was determined to be the stabilizing factor present in the cell cytosol responsible for increasing the effective chemical half-life of LTA4. Rat epithelial fatty acid-binding protein (E-FABP) was isolated using partial protein purification and immunoprecipitation. In-gel digestion with trypsin followed by peptide fingerprint analysis using matrix-assisted laser desorption ionization mass spectrometry and sequencing the major tryptic peptide obtained from liquid chromatography/mass spectrometry/mass spectrometry analysis identified E-FABP in the active fraction. Semi-quantitative Western blot analysis indicated that E-FABP in the cytosolic fraction of RBL-1 cells was present at ∼1-3 pmol/106 cells. E-FABP (9 μM) was tested for its ability to stabilize LTA4, and at 37 °C E-FABP was able to increase the half-life of LTA4 from the previously reported half-life less than 3 s to a half-life of ∼7 min. These results present a novel function for the well studied fatty acid-binding protein as a participant in leukotriene biosynthesis that permits LTA4 to be available for further enzymatic processing in various cellular regions.

Original languageEnglish (US)
Pages (from-to)7420-7426
Number of pages7
JournalJournal of Biological Chemistry
Volume279
Issue number9
DOIs
StatePublished - Feb 27 2004

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