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

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.