Rostovtseva T K, Aguilella V M, Vodyanoy I, Bezrukov S M, Parsegian V A
Laboratory of Physical and Structural Biology, NICHD, National Institutes of Health, Bethesda, Maryland 20892-5626 USA.
Biophys J. 1998 Oct;75(4):1783-92. doi: 10.1016/S0006-3495(98)77620-9.
We manipulate lipid bilayer surface charge and gauge its influence on gramicidin A channel conductance by two strategies: titration of the lipid charge through bulk solution pH and dilution of a charged lipid by neutral. Using diphytanoyl phosphatidylserine (PS) bilayers with CsCl aqueous solutions, we show that the effects of lipid charge titration on channel conductance are masked 1) by conductance saturation with Cs+ ions in the neutral pH range and 2) by increased proton concentration when the bathing solution pH is less than 3. A smeared charge model permits us to separate different contributions to the channel conductance and to introduce a new method for "bilayer pKa" determination. We use the Gouy-Chapman expression for the charged surface potential to obtain equilibria of protons and cations with lipid charges. To calculate cation concentration at the channel mouth, we compare different models for the ion distribution, exact and linearized forms of the planar Poisson-Boltzmann equation, as well as the construction of a "Gibbs dividing surface" between salt bath and charged membrane. All approximations yield the intrinsic pKain of PS lipid in 0.1 M CsCl to be in the range 2.5-3.0. By diluting PS surface charge at a fixed pH with admixed neutral diphytanoyl phosphatidylcholine (PC), we obtain a conductance decrease in magnitude greater than expected from the electrostatic model. This observation is in accord with the different conductance saturation values for PS and PC lipids reported earlier (, Biochim. Biophys. Acta. 552:369-378) and verified in the present work for solvent-free membranes. In addition to electrostatic effects of surface charge, gramicidin A channel conductance is also influenced by lipid-dependent structural factors.
我们通过两种策略来调控脂质双分子层表面电荷并评估其对短杆菌肽A通道电导的影响：通过改变本体溶液pH值来滴定脂质电荷，以及用中性脂质稀释带电荷脂质。使用含有CsCl水溶液的二植烷酰磷脂酰丝氨酸（PS）双分子层，我们发现脂质电荷滴定对通道电导的影响被掩盖了：一是在中性pH范围内Cs⁺离子使电导饱和，二是当浴液pH小于3时质子浓度增加。一个涂抹电荷模型使我们能够区分对通道电导的不同贡献，并引入一种测定“双分子层pKa”的新方法。我们使用带电表面电位的古依 - 查普曼表达式来获得质子和阳离子与脂质电荷的平衡。为了计算通道口处的阳离子浓度，我们比较了离子分布的不同模型、平面泊松 - 玻尔兹曼方程的精确形式和线性化形式，以及在盐浴和带电膜之间构建“吉布斯分界面”。所有近似结果都表明，在0.1 M CsCl中PS脂质的固有pKain在2.5 - 3.0范围内。通过在固定pH下用混合的中性二植烷酰磷脂酰胆碱（PC）稀释PS表面电荷，我们观察到电导降低的幅度大于静电模型的预期。这一观察结果与之前报道的PS和PC脂质不同的电导饱和值一致（Biochim. Biophys. Acta. 552:369 - 378），并且在本工作中对无溶剂膜进行了验证。除了表面电荷的静电效应外，短杆菌肽A通道电导还受脂质依赖性结构因素的影响。
