Worley J F, French R J, Pailthorpe B A, Krueger B K
Department of Physiology, University of Maryland School of Medicine, Baltimore 21201.
Biophys J. 1992 May;61(5):1353-63. doi: 10.1016/S0006-3495(92)81942-2.
We have studied the effects of membrane surface charge on Na+ ion permeation and Ca2+ block in single, batrachotoxin-activated Na channels from rat brain, incorporated into planar lipid bilayers. In phospholipid membranes with no net charge (phosphatidylethanolamine, PE), at low divalent cation concentrations (approximately 100 microM Mg2+), the single channel current-voltage relation was linear and the single channel conductance saturated with increasing [Na+] and ionic strength, reaching a maximum (gamma max) of 31.8 pS, with an apparent dissociation constant (K0.5) of 40.5 mM. The data could be approximated by a rectangular hyperbola. In negatively charged bilayers (70% phosphatidylserine, PS; 30% PE) slightly larger conductances were observed at each concentration, but the hyperbolic form of the conductance-concentration relation was retained (gamma max = 32.9 pS and K0.5 = 31.5 mM) without any preferential increase in conductance at lower ionic strengths. Symmetrical application of Ca2+ caused a voltage-dependent block of the single channel current, with the block being greater at negative potentials. For any given voltage and [Na+] this block was identical in neutral and negatively charged membranes. These observations suggest that both the conduction pathway and the site(s) of Ca2+ block of the rat brain Na channel protein are electrostatically isolated from the negatively charged headgroups on the membrane lipids.
我们研究了膜表面电荷对整合于平面脂质双分子层中的大鼠脑单条、经蟾毒素激活的钠通道中钠离子渗透和钙离子阻滞的影响。在无净电荷的磷脂膜（磷脂酰乙醇胺，PE）中，在低二价阳离子浓度（约100 microM镁离子）下，单通道电流-电压关系呈线性，单通道电导随[钠离子]和离子强度增加而饱和，达到最大值（γ max）31.8 pS，表观解离常数（K0.5）为40.5 mM。数据可用矩形双曲线近似。在带负电荷的双分子层（70%磷脂酰丝氨酸，PS；30% PE）中，在每个浓度下观察到的电导略大，但电导-浓度关系的双曲线形式得以保留（γ max = 32.9 pS，K0.5 = 31.5 mM），在较低离子强度下电导没有任何优先增加。对称施加钙离子导致单通道电流的电压依赖性阻滞，在负电位下阻滞更大。对于任何给定的电压和[钠离子]，这种阻滞在中性和带负电荷的膜中是相同的。这些观察结果表明，大鼠脑钠通道蛋白的传导途径和钙离子阻滞位点在静电上与膜脂质上带负电荷的头部基团隔离。
