Giangiacomo K M, Garcia M L, McManus O B
Department of Membrane Biochemistry and Biophysics, Merck Institute for Therapeutic Research, Rahway, New Jersey 07065.
Biochemistry. 1992 Jul 28;31(29):6719-27. doi: 10.1021/bi00144a011.
The interaction of iberiotoxin (IbTX) with the large-conductance calcium-activated potassium (maxi-K) channel was examined by measuring single-channel currents from maxi-K channels incorporated into planar lipid bilayers. Addition of nanomolar concentrations of IbTX to the external side of the channel produced long nonconducting silent periods, which were interrupted by periods of normal channel activity. The distributions of durations of blocked and unblocked periods were both described by single exponentials. The mean duration of the unblocked periods decreased in proportion with the external concentration of IbTX, while the mean duration of the blocked periods was not affected. These results suggest that IbTX blocks the maxi-K channel through a simple bimolecular binding reaction where the silent periods represent times when a single toxin molecule is bound to the channel. In symmetric solutions of 150 mM KCl, with a membrane potential of 40 mV, the mean duration of the blocked periods produced by IbTX was 840 s, and the association rate was 1.3 x 10(6) M-1 s-1, yielding an equilibrium dissociation constant of about 1 nM. Raising the internal potassium concentration increased the dissociation rate constant of IbTX in a manner which was well described by a saturable binding function for potassium. External tetraethylammonium ion increased the average duration of the unblocked periods without affecting the blocked periods, suggesting that tetraethylammonium and IbTX compete for the same site near the conductance pathway of the channel. Increasing the external concentration of monovalent cations from 25 to 300 mM with either potassium or sodium decreased the rate of binding of IbTX to the channel by approximately 24-fold, with little effect on the rate of toxin dissociation.(ABSTRACT TRUNCATED AT 250 WORDS)
通过测量整合到平面脂质双分子层中的大电导钙激活钾通道（maxi-K通道）的单通道电流，研究了埃博毒素（IbTX）与该通道的相互作用。向通道外侧添加纳摩尔浓度的IbTX会产生长时间的非传导性沉默期，这些沉默期会被正常通道活动期打断。阻断期和未阻断期的持续时间分布均可用单指数函数描述。未阻断期的平均持续时间随IbTX外部浓度成比例降低，而阻断期的平均持续时间不受影响。这些结果表明，IbTX通过简单的双分子结合反应阻断maxi-K通道，其中沉默期代表单个毒素分子与通道结合的时间。在150 mM KCl的对称溶液中，膜电位为40 mV时，IbTX产生的阻断期平均持续时间为840秒，结合速率为1.3×10⁶ M⁻¹ s⁻¹，平衡解离常数约为1 nM。提高内部钾浓度会以钾的饱和结合函数很好描述的方式增加IbTX的解离速率常数。外部四乙铵离子增加了未阻断期的平均持续时间，而不影响阻断期，这表明四乙铵和IbTX竞争通道传导途径附近的同一位点。用钾或钠将单价阳离子的外部浓度从25 mM增加到300 mM，会使IbTX与通道的结合速率降低约24倍，对毒素解离速率影响很小。（摘要截短于250字）
