Institute of Physiology, Pathophysiology and Biophysics, Department of Biomedical Sciences, University of Veterinary Medicine, Vienna, Austria.
Division of Organic Chemistry and Biochemistry, Rudjer Bošković Institute, Zagreb, Croatia.
Biophys J. 2018 May 8;114(9):2142-2151. doi: 10.1016/j.bpj.2018.04.011.
Long-chain free fatty acids (FFAs) play an important role in several physiological and pathological processes such as lipid fusion, adjustments of membrane permeability and fluidity, and the regulation of enzyme and protein activities. FFA-facilitated membrane proton transport (flip-flop) and FFA-dependent proton transport by membrane proteins (e.g., mitochondrial uncoupling proteins) are governed by the difference between FFA's intrinsic pK value and the pH in the immediate membrane vicinity. Thus far, a quantitative understanding of the process has been hampered, because the pK value shifts upon moving the FFA from the aqueous solution into the membrane. For the same FFA, pK values between 5 and 10.5 were reported. Here, we systematically evaluated the dependence of pK values on chain length and number of double bonds by measuring the ζ-potential of liposomes reconstituted with FFA at different pH values. The experimentally obtained intrinsic pK values (6.25, 6.93, and 7.28 for DOPC membranes) increased with FFA chain length (C16, C18, and C20), indicating that the hydrophobic energy of transfer into the bilayer is an important pK determinant. The observed pK decrease in DOPC with increasing number of FFA double bonds (7.28, 6.49, 6.16, and 6.13 for C20:0, C20:1, C20:2, and C20:4, respectively) is in line with a decrease in transfer energy. Molecular dynamic simulations revealed that the ionized carboxylic group of the FFAs occupied a fixed position in the bilayer independent of chain length, underlining the importance of Born energy. We conclude that pK is determined by the interplay between the energetic costs for 1) burying the charged moiety into the lipid bilayer and 2) transferring the hydrophobic protonated FFA into the bilayer.
长链游离脂肪酸 (FFA) 在多种生理和病理过程中发挥着重要作用，例如脂质融合、膜通透性和流动性的调节，以及酶和蛋白质活性的调节。FFA 促进的膜质子转运（翻转）和膜蛋白（如线粒体解偶联蛋白）依赖的 FFA 质子转运受 FFA 固有 pK 值与紧邻膜处 pH 值之间的差异控制。到目前为止，由于 FFA 从水溶液转移到膜中时 pK 值会发生变化，因此对该过程的定量理解一直受到阻碍。对于相同的 FFA，报道的 pK 值在 5 到 10.5 之间。在这里，我们通过测量不同 pH 值下用 FFA 重建的脂质体的 ζ-电势，系统地评估了 pK 值对链长和双键数的依赖性。实验获得的固有 pK 值（DOPC 膜为 6.25、6.93 和 7.28）随 FFA 链长（C16、C18 和 C20）增加而增加，表明转移到双层中的疏水性能量是重要的 pK 决定因素。在 DOPC 中观察到的随着 FFA 双键数增加而 pK 值降低（C20:0、C20:1、C20:2 和 C20:4 分别为 7.28、6.49、6.16 和 6.13）与转移能量降低一致。分子动力学模拟表明，FFA 的离子化羧酸基团在双层中占据固定位置，与链长无关，这强调了 Born 能量的重要性。我们得出结论，pK 值由以下两个因素之间的相互作用决定：1）将带电部分埋入脂质双层中的能量成本，以及 2）将疏水性质子化 FFA 转移到双层中的能量成本。
