SURFACE LYSINE RESIDUES MODULATE THE COLLISIONAL TRANSFER OF FATTY-ACID FROM ADIPOCYTE FATTY-ACID-BINDING PROTEIN TO MEMBRANES

The transfer of-unesterified fatty acids (FA) from adipocyte fatty aci d binding protein (A-FABP) to phospholipid membranes is proposed to oc cur via a collisional mechanism involving transient ionic and hydropho bic interactions [Wootan & Storch (1994) J. Biol. Chem. 269, 10517-105 23]. In particular, it was suggested that membrane acidic phospholipid s might specifically interact with basic residues on the surface of A- FABP. Here we addressed whether lysine residues on the surface of the protein are involved in this collisional transfer mechanism. Recombina nt A-FABP was acetylated to neutralize all positively charged surface lysine residues. Protein fluorescence, CD spectra, and chemical denatu rant data indicate that acetylation did not substantially alter the co nformational integrity of the protein, and nearly identical affinities were obtained for binding of the fluorescently labeled FA [12-(9-anth royloxy)oleate] to native and acetylated protein. Transfer of 2-(9-ant hroyloxy)palmitate (2AP) from acetylated A-FABP to small unilamellar v esicles (SW) was 35-fold slower than from native protein. In addition, whereas the 2AP transfer rate from native A-FABP was directly depende nt on SUV concentration, 2AP transfer from acetylated protein was inde pendent of the concentration of acceptor membranes. Factors which alte r aqueous-phase solubility of FA, such as ionic strength and acyl chai n length and saturation, affected the AOFA transfer rate from acetylat ed but not native A-FABP. Finally, an increase in the negative charge density of the acceptor SUV resulted in a marked increase in the rate of transfer from native A-FABP but did not increase the rate from acet ylated A-FABP. Collectively, these studies indicate that positively ch arged lysine residues on A-FABP are important for effective collisiona l transfer of FA between A-FABP and phospholipid bilayers. In the abse nce of rapid collisional transfer, FA movement to membranes occurs by a slower, aqueous diffusion-mediated process. Thus ionic interactions between A-FABP and membranes may play a key role in intracellular FA t rafficking.