Li Libo, Vorobyov Igor, Allen Toby W
Department of Chemistry, University of California, One Shields Avenue, Davis, California 95616, USA.
J Phys Chem B. 2008 Aug 14;112(32):9574-87. doi: 10.1021/jp7114912. Epub 2008 Jul 18.
The issue of ionizable protein side chains interacting with lipid membranes has been the focus of much attention since the proposal of the paddle model of voltage-gated ion channels, which suggested multiple arginine (Arg) side chains may move through the hydrocarbon core of a lipid membrane. Recent cell biology experiments have also been interpreted to suggest that these side chains would face only small free energy penalties to cross membranes, challenging a long-standing view in membrane biophysics. Here, we employ side chain analog and transmembrane helix models to determine the free energy of an Arg side chain, as a function of protonation state, across a membrane. We observe high free energy barriers for both the charged and neutral states that would prohibit lipid-exposed movement. The mechanisms for charged and neutral Arg transport are, however, very different, with the neutral state experiencing simple dehydration, whereas the charged state experiences a complex mechanism involving connections to the bilayer interfaces that deform the local membrane structure. We employ special methods to ensure sampling of these interfacial connections and decompose the free energy to shed light on the mechanisms. These deformations are found to preferentially stabilize the protonated form, such that the Arg side chain remains almost exclusively charged inside the membrane, with a pKa shift of <or=4.5 units. In contrast, the analog models are found to exaggerate the variations in energetics across the membrane and have larger pKa shifts. These results have implications for models of voltage gated ion channels, suggesting that although Arg side chains are ideally suited for carrying charge, the thermodynamics dictate that they must remain sequestered from the lipid bilayer environment.
自从电压门控离子通道的桨状模型提出以来，可电离蛋白质侧链与脂质膜相互作用的问题一直备受关注，该模型表明多个精氨酸（Arg）侧链可能穿过脂质膜的烃核。最近的细胞生物学实验也被解释为表明这些侧链穿过膜时仅面临较小的自由能惩罚，这对膜生物物理学中的一个长期观点提出了挑战。在这里，我们采用侧链类似物和跨膜螺旋模型来确定Arg侧链在膜上的自由能，作为质子化状态的函数。我们观察到带电和中性状态都存在高自由能障碍，这将阻止脂质暴露的移动。然而，带电和中性Arg转运的机制非常不同，中性状态经历简单的脱水，而带电状态经历涉及与双层界面连接的复杂机制，这会使局部膜结构变形。我们采用特殊方法来确保对这些界面连接进行采样，并分解自由能以阐明机制。发现这些变形优先稳定质子化形式，使得Arg侧链在膜内几乎完全带电荷，pKa偏移≤4.5个单位。相比之下，发现类似物模型夸大了跨膜能量学的变化并且具有更大的pKa偏移。这些结果对电压门控离子通道模型有影响，表明尽管Arg侧链非常适合携带电荷，但热力学表明它们必须与脂质双层环境隔离。
