Peptide-Induced Lipid Flip-Flop in Asymmetric Liposomes Measured by Small Angle Neutron Scattering
Peptide-Induced Lipid Flip-Flop in Asymmetric Liposomes Measured by Small Angle Neutron Scattering
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DOI：
10.1021/acs.langmuir.9b01625
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发表时间：
2019-09-10
期刊：
影响因子：
3.9
通讯作者：
Marquardt, Drew
中科院分区：
文献类型：
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作者：
Nguyen, Michael H. L.;DiPasquale, Mitchell;Marquardt, Drew
尽管天然质膜中普遍存在脂质跨双分子层不对称，但大多数研究的仿生模型膜是对称的。最近的进展有助于克服在体外制备不对称脂质体的困难，允许检查更大的相关生物物理问题集。在这里，我们通过时间分辨小角中子散射（SANS）测量脂质触发器来研究不对称双分子层的稳定性。不对称的大单层小泡在37℃（半衰期，t(1/2) = 140 h）表现出缓慢的自发翻转，其内部双层小叶主要含有1-棕榈酰-2-油基-sn-甘油-3-磷酸胆碱（POPC），外部小叶主要含有1,2-二myristoyl-sn-甘油-3-磷酸胆碱（DMPC）。然而，肽的包含，即gramicidin、alamethicin、melittin或pHLIP（即低ph插入肽），加速了脂质翻转。其中三种多肽（即菲利普肽、alamethicin肽和蜂蜂素肽）分别被添加到预先形成的不对称囊泡中，我们在不到2小时的时间内观察到一个完全混乱的双分子层。另一方面，Gramicidin在不对称脂质体形成过程中被预先掺入，并显示出脂质不对称损失速率增加了8倍。这些结果表明，膜表面相关（例如，吸附/插入）事件是本研究中不对称模型膜中脂质混乱的主要驱动因素。我们讨论了膜肽结合、构象和插入对脂质不对称的影响。
Despite the prevalence of lipid transbilayer asymmetry in natural plasma membranes, most biomimetic model membranes studied are symmetric. Recent advances have helped to overcome the difficulties in preparing asymmetric liposomes in vitro, allowing for the examination of a larger set of relevant biophysical questions. Here, we investigate the stability of asymmetric bilayers by measuring lipid flip-flop with time-resolved small-angle neutron scattering (SANS). Asymmetric large unilamellar vesicles with inner bilayer leaflets containing predominantly 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and outer leaflets composed mainly of 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC) displayed slow spontaneous flip-flop at 37 degrees C (half-time, t(1/2) = 140 h). However, inclusion of peptides, namely, gramicidin, alamethicin, melittin, or pHLIP (i.e., pH-low insertion peptide), accelerated lipid flip-flop. For three of these peptides (i.e., pHLIP, alamethicin, and melittin), each of which was added externally to preformed asymmetric vesicles, we observed a completely scrambled bilayer in less than 2 h. Gramicidin, on the other hand, was preincorporated during the formation of the asymmetric liposomes and showed a time resolvable 8-fold increase in the rate of lipid asymmetry loss. These results point to a membrane surface-related (e.g., adsorption/insertion) event as the primary driver of lipid scrambling in the asymmetric model membranes of this study. We discuss the implications of membrane peptide binding, conformation, and insertion on lipid asymmetry.