Rostovtseva T K, Bezrukov S M
Laboratory of Physical and Structural Biology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland 20892, USA.
Biophys J. 1998 May;74(5):2365-73. doi: 10.1016/S0006-3495(98)77945-7.
The "molecular Coulter counter" concept has been used to study transport of ATP molecules through the nanometer-scale aqueous pore of the voltage-dependent mitochondrial ion channel, VDAC. We examine the ATP-induced current fluctuations and the change in average current through a single fully open channel reconstituted into a planar lipid bilayer. At high salt concentration (1 M NaCl), the addition of ATP reduces both solution conductivity and channel conductance, but the effect on the channel is several times stronger and shows saturation behavior even at 50 mM ATP concentration. These results and simple steric considerations indicate pronounced attraction of ATP molecules to VDAC's aqueous pore and permit us to evaluate the effect of a single ATP molecule on channel conductance. ATP addition also generates an excess noise in the ionic current through the channel. Analysis of this excess noise shows that its spectrum is flat in the accessible frequency interval up to several kilohertz. ATP exchange between the pore and the bulk is fast enough not to display any dispersion at these frequencies. By relating the low-frequency spectral density of the noise to the equilibrium diffusion of ATP molecules in the aqueous pore, we calculate a diffusion coefficient D = (1.6-3.3)10(-11) m2/s. This is one order of magnitude smaller than the ATP diffusion coefficient in the bulk, but it agrees with recent results on ATP flux measurements in multichannel membranes using the luciferin/luciferase method.
“分子库尔特计数器”概念已被用于研究ATP分子通过电压依赖性线粒体离子通道VDAC的纳米级水孔的运输。我们研究了ATP诱导的电流波动以及通过重构到平面脂质双分子层中的单个完全开放通道的平均电流变化。在高盐浓度（1M NaCl）下，添加ATP会降低溶液电导率和通道电导，但对通道的影响要强几倍，并且即使在50mM ATP浓度下也表现出饱和行为。这些结果和简单的空间位阻考虑表明ATP分子对VDAC水孔有明显的吸引力，并使我们能够评估单个ATP分子对通道电导的影响。添加ATP还会在通过通道的离子电流中产生过量噪声。对这种过量噪声的分析表明，其频谱在高达几千赫兹的可及频率区间内是平坦的。在这些频率下，孔与主体之间的ATP交换足够快，不会显示任何色散。通过将噪声的低频谱密度与ATP分子在水孔中的平衡扩散相关联，我们计算出扩散系数D = (1.6 - 3.3)×10⁻¹¹ m²/s。这比ATP在主体中的扩散系数小一个数量级，但与最近使用荧光素/荧光素酶方法在多通道膜中进行的ATP通量测量结果一致。
