The patch-clamp technique was used to examine the plasma membranes of sensitive yeast spheroplasts exposed to partially purified killer toxin preparations. Asolectin liposomes in which the toxin was incorporated were also examined. Excised inside-out patches from these preparations often revealed at 118 pS conductance appearing in pairs. The current through this conductance flickered rapidly among three states: dwelling mostly at the unit-open state, less frequently at the two-unit-open state, and more rarely at the closed state. Membrane voltages from -80 to 80 mV had little influence on the opening probability. The current reversed near the equilibrium potential of K+ in asymmetric KCl solutions and also reversed near O mV at symmetric NaCl vs. KCl solutions. The two levels of the conductance were likely due to the toxin protein, as treatment of spheroplasts or liposomes with extracellular protein preparations from isogenic yeasts deleted for the toxin gene gave no such conductance levels. These results show that in vivo the killer-toxin fraction can form a cation channel that seldom closes regardless of membrane voltage. We suggest that this channel causes the death of sensitive yeast cells.

译文

:使用膜片钳技术检查暴露于部分纯化的杀伤性毒素制剂的敏感酵母原生质球的质膜。还检查了其中掺入了毒素的Asolectin脂质体。从这些制剂中切出的由内而外的补丁通常以118 pS的电导率成对出现。通过该电导的电流在三个状态之间快速闪烁:主要在单元打开状态下居住,在两个单元打开状态下较少居住,而在闭合状态下很少居住。从-80到80 mV的膜电压对打开概率几乎没有影响。在非对称KCl溶液中,电流在K的平衡电位附近反转,并且在对称NaCl与KCl溶液中,在O mV附近也反转。电导的两个水平可能是由于毒素蛋白引起的,因为用针对毒素基因缺失的等基因酵母中的胞外蛋白制剂处理原生质球或脂质体并没有得到这样的电导水平。这些结果表明,在体内,无论膜电压如何,杀伤毒素部分都可以形成很少关闭的阳离子通道。我们建议该通道导致敏感酵母细胞死亡。

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