Winters C J, Reeves W B, Andreoli T E
Department of Internal Medicine, University of Arkansas College of Medicine, Little Rock 72205.
J Membr Biol. 1990 Dec;118(3):269-78. doi: 10.1007/BF01868611.
We evaluated the effects of varying aqueous Cl- concentrations, and of the arginyl- and lysyl-specific reagent phenylglyoxal (PGO), on the properties of Cl- channels fused from basolaterally enriched renal medullary vesicles into planar lipid bilayers. The major channel properties studied were the anion selectivity sequence, anionic requirements for channel activity, and the effects of varying Cl- concentrations and/or PGO on the relation between holding voltage (VH, mV) and open-time probability (Po). Reducing cis Cl- concentrations, in the range 50-320 mM, produced a linear reduction in fractional open time (Po) with a half-maximal reduction in Po at cis Cl- approximately 170 mM. Channel activity was sustained by equimolar replacement of cis Cl- with F-, but not with impermeant isethionate. For trans solutions, the relation between Cl- concentration and Po was negatively cooperative, with 50% reduction in po at 10 mM Cl-. Reducing cis Cl- had no effect on the gating charge (Z) for channel opening, but altered significantly the voltage-independent energy (delta G) for channel opening. Phenylglyoxal (PGO) reduced Z and altered delta G for Cl- channel activity when added to cis, but not trans solutions. Furthermore, in the presence of cis PGO, reducing the cis Cl- concentration had no effect on Z but altered delta G. Thus we propose that cis PGO and cis Cl- concentrations affect separate sites determining channel activity at the extracellular faces of these Cl- channels.
我们评估了不同的氯离子（Cl⁻）水溶液浓度以及精氨酰和赖氨酰特异性试剂苯乙二醛（PGO）对从肾髓质基底外侧富集囊泡融合到平面脂质双分子层中的氯离子通道特性的影响。所研究的主要通道特性包括阴离子选择性序列、通道活性的阴离子需求，以及不同Cl⁻浓度和/或PGO对保持电压（VH，mV）与开放时间概率（Po）之间关系的影响。在50 - 320 mM范围内降低顺式Cl⁻浓度，会使开放时间分数（Po）呈线性降低，在顺式Cl⁻约为170 mM时Po降低至最大值的一半。用氟离子（F⁻）等摩尔替代顺式Cl⁻可维持通道活性，但用不可渗透的羟乙基磺酸则不行。对于反式溶液，Cl⁻浓度与Po之间的关系呈负协同性，在Cl⁻浓度为10 mM时Po降低50%。降低顺式Cl⁻对通道开放的门控电荷（Z）没有影响，但显著改变了通道开放的电压无关能量（ΔG）。当添加到顺式溶液而非反式溶液中时，苯乙二醛（PGO）会降低Cl⁻通道活性的Z并改变ΔG。此外，在存在顺式PGO的情况下，降低顺式Cl⁻浓度对Z没有影响，但会改变ΔG。因此，我们提出顺式PGO和顺式Cl⁻浓度会影响这些Cl⁻通道细胞外表面决定通道活性的不同位点。
